Lobster blood chemistry, and gruesome infestations.

I started talking about a little parasite in my previous blog, Nicothöe astaci. I realised it was getting a bit long so decided to split it into two - the first about histology of infected animals, and another, this, about physiological effects of the parasite on my lobster hosts!

So, after hearing more about this fascinating creature I wanted to know what it did and whether the parasite load, like the French scientist had mentioned, had an effect on the physiology or even the life, of the host...

I set off to Ilfracombe and Lundy, a place we had sampled before and knew for sure that there were pretty high levels of Nicothöe. It was here that the fishermen had pointed out the parasites to our research group in the first place! I joined forces with our favourite lobster fisherman Geoff and came back to Swansea with 18 lobsters (about 10 kilos) from various points around the Ilfracombe and Lundy coast. I let them acclimate for a few weeks in the aquarium to get used to the conditions before starting any experiments. Lots of things can stress a lobster out, including being caught in a lobster pot, handling and transportation so it's always good to do this when working with live animals from the wild.

As you can see from below, we had quite a range of parasite loads on our lobsters. It ranged from just a few to alot - infestation!!!

Photographs showing examples of (A,B) low and (C,D) high levels of Nicothoë astaci (arrows) in the gills of European lobster before (A,C) and after (B,D) excision. Inset shows the structure of the parasites. Note the high numbers of parasites at the base of the gills in the lobster with high parasite load (arrows). The excised gills show the arrangement of gills into outer, middle and inner sets. Photo taken from Davies et al. (2015)

A good way to test levels of stress or changes in a lobster (or any crustacean) physiology is by testing for changes in the composition of blood, or haemolymph. This was especially true in our case, since Nicothöe astaci is haematophagous, or blood sucking! I decided to test our lobster blood for 4 key components; haemocyanin, ammonia, glucose and total protein. Haemocyanins (sometimes spelled hemocyanin) are the crustacean version of our haemoglobin; proteins that transport oxygen throughout the body. Haemocyanins  contain two copper atoms that bind a single oxygen molecule (remember it's O2) and the reason that you hear many people saying that lobster/crab blood is blue (this is not strictly true - more of this later!). Unlike the haemoglobin in red blood cells found in vertebrates, haemocyanins are not bound to blood cells but are instead suspended directly in the haemolymph. 

We also tested for total haemolymph protein - this is because haemocyanins are not just oxygen carriers. They make up approximate 80-90% of total haemolymph protein (although this changes depending on whose papers you read!) and are an important component in some invertebrate immune systems. In arthropods (crabs, lobsters etc.) the haemocyanin family includes phenoloxidases, hexamerins, pseudohemocyanins or cryptocyanins and (dipteran) hexamerin receptors. Phenoloxidase are copper containing tyrosinases,  proteins involved in the process of sclerotization of arthropod cuticle, wound healing, and humoral immune defenses. For me, testing for haemocyanin is a win-win, not only are our parasites located on the gills, where key oxygen exchange occurs, but then suck the blood, so we hoped that testing for this would give us some answers. Questions here were:

1. Does the presence of the parasite hinder oxygen transfer across the gills?

2. Does the blood sucking activity of the parasite deplete oxygen levels in the haemolymph?

3. Does the presence of the parasite deplete haemocyanin (i.e. is the % of haemocyanin in total protein higher or lower than averages)

Aquatic crustaceans excrete the nitrogen derived from protein and amino acid catabolism primarily through the gills, the gut and the antennal/green glands. Nitrogenous waste in lobsters is made up of urea, ammonia and amino acid compounds; the major excretory product is ammonia. The concentration of this waste in the haemolymph changes in response to stress and ecdysis wherefore we tested for changes in ammonia levels.

Questions included:
1. Does the presence of the parasite hinder ammonia excretion?

2. Is the presence of the parasite increasing stress-induced ammonia levels?

Finally, we tested for glucose. Glucose levels have been shown to change in line with lobster stress levels and we thought it might be affected by the parasites attaching to the gills.

So, what did we find? Safe so say, as expected, there was a positive correlation with the amount of parasites and total protein. This means that as the number of parasites on a lobster increases, so does the amount of protein in the blood. Sounds weird, until you see that the haemocyanin also increases, and it makes up 84% of the total protein in the haemolymph we tested. So, the real story here is an increase in haemocyanin as parasite load increases. We think this is the lobster most likely compensating for reduced respiratory function due to gill damage caused by the parasite. Increased haemocyanin, may therefore be advantageous for infected lobsters.

There was also a slight, but not significant, correlation with ammonia and glucose (see figure below). It could be that ammonia and glucose are not really affected by the parasites, or, as in another study, parasites can absorb glucose from the haemolymph, thereby forcing the host to resupply tissues with this sugar from glycogen reserves in the hepatopancreas in order to maintain carbohydrate homeostasis. As for the ammonia, some studies have shown a switch in nitrogenous wastes to products such as urate or urea... which we didn't test for.

This figure, taken from my paper Davies et al. (2015) shows the results of a Spearman’s correlation coefficient analysis.  You can see correlations between parasite numbers and haemolymph concentrations of (A) total protein (p = 0.02), (B) haemocyanin (p = 0.0065), (C) glucose (p = 0.2112) and (D) ammonia (p = 0.1290). Asterisks denote significance. 

As always, you can email me, tweet me, or add me on LinkedIn. I am happy to send over copies of my papers or answer questions! 

Team lobster! A meeting at the National Lobster Hatchery

The National Lobster Hatchery, Padstow

Last Friday, I was asked to attend a meeting at the National Lobster Hatchery in Padstow, Cornwall. I know I've blogged about it in the past, but collaboration with other research groups is, to me, one of the most important factors when conducting a project. As well as a hub for lobster science in the UK, the national lobster hatchery has great ties with the local fishermen, the Padstow community and restaurants in the area, with their very successful 'buy one set one free' campaign.

I've liaised with Padstow in the past - when I had visiting researchers in my laboratory from the New England Aquarium, Boston, this was on our list of places to visit; and when I needed juvenile lobsters for exposure studies, I would contact the hatchery. It was only at the 10th International Conferenceon Lobster Biology and Management, that I really got to know the 'hatchery lot' and we became great friends, and colleagues. We vowed that we would stay in touch and try to meet as often as possible in order to discuss the future of European lobster research in the UK.

In attendance at the meeting on Friday were hatchery staff Dom Boothroyd, the general manager; Research & Development Officer Dr Carly Daniels; Business Development Officer, Clare Stanley and PhD student Charlie Ellis, who is part of the University of Exeter's Falmouth Campus, but works closely with the hatchery on his research project. 

A lobster with one of Dans tags on it's 'arm' 

First up to talk was Dr. Daniel Skerritt, who completed his PhD at Newcastle University last year, investigating lobster abundance and movements in Northumberland. Dan now works as a consultant for MRAG in London and gave us a talk about his research findings both during and post- PhD. For his project, Dan monitored lobster behaviour in and around baited pots (used to catch lobsters), and their interactions with habitat using acoustic telemetry. Perhaps his most significant findings which may have the greatest implication to management, concern differences between the sexes. From mark-recapture studies (where a lobster is tagged, released, and caught again) he found that males have a much higher catchability than females. This means that a lot more male lobsters were recaptured – but why? The acoustic telemetry work revealed further differences between the sexes; males use a much larger area of seafloor than females, which could account for this increased catchability due to greater probability of pot-interaction. However, overall this work focused on the utilisation and behavioural changes over substrate. Dan has a publication in press for Marine Ecology Progress Series; “Fine-scale movement, activity patterns and home-ranges of European lobster Homarus gammarus” the prepress abstract can be viewed here.

Aside from his science, Dan has also been involved in some outreach work. He struck up an interesting collaboration with a graphic designer and the Great North Museum. They put on an exhibition with input from Natural History Museum called ‘Spineless’, with Dan’s work being the subject of one of the exhibits. The aim of the collaboration was to make the kids of the northeast aware of the importance of the lobster fishery; you can see more about the exhibit here.

Check out this great little video of Dan, talking about his research.

Lobsters from Roots and Wings on Vimeo.

A snippet from my lecture 

Up next, I gave a talk about the main findings from my PhD; I have talked mainly at conferences about my shell disease susceptibility work but my lesser known research concerning parasites (see last weeks blog post... and more in next weeks!) and MPAs, were very interesting to share. It's great to talk informally about this, and to get some ideas together for future work. 

Charlie is currently writing up his PhD and gave us a short overview of his findings so far. The National Lobster Hatchery's main mission is to create a sustainable lobster fishery in Cornwall and in order to do this, the number one research priority is to monitor the success of it's primary charitable objective (i.e. the stock enhancement program). In order to do this, they must be able to estimate survival rates for hatchery reared lobsters in the wild, as well as their contribution to catches of landing-sized European lobster. To do this, genetic analysis of Cornish lobster stocks is essential, and something that Charlie has been working on. He has also been examining tagging systems that will enable stakeholders to easily identify hatchery reared animals. So far, Charlie has found that the lobsters around the Cornish coast all seem to come from one gene pool, which is good for the release programme which relies on volunteers to bring in berried hens (expectant lobster mums) from various locations. 

Spot the baby lobsters!

I think we are a very talkative lot so we didn't have much time for poor Carly to talk to us about her new and exciting project which focuses on developing sea based culture of lobsters in containers, a rearing technique that exhibits the potential for a low carbon form of rearing with no feed costs. This is a consortium project, led by the National Lobster Hatchery, which follows on from an earlier project also funnded by Innovate UK/BBSRC. Carly completed both her BSc and PhD projects at the hatchery, concentrating on the optimisation of the rearing diets for early life stages of the European lobster, in order to enhance growth, survival and health using biotic dietary supplements.

The hatchery also hosts students who work on small but important projects and so we also heard interesting presentations from Dan Sankey, who is working on lobster behaviour and is soon to begin an MRes at Swansea University; and Grace Dugdale, a BSc student at Cardiff University who is working on a placement year alongside Carly at the hatchery. Grace is looking into the effects of probiotics on lobster juveniles. Also in attendance were Adam Bates, who is working towards an MPhil in European lobster genomics and Joe Augier who previously completed his undergraduate project at the hatchery and is going on to do an MRes.

In all, it was a great way to reconnect with the lobster team, over a year after meeting at the ICWL in Mexico. I would like to acknowledge all in #Teamlobster for helping me to write this blog post… lobster scientists, unite!

Ultimo pez

I know, I know, I have been slacking in the blogging department. Although, I do have a good excuse! This weekend I finished my thesis. Yup, you read that right. FINISHED. THESIS. It feels good to say that. The last few months have been a bit of a whirlwind, I'm not quite sure where all the time has gone (or whether I have a social life anymore?) but all I know is that I am done. I'm not quite sure if I can feel the relief yet.

Another reason for me being so busy these past few weeks has been because it's the start of term again, and you know what that means? Yup, the start of the teaching semester! As you will know if you have read my blog in the past, I am a 'self-funded' PhD student, so teaching has been a major part of my PhD. This year I have been helping out my supervisor with Cell and Immunobiology again, and started assisting with a new module; Molecular Ecology! We are teaching the second years how to do DNA extractions and PCR - wahoo - wish they did that when I was doing my undergraduate degree! I have also been helping out teaching an R module to the new cohort of Masters Students and a plant module to the first years.

I have also been guest lecturing again for the third year Diseases of Aquatic Organisms module, my favourite! Luckily, I already had a talk prepared, because in September I was invited to speak at a very cool conference. The Annual Meeting of the National Reference Laboratories for Crustacean Diseases takes place, as the name suggests, annually. This year was the 6th meeting, which took place at Cefas, Weymouth. The Weymouth lab is the European Union Reference Laboratory for Crustacean Diseases and I was invited by the director, Dr. Grant Stentiford, who will also be the external examiner for my PhD. I was asked to give an overview talk on shell disease, as part of the 'eDNA and invertebrate pathogens' session, which was really exciting. I was able to meet a bunch of international researchers interested in the same subject area as me. I was also able to get a tour of the department from part-time PhD student and EURL Coordinator, Kelly Bateman. Weymouth is a lovely little town and I had a little time to explore before catching my train home on the Friday evening.

So what next?! Well that's the big question. Obviously I have to wait until December for my viva (or 'defender', as the rest of the world calls it) and I have a few ideas up my sleeve and a couple more papers to send off. I have already applied for funding for some work abroad next year, will keep everyone posted. I will also be applying for some more in the coming weeks so it's just a case of waiting on decisions... I did get a little over excited the other night and treated myself to a new tripod and aperture timer so watch this space for some cool time-lapse videos (spare time... what are you!?) Before I go, here are some photos of a contented PhD student in her natural habitat: