Thursday, 31 January 2013

Evidence for a core gut microbiota in zebrafish




“Evidence for a core gut microbiota in the zebrafish”

In this paper by Roeselelers et al zebrafish were used in a study to examine in detail their host gut microbiota, this was achieved mainly by using 16srRNA sequencing. Comparisons were made between specimens from varied natural habitats and those raised domestically in laboratory conditions in order to acetate whether the communities are a selective decision or whether it is based on environmental or locality change.  An initial look at the gut microbiota of the zebrafish with in laboratory conditions, through DNA sequencing, shows that it is predominantly bacteria from the phylum proteobacteria, specifically  Firmicutes and Fusobacteria seemingly present during larva and adult stages.

Standard microbiological testing techniques were used in the methodology of this paper, they began with ensuring the laboratory conditions are standardised in order to maintain conformity across the board, vital for accurate results. Once samples were collected from the  Shutunga River in Mathabhanga subdivision of Cooch Behar district in the Indian state of West Bengal, Roeselers et al first began with DNA extraction using a classic blood/tissue DNA extraction followed by T-RFLP in order to analyse 16srRNA gene amplification, this in turn led to  sequencing of bacteria followed by standard phylogenic analysis to identify individual strains.

The results were quite interesting, firstly looking at the comparisons of the gut microbiota of domesticated (5 labs, multiple generations) and recently caught fish (using methods previously discussed) findings showed a significant similarity between communities. T-RFLP analysis also backed these results up, taxon based study was observed, the Shannon-weaver diversity index again showed significant similarities between the  domesticated and recently caught samples this was based on  OTU sequences defined by 97% pairwise sequence identity.

As well as the above Roestelers et al wanted to make phylogenetic comparisons, in order to achieve this they used the Uniifrac tree technique and supplemented 16srRNA sequences from the zebrafish against that of mammalian (mouse and human) samples as well as 7 other fish species in order to clear differences. Distinct clusters were seen, as expected the human and mouse libraries were clustering quite distinctively, denoting the obvious difference between mammalian species and fish species gut microbiota.

When analysing the fish libraries a distinct cluster was also seen with all the zebra fish samples and the yellow cat fish alike, further follow up studies subjecting 16srRNA gene sequences to PCoA, similarly to previous tests,  it showed a high similarity between recently caught zebrafish and the yellow cat fish. Interestingly two bacterial classes were found consistently in the gut of the zebrafish (both domesticated and terrestrial) these two classes were ƴ-proteobacteria and Fusobacteria, this could suggest that these classes have become  adapted for life in the fish intestine and utilising this as a niche environment.

One last study was looking at the change in community structure from different aquaculture laboratories, results demonstrated that the fish with in the same labs displayed similar community microbial communities but differed from that of other laboratories, this could be that they are displaying historical connections between bacterial communities and individual locations. The difference can be seen when looking at the 3 lab samples 525 OTU’s were discovered but only 21 were found in all 3, not displaying a high enough quantity to be significant.

I think that this is an interesting paper, perhaps a little wordy in the methods section, as most people who study microbiology are familiar with these techniques. As far as the study is concerned I found it a very interesting area, it could be particularly beneficial for the aquaculture industry and a further increase study could be expanded to other farmed fish such as salmon or tuna to help in the eradication of harmful bacteria released in fish fecal matter. It would also help in ornamental aquaculture industries as many fish are of muti-generations and these bacteria may be transferred during cross breeding from various other fish farms.
There is much more information in the paper, what I have talked about is only about two thirds of its depth, those interested in aquaculture may find it interesting so have a read I will put the paper below. Hope it makes sense and thanks for reading.   

 Roeselers, Guus, et al. "Evidence for a core gut microbiota in the zebrafish." The ISME journal 5.10 (2011): 1595-1608.

Wednesday, 30 January 2013

Shedding light on the function of marine sponge microbiota


Sponges are the most ancient metazoan animal phylum (approximately 600 million years old) and are found in tropical and subtropical oceans, the polar regions, the deep sea and in freshwater lakes and streams. Marine demosponges harbour complex microbial communities at densities exceeding the surrounding seawater by several orders of magnitude and sometimes these microorganisms make up 40-60% of the animals biomass. While the majority of the microorganisms remain unculturable, Grozdanov & Hentschel (2007) review several of the discoveries made using environmental genomic techniques shedding light on the function of sponge-associated microbiota.

The archaeon Cenarchaeum symbiosum (C. archaeum lineage) has now been found in more than 19 shallow water sponge species, and in an arctic deep water sponge, making up 70-90% of the microbial biomass. Interestingly, it is thought that the C. archaeum lineage consistently affiliated with the sponges is not thermophillic; sequence comparison of several clones carrying the ribosomal operon and subsequent biochemical characterization of the enzyme revealed enzyme instability above 40oC. Also, the presence of molecular machinery encoding a modified version of the 3-hydroxypropionate cycle for CO2 assimilation and the identification of genes using reduced nitrogen compounds and ammonia for energy production might be evidence for a symbiotic relationship between the sponge host and the archaeon, in which removal of sponge waste products such as ammonia would provide energy for the symbiont while CO2 fixation by the archaea would supply organic carbon to the sponge. A complete genome of C. symbiosum was generated by systematical selection of overlapping clones from environmental fosmid libraries and this has shown that many predicted genes appear to be specific and Grozdanov & Hentschel (2007) suggest that these may perform symbiosis-related functions such as mediation of cell-to-cell contact, degradation of extracellular matrix proteins or evasion of the host defences.

Several of the studies reviewed give evidence for a bacterial origin of onnamides, a group of polyketide compounds that exhibit powerful anti-tumour activities. The onnamides gene cluster follows the co-linearity rule in that the domain architecture of the involved genes mirrors the chemical structure of the onnamides nearly perfectly. Research on the diversity of PKSI genes led to the discovery of unusually small monomodular and widely distributed PKSI operons in sponge microbiota that are similar to methyl-branched fatty-acid-encoding PKS systems from Mycobacteria. The biological function of these in sponges is unclear but Grozdanov & Hentschel thought that these methylated fatty acids may be important for the establishment and maintenance of symbiosis by protecting the bacterial cells from phagocytosis. Several other compounds that are known to be secondary metabolites found in sponges as well as biotechnologically relevant compounds have been found to be produced by the sponge microbiota also.

This was an interesting paper highlighting some of the main findings surrounding marine sponges and the potential applications for some of the compounds produced by their microbial communities. These are of interest both medically and for biotechnology. Much of the information given for each point was from several separate studies and references for each can be found within the Grozdanov & Hentschel (2007) paper.

 

Grozdanov, L. & Hentschel, U. (2007) An environmental genomics perspective on the diversity and function of marine sponge-associated microbiota. Ecology and industrial microbiology. 10, 215-220

Tuesday, 29 January 2013

Algal biotechnology: An answer for the biofouling?


Quorum Sensing Inhibition by Asparagopsis taxiformis, a Marine Macro Alga: Separation of the Compound that Interrupts Bacterial Communication

 
Biofilms are communities of microbes sheathed in extracellular polymeric substances (see Munn’s book for more). Biofilm formation is controlled by quorum sensing (QS), as we know QS is density-dependent gene regulation through signals produced by bacteria. Many pathogenic bacteria in biofilms use QS as a regulatory mechanism for their production of virulence factors. Therefore, QS can be used as a drug target instead of antibiotics which has lead to emergence of drug resistance. It is known that quorum sensing inhibitors increase the susceptibility of bacterial biofilms to antibiotics; as a consequence this has led to the scientific community concentrating on QS inhibitors. We all know of some of the implications of biofilms, for instance harmful contamination of bacteria on important farmed animals and fouling of important coastal structures. Traditionally we solved this problem with organotins, but they are highly toxic, non-biodegradable and have long-term effects. There is an urgent need for nontoxic antifouling compounds. The occurrence of fully grown biofilm on marine algae is a rare event. Marine algae have effective defence mechanisms to prevent biofilm formation, such as QS inhibition, some of which have already been described. However, the authors believe we need to screen more seaweed for identification of new QS inhibitors.
 
In this investigation, Jha et al screened 30 marine algae extracts for their QS inhibitory potential by using a reporter strain Chromobacterium violaceum CV026. The C. violaceum CV026 is a mutant strain incapable of producing AHL signals making it a versatile and easy-to-use reporter that responds to extracellular AHLs. Samples that showed growth inhibition, as well as QS inhibition, were fractionated using five different solid phase extraction (SPE) cartridges, in order to separate two distinct activities, antibacterial and quorum sensing inhibition.
 
Amongst the 30 different algae seaweeds tested for quorum sensing inhibition using CV026 Jha et al found that Asparagopsis taxiformis showed the greatest QS inhibition. This was done by analysing the zones of inhibition found in the samples. The anti quorum sensing activity of A. taxiformis was further confirmed using the sensor strain, Serratia liquefaciens MG44.  This is the first report of the quorum sensing inhibition property of A. taxiformis and to successfully report capability of different SPE cartridges to separate quorum sensing inhibitor and antibacterial compounds which is important because it supports the notion that the QSI is based on the interference of bacterial signalling, rather than antibacterial activity. These results could be huge but I feel the authors have left their work unfinished here, therefore making the huge potential investigation negligible.  To my understanding of this study the Authors have not distinguished what compound is causing the QS inhibition that is reported. Jha et al successfully identified a compound which caused this inhibition but this was not identified, they suspect it is  “2-dodecanoyloxyethanesulfonate”. But what is it and was it in the other 29 algae? Is it completely new to science or is it a compound we already know of? If the latter then I feel we are back to square 1. What do you think?

Rotting wood never looked so good


The deep sea is completely devoid of sunlight, wind and other inputs of energy from the cosmos, and must rely on the leftovers of the world above it for energy and sustenance. These inputs are essential to life in this harsh environment and arrive in the form of sunken wood, whale carcasses, kelp and other debris soon forgotten or overlooked by sentient beings above.

When a piece of wood sinks to the deep ocean they form hotspots of biodiversity which have a high sulphide concentration and high reducing power, thus attracting chemoautotrophic bacteria. It is thought that these hotspots of sunken debris acted as stepping stones for the spread of life after its origin on hydrothermal vents through providing sulphide- and methane- rich niches on which overspill from crowded hydrothermal vents could thrive. This is an interesting concept and the study provides respectable evidence to support it and addresses several inconsistencies.

The authors aim to address several questions in this study, the most prominent of these being the question of how exactly the sulphic habitats attract chemosynthetic bacteria and the effect of this on the surrounding benthic communities. Ultimately this paper contributes to a better understanding of microbial ecology and biogeochemistry of the unique ecosystem created by falling wood and the role of these hotspots.

In order to study this, the authors deployed four replicate wood falls of a similar size, depth, weighting and branches at different distances to an active cold seep of the Central Nile deep-sea fan (Eastern Mediterranean). The replicates were revisited one year after deployment and smaller parts of the wood were enclosed for analysis. This method is interesting as the surrounding water was captured and analysed for sulphide levels, which is a good method, however laterally think it is easy to see that the clumsy useage of remote vehicles to collect the sample could easily brush off a significant proportion of the surface bacteria present and indeed, the authors do state that “Due to the sampling strategy on board, very small or rare organisms may have not been captured”. Subsequent biochemical tests performed include bacteria cell counts, DNA analysis and Automated RISA (ARISA) fingerprinting performed.  

The results of these tests show that despite major differences in community between the wood replicates, a core bacterial community is observable; predominantly sulphur reducing bacteria, which facilitates the development of sulfidic niches, building stepping stones for chemosynthetic life at these allochthonous habitats in the deep sea.

REF: Bienhold C, Pop Ristova P, Wenzhöfer F, Dittmar T, Boetius A (2013) How Deep-Sea Wood Falls Sustain Chemosynthetic Life. PLoS ONE 8(1): e53590. doi:10.1371/journal.pone.0053590
Accessed From: http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0053590

Sunday, 27 January 2013

A day in the life of wild planktonic microbial species



Planktonic microbial communities are able to adjust to environmental fluctuations by regulation gene expression in response to nutrient availability. However, detailed patterns of the dynamic responses of planktonic communities in situ are not well-described. Here Ottesen et al. face the challenge to study activities of a discrete picoplankton community over a time period of two days off the coast of California. They had to find a way to monitor gene expression of a diverse community over time, while taking into account the spatial heterogeneity of the ocean resulting from hydrodynamic processes, because samples from a fixed location may not represent the same coherent microbial population over time through the action of ocean currents. 

A robotic sampler, attached to a buoy, allowed drifting over 50 km with the currents and results confirmed that samples showed greater overall similarity in taxonomic composition to one another than did samples collected at a fixed station.  Every 4 hours samples for community RNA sequencing were collected and preserved. This analysis allows creating genome-wide transcriptome profiles of co-occurring taxa within a microbial assemblage; so basically, by matching gene sequences to a adequate database and by using various statistics (cluster analysis) and software packages (geneARMA), you are able to identify the genes expressed by each taxon and to create profiles showing the expression of those genes over time. 

Ottesen et al. focused on five microbial populations representing ecologically important clades of marine picoplankton:  Ostreococcus, Synechococcus, Pelagibacter,SAR86 cluster  Gammaproteobacteria (SAR86), and marine group II Euryarchaea (MGII). Consistent with previous laboratory studies, the photosynthetic Ostrococcus and Synechococcus exhibited strong diel rhythms over thousands of gene transcripts. For instance genes regulating carbon fixation, photosynthesis, oxidative phosphorylation, and respiration had periodic trends in transcript abundance, which relates to the photoautotrophic lifestyle of these organisms.

In contrast, none of the transcript from the heterotrophic populations was identified to be periodic. Instead Ottesen et al. observed “well-orchestrated”, genome-wide transcriptional regulation (particularly of genes involved in growth and nutrient acquisition) with strong correlation between some major metabolic pathways in Pelagibacter. Interestingly, SAR86 and MGII showed the same patterns of gene regulation, which seemed to reflect synchronous responses to the same environmental signal. Either each species population responds independently to the same environmental cues, or the transcriptional patterns are the result of interspecific communication and signalling e.g. by using auto-inducer molecules. Possibly, only one species senses the cues, which are then indirectly broadcast to others. But more transcriptional studies are required to determine the mechanism of these synchronised patterns. 

This study provides an insight into daily patterns of gene expression of dominant members of the picoplankton community and puts it into a temporal context. I was surprised by the high level of gene regulation occurring on a daily basis and in such a short period of time and it just shows how rapidly these organisms can respond to environmental changes. The suggestion that specific cues elicit cross-species coordination of gene expression among diverse microbial groups sounds promising and is worth following up. 

The study of these short time-scale microbe-environment and microbe-microbe interactions are crucial to understand the large scale patterns observed within populations and communities. 




Ottesen, E. A., Young, C. R., Eppley, J. M., Ryan, J. P., Chavez, F. P., Scholin, C. A. & DeLong, E. F. 2013. Pattern and synchrony of gene expression among sympatric marine microbial populations. Proceedings of the National Academy of Sciences, 1–10. Available at: http://www.pnas.org/cgi/doi/10.1073/pnas.1222099110

Vectors of neurotoxin: Krill and the diatoms they eat


The diatom Pseudo-nitzschia australis produces the neurotoxin domoic acid (DA), which has been responsible for the death of many marine mammals and birds. The authors of this paper have outlined the need to study the organisms which can act as vectors for the DA through the trophic levels and have elected to study Krill as an attempt to quantify whether or not enough DA could pass through the food web in high enough levels to constitute a threat to marine mammals. The study assayed the homogenised samples of Krill in order to determine the toxicity levels and found that the toxin was only present in Euphausia pacifica when there were toxic Pseudo-nitzschia spp in the water column. This implies that the toxins in these Krill pass through the digestive tract of the Krill without being re-distributed throughout the rest of the body. However, this does not mean that the Krill are not effective vectors of DA as the levels of toxicity within the sampled individuals was measured to reach 44µg which far exceeds the acceptable human allowance for DA intake. This represents a problem for marine mammals which prey upon Krill, such as some cetaceans, which can ingest, in some cases, 2 tonnes of Krill at a time. The authors pointed out that although in some cases, ie whales, this may still not deliver a high enough dosage of DA to be harmful, however, during diving alot of the blood will flow to the heart and brain, as per the mammalian diving adaptations. This will also deliver alot of the toxin away from detoxification systems and directly into contact with the more important organs of the animal, which can most likely result in death. The resulting work of this paper highlights the need for further study into vectors of toxic algae during bloom times as this will allow for a greater understanding of the ecology of harmful algal blooms and the possible effects that they could have for marine mammals and birds in the future. 

Bargu S., Powell CL., Coale SL., Busman M., Doucette GJ., Silver MW., 2002, Krill: A potential vector for domoic acid in marine food webs, Marine Ecology Progress Series, 237, 209-216

Harmful dinoflagellate blooms caused by Cochlodinium sp.: Global expansion and ecological strategies facilitating bloom formation:

 

 
As algal blooms are currently being studied in our lectures I figured this study/review was relevant. As dinoflagellates are one of the ever present causes of blooms and the Cochlodinium sp. is no exception, over the past two decades it has been witnessed that there has been a massive and global expansion of harmful algal blooms by this genus. These blooms are characterised by large red blooms that are easy to distinguish and recognise.

 

There are over 40 different species within this group; however this review is looking at only two of them as these are the ones that are the main harmful algal bloom producers, C.polykrikoides and C.fulvescens.  In the 1990’s these blooms were observed and recorded only in South East Asia and North America, nowadays the blooms have spread globally occupying areas such as the whole of Asia and Europe so a mass expansion has occurred within the past two decades.

 

The blooms caused by these two species are strongly ichthytoxic which essentially means poisonous to fish, however they do still cause mortality within other marine organisms. The compounds responsible for these blooms are still unknown and due to the recent expansion and sheer lack of evidence there is a lot less known about this species compared to other harmful algal bloom causing species.

 

This review in essence compares lots of studies and aspects that have been done on this species to try and come to a conclusion as to why this expansion has occurred so suddenly, obviously the key observations made are that there is an increase in anthropogenic factors and/or climate change, and this may be the case but as mentioned previously due to the sheer lack of evidence no firm conclusions can be drawn as yet.  The general methods used to study these blooms fall into 3 categories; microscopy, molecular techniques and remote sensing.

 

The areas this review looked at to study were temperature, salinity and grazing. The comparisons found that both species showed clear overlaps in temperature and salinity tolerances, with the general habitat and conditions for optimum growth being moderate temperatures and high salinities, these conditions are often associated with offshore tropical and sub-tropical areas. It was found that by altering their chain length these species could survive in altered conditions down to a low temperature of ten degrees but no lower, and all that they showed was mild stress. To further back this up it was observed in North America that some species preferred and could survive in areas that had cool water intrusions and more brackish salinities. So the results present the fact that just due to alterations physically these species have been able to relocate to new niches and survive with only a minor metabolic cost.

 

Regarding the grazing concept, as in any environment this species are mildly grazed but it was seen that when there is a lack of top down control grazing, or even a minor amount of grazing this species thrived by upping their chain length and therefore size, so it doesn’t appear that grazing or predation affects the growth or location of these blooms, evidently if there were predators in a large scale this might affect the locations but there haven’t been any noted. Finally it has been noted that under different conditions these two species have a wide range of nutrient strategies and so have a lot of flexibility, this leads to the hypothesis that with the fact that nutrients then shouldn’t be a problem they can survive and move out of their general habitat.

 

This hasn’t been studied as to whether this was a recently inherited thing or whether it has always been present but it poses a possible explanation with the fact that there is a such a large window of variability with optimum conditions I believe (and so do the authors) that this is how and why the dinoflagellate blooms have been able to thrive elsewhere. Especially if climate change is a driver, then an increase in water temperature will lead to this species being able to thrive a lot more, it obviously depends on what the salinity levels do but all that can be hypothesised is that with an increase in temperature there will be a dramatic increase in the abundance of these species and therefore algal blooms.

 

Although it is only a review and to be frank there isn’t that much out there as this review is from 2012 it is still important and poses a lot of useful information and potential for future studies. It’s only an up and coming field of study, but if these blooms are spreading and getting a sort of resistance and tolerance to changing conditions then it needs to be heavily studied before the whole range of oceans and their inhabitants are in trouble.

 

Reference:

 

Kudela,R., Gobler,C. (2012)  Harmful dinoflagellate blooms caused by Cochlodinium sp.: Global expansion and ecological strategies facilitating bloom formation. Harmful Algae 14, 71-86.

 

 

The paper can be located at this website:

 


 

 


Domoic acid contamination within eight representative species from the benthic food web of Monterey Bay, California, USA


Phytoplankton are at the base of many food webs, hence if these produce toxins that are harmful to organisms at higher trophic levels, and these toxins can be passed up via the food chain, they will have major implications for the health of those organisms. Domoic acid has been named as a toxin produced by phytoplankton that causes disorientation, memory loss, seizures, coma and death. It is possible for organisms to contain low levels of DA without exhibiting these effects. This allows for low levels in prey to be passed up to organisms through the food chain, resulting in higher concentrations at higher trophic levels. This process is understood in pelagic food webs but not benthic food webs. Kvitek1 et al (2008) investigated the levels of DA in organisms at different trophic levels of a benthic food web, using Pseudonitschia species as reference due to their common occurrence in the sampling area, and ability to produce DA.

Samples of organisms, and seawater from the top, middle, and bottom, were collected from Del Monte Beach in the southern bight of Monterey Bay, California. Eight benthic species, Urechis caupo, Emerita analoga, Nassarius fossatus, Citharichthys sordidus, Neotrypaea californiensis, Pagurus samuelis, Dendraster excentricus and Olivella biplicata, were chosen as they represent several different trophic levels, feeding styles and a link to higher organisms such as sea birds. Collection occurred 3 times a week in Pseudo-nitzschia australis bloom conditions and every 2 weeks when a bloom was lacking. Sampling spanned 2 distinct blooming events and one small bloom, from autumn 2000 to autumn 2001.
 
Pseudo-nitzschia species were identified by incubating small aliquots of water with fluorescently P. australis and P. multiseries (the local species) species-specific probes, then observed under a compound microscope and florescent light, from which volume-specific abundances were calculated. The concentration of DA was determined using high-performance liquid chromatography (HPLC) on filtered water column samples and extracts of homogenised animals. Emerita analoga was analysed used a reverse phase of HPLC. Several small organisms were homogenised from the same species to create equal volumes across all species. The efficiency of this method was tested by injected known amounts of DA standard into tissue of Urechis caupo and Dendraster excentricus, which had not been tested in previous research, and following the method of this experiment.Further investigation occurred into Urechis caupo due to no previous research confirming the presence of DA in their phylum, Echiura, despite DA detection for all samples during this experiment. Isolation of the peak fractions and selective ion monitoring of mass spectrometry data, then assays, confirmed the identity of DA and toxicity, respectively.

Statistical analysis occurred using Pearson correlation and paired t-tests to compare: water column DA concentrations and cell density of Pseudonitschia; water depth and water column DA concentration; and DA tissue concentrations during blooms and when a bloom was lacking. Additionally, linear regression was used to determine strength of the relationship between water column DA concentrations and cell density of Pseudonitschia.

The abundance of P. australis and P. multiseries peaked during blooms in autumn 2000, and spring and autumn 2001. Overall, the strength of the relationship between water column DA concentrations and cell density of Pseudonitschia was weak. Kvitek1 et al (2008) presumed that the DA was produced by pseudo-nitzschia species, however it is possible for other sources to produce these, such as other diatoms. It seems likely due to the independence of DA concentrations and P. australis and P. multiseries densities.  

The relationship between toxin levels and that of the water column, and toxin retention levels, varies between organisms of different trophic levels. If the hypothesis stated for the pelagic food web was also true for the benthic food web, an increase in toxin levels would be observed the higher the food web. However this is not the case. The average DA concentrations of the organisms during blooms were the highest for 2 filter-feeding species; moderate levels for the 2 scavengers, a predator, and a deposit-feeding shrimp; lowest levels for the remaining 2 deposit-feeders, with 1 also having the second feeding method of filter feeding. Logically speaking, the filter feeders should have high levels considering their direct ingestion of the Pseudonitschia species and toxins. The concentration appears to reduce further up the food chain, which could be a result of higher organisms feeding on other species that do not accumulate DA that have not been investigated in this study.
 
There was a low extraction efficiency noted for Dendraster excentricus, hence it might have been falsely identified as having lower levels than the other 2 filter feeders. On the same note, the efficiencies for the other species were not checked. Considering the validity of this method was shown as questionable, surely Kvitek1 et al (2008) would have checked.  They also state that 5 species contained significantly higher concentrations of DA during the blooms, and 3 species appear to retain the toxin momentarily after a bloom. This may not be retention but evidence that there is another source of DA in the environment.
 
They stated that DA was distributed evenly over all depths, therefore used the values of DA at the surface in comparisons between water content and animal content. When I looked at the data there did seem to be a peak of DA at 7m approximately twice the DA concentration of the surface and 25m. The species investigated inhabit different depths, and considering the variation in DA they should have compared the DA levels in the water at the same depth as the organism.

No comments were made on the health of the organisms when they were collected, and some collection techniques even involved killing the organism whilst in the water still. Considering DA is a neurotoxin and can cause behavioural changes, surely the behaviour of the animal should have been investigated even in brief to see if the levels present where effecting them.  

They also did not comment on the abundance of the animals present, neither those investigated or others, and the number of species considered is small. The weakness of the relationship could be due to the presence of organisms that interact with DA in some way. For example: uptake DA from the environment or alter the levels via some form of external detoxification system. 6 species exceeded levels of DA thought to be safe for higher level consumers. They could compare their findings to data collected on the abundance of higher trophic level organisms known to feed to these, or consider any reported mortalities that occurred at the time.

This study highlights the ability of domoic acid to manifest in these marine organisms of varying feeding styles and trophic levels, still within low levels overall. It suggests the transfer of the toxin is different in the benthic food web than in the pelagic food web in that concentrations of DA do not increase further up the trophic levels. However several more studies using different organisms, and considering different sources of the toxin, would be necessary to confirm the findings. The extraction method may also need changing as the one used is not efficient for all species. There is potential for cross-referencing to occur of this data with other data collected at the same time about other organisms inhabiting the area, and environmental changes. Due to the complexity of the food web this would be largely beneficial, although time consuming and impossible to do for all species.

Kvitek1, R. G; Goldberg, J. D; Smith, G. J; Doucette, G. J; Silver, M. W (2008) Domoic acid contamination within eight representative species from the benthic food web of Monterey Bay, California, USA, MARINE ECOLOGY PROGRESS SERIES, 367: 35–47