Good morning, everyone. Thank you for joining us in this session on fish medicine 101, focusing on the diagnosis of diseases for the fish patient. My name is Esteban Soto.
I am a professor of aquatic animal health at the University of California, Davis. And my work focuses on a clinical research and didactic on aquatic animal medicine. So I appreciate the opportunity to share with you some of these topics today.
The material that is going to be presented has been acquired from a multitude of great collaborators as well as online and Polish sources, and it's intended for educational purposes. Very straightforward, the learning objectives, you know, for this session, although we're not gonna have, the opportunity to perform a laboratory, we are going to cover basically the practises, you know, for how to collect, you know, diagnostic samples, the value of these diagnostic samples, how to process them, and, of course, you know, this will entitle collection samples, you know, from In vivo, so basically a live patient as well as how to perform a full necropsy in the case, you know, where you have to perform such a procedure on on a fish. We're gonna outline methodologies, you know, for this sample collection and a submission for histology, microscopic, microbiological, and molecular analysis.
And we're gonna also cover basically the principles of anaesthesia and euthanasia, particularly for the most common methods, you know, that we utilise in fish. So very briefly and in very generalistic terms, this is kind of the layout that we follow for diagnosis of diseases in fish. So to approach our patient, you know, when it's presented to us, OK?
As in many, many, many, many cases, you know, of course, you know, for most of you, it might be challenging, you know, to get a complete history from your patient, you know, but that initial discussion with your client is absolutely critical in the United States, you know. As in many other countries, we have to follow have first, you know, a veterinary client patient relationship in order to be able to provide veterinary services, you know, to, our, patients. So acquiring a thorough history is of absolute critical importance.
OK? For this, and as I'm gonna present in the next slide, you know, it will be a deal for you to develop a, . A document, you know, where you can go in an organised matter, collecting all of the information, you know, from the system, you know, and the water, the environment where the fish lives, and of course, you know, then on your patient.
With fish medicine, we deal with two different patients. We deal with the water and we deal with the fish. And the fish veterinarian needs to be familiar with some of these chemistry on on water samples.
Probably some of you, Always wonder, as I did, you know, when I was in high school and in college, you know, what good chemistry will bring to us. Well, if you want to work with fish medicine, you know, some of this chemistry does have to come back because a lot of these parameters do interact, you know, within each other and it is important that we have an understanding on how this interaction works and on how it can affect the health of the animal. OK?
On site, so basically, on the aquarium or the pond, you know, whatever that system is that is holding your patient, there will be some parameters that you need to collect, on site, like the oxygen and the temperature. For the other parameters, they can collect the water sample and bring to your clinic and you can evaluate them in there. But we're gonna talk a little bit more in depth on history in just a couple of minutes.
The water sampled, you know, the water quality parameters that we typically measure are presented in here. However, those are not all of the ones that we can evaluate. You know, in some cases, we have to go a little bit deeper.
For example, if there's a suspicion of gas bubble disease, you know, or supersaturation of the water. You need to be able to evaluate total dissolved gases, you know, which could be a little bit complicated. So, of course, you know, if you go into this line of work, and it's gonna be important also to familiarise, you know, as to which laboratories can run, you know, a full analysis of the water quality.
The evaluation of the fish on site is also very important. So before they bring the animal to your clinic, in some cases, this is possible. In some cases, of course, you know, the veterinarian will have to visit, you know, the facility.
But evaluating that animal in their natural setting is going to be very important, you know, to really assess, you know, that behaviour of the animal and start seeing some of the physical abnormalities. If you're not able to visit your client, of course, collecting photos or videos, you know, will be highly, highly valuable for you to start generating a differential diagnosis and particularly also very important for you to then communicate with your client in If it is, you know, what the prognosis might be for that patient. In some cases, you have to be, of course, you know, a little bit, not a little bit, you always have to be like, you know, very sincere and share what the prognosis is going to be.
In some cases, unfortunately, the recommendation will be to euthanize the animal. But then the euthanasia of this animal can provide you a great source of material when you perform the necrosis that will allow you Particularly when you're dealing with population level cases, you know, to really, really found, you know, the culprit of of the mortality events, OK? So, again, sometimes, you know, you have to approach, you know, these cases at a population level, in some cases, you know, as the individual level.
In a live animal, there's a number of samples, of course, you know, that you can collect, you know, skin g biopsies, blood that can be used for serology, can be used for clean paths, for microbiology, etc. You can do a number of imaging methods, endoscopy, radiologicalology trasonography, even CT scans nowadays. But again, the veterinarian has to familiarise itself, you know, with the anatomy of the animal and how to conduct, you know, a necropsy and collect, you know, good samples, you know, for them moving downstream for other diagnostic methods, microbiology, molecular histological, etc.
Analysis. OK? There are some, there are most of of all of these methods, you know, are, you know, generalistic and very similar as to what you do in mammalians and avian species, you know, but of course, with the poison patient, you know, there are some intrinsic differences, you know, for example, the blood clots very fast in the fish patient.
Alysis occurs extremely fast, you know, so, once an animal dies, you know, you need to perform that necropsy ASAP. You cannot just put in the refrigerator and do it the next day. As you know, it can happen, you know, with some of the mammalian patients.
Microbiology, of course, you know, they need to be familiarised, you know, with what type of media to use, you know, for the fish patients because the bacteria and viruses will not grow in some of the media that we use, you know, for mammalians. And of course, you know, the temperature, you know, of incubation, you know, The fish, most of the fish organisms will not grow above 30 °C. If you're dealing with a cold water species, you know, the ideal temperature will be below 20 °C, whereas if you're dealing with a warmer fish species, you know, probably between 25 and 30.
So all of these intrinsic differences are important that the veterinary communicates very clearly with the Diagnostic laboratory so that the samples are processed in the right manner. OK. This is the aquatic Animal Health Forum, for example, that we utilise at UC Davis, and it is important, you know, that we have this, you know, we share it with our residents, with the house officers and the fellows, etc.
For them to be able to go in a systematic way on this collection of the history. So we shared this document with our clients beforehand, you know. Have them fill it out, you know, and then, share it with us together with videos and photos, you know, of our patient.
Then we call them and we go through this form, you know, to basically provide them a quote, you know, and of course, you know, before we see the patient. This is important, you know, because, of course, it allows us to prepare a potential, differential diagnosis. And then better, assess, you know, kind of, you know, the future of that animal, the prognosis, you know, of the case, but also to provide a reliable estimate, you know, for our clients to know, again, you know, how much, you know, it will cost, you know, the analysis, you know, and of course, you know, let them make the decision if moving forward or Not.
OK? So again, it includes information of the actual system, you know, on the nutrition, you know, aquatic environment, you know, and of course, you know, then on the reasons for the visit. I would love to say, you know, that a lot of our clients, you know, are, you know, providing a preventative care, you know, for the animals, although.
You know, this has been changing, I think, you know, as they gain, you know, a better understanding on the importance, you know, of the value of the veterinarian in this field. Still, you know, most of this is a reactionary, and they will call us only when there is an issue. OK.
So, of course, you know, we have to provide value for our services, you know, and that, you know, starts with education, etc. So it has improved, but there is still a long way to go. So, most of the times, you know, again, you know, it's more reactionary, and at this point, you know, it's where, you know, the reason why they're calling you, our fish are dying, the fish are not eating, the fish are not reproducing, etc.
Etc. OK. So, as I mentioned again, you know, these are gonna be the steps, you know, so we're gonna go one by one.
In the case history, again, you know, one of the most important pieces of information that we need to get are these morbidity or mortality patterns, OK? And as with other animal species, you know, they will start allowing us, you know, to formulate a differential diagnosis. For example, if we have a case.
You know, where there is an overnight death of animals, you know, such as in the lower figure in here, where we have hundreds if not thousands of dead fish overnight, it is very unlikely that it's going to be an infectious agent, you know, causing that overnight mortality. In most cases, you know, that will, occur if there is a toxic problem. So, for example, bleach that may, may, leak you know, into the ponds, or for example, copper treatment, you know, in, in an incorrect manner.
Or in a very commonly, oxygen depletion, for example, in this case, you know, in the southeastern United States, during the summer, very common to have algae blooms. And if there is an algae bloom, you know, and there is no extra oxygenation presented, you know, for these systems, you know, during the night, that algae will consume all of the oxygen and will cause, you know, a level of anoxia that will then kill all of the, the fish in the system. OK?
Most of our infectious organisms, we actually see them more in an acute to super acute manner. So, basically, you know, within weeks, you know, of a stressor event. And this stressor event could be like handling, could be introducing new animals into the system.
Etc. And that again, you know, of course, you know, provides an immune suppression and then typically, you know, portunistic agents, you know, like aeromonas, you know, that can then go ahead and cause an infection and mortalities in the system. But also, there could be, you know, low mortalities, you know, that, will occur over an extended period of time, you know, there's some, Microorganisms that of course we have to consider, such as mycobacterium or bacterium in different species, but also could be again, some of these water quality values or poor nutrition or some toxins present in the water, that again, you know, can cause consistent, like, you know, more chronic, you know, immunosuppression and of course, later on, you know, the death of the animal.
So all of this is important again, you know, to acquire during the history because it starts telling us what type of analysis I really need to perform and how urgent it will be. For example, in this case, again, you know, with the hyper acute or per acute mortality events overnight, measuring the amount of dissolved oxygen early in the morning or collecting water samples and submit for the lab for toxicological screening of chlorine will be, of course, you know, pretty important. OK, so what do our clients need, you know, if they are interested in either bringing us, you know, some samples or, you know, basically, contacting us, you know, for a submission.
OK? So there's different ways, again, you know, if the client is able to bring the fish, you know, there will be one to the clinic, that will be one option, you know, in some cases, we need to go ahead and visit, you know, the client, you know, in their house, you know, because they're dealing with very large number of animals or a very large patient. For example, we have Dealt with a sturgeons or a koi that are pretty large, you know, in size and very difficult to move.
So, if our clients are bringing fish patients, you know, to the clinic, what do they need? Well, if their fish are small in a cooler, such as this one, or particularly one with wheels that could be beneficially easier to transport. They can bring, you know, the patient, you know, in, in this, this manner.
Very important, you know, important to bring it, you know, with the proper aeration. So, some of these portable pumps, you know, and that will provide, you know, you know. Oxygenation of the water as the fish is moved, you know, from their house, you know, into the clinic.
OK? And always important, you know, to recommend bringing a sample of the water from the system so that we can evaluate also the water quality parameters. In larger size fish, you know, well, this changes a little bit just by size, you know, but there's also other options, you know, they can bring, you know, a larger cooler, such as this one, which, of course, in the caveat that, you know, the folks moving the fish, you know, need to be strong enough, you know, to really be able to move this volume of water, which could be, you know, pretty heavy.
And in some cases, even Using a tarp cover, for example, you know, in the, in the bed, you know, the pickup truck has worked, you know, to move, you know, large number of fish as one, you know, for evaluation. So, again, a little bit of a MacGyver, type situations in here where, you know, imagination has to be let loose, you know, of course, with guidance, so that these animals, you know, can be transported in a safe manner. If we visit our clients, you know, what do we need, you know, to go ahead and provide veterinary services in the field.
We typically bring a table, you know, again, you know, just, you know, to put our equipment, the microscope, etc. In some cases, even the fish patient, you know, will go in this table, nets and buckets, you know, in the, in the different buckets, you know, systems, you know, we need for an anaesthesia, of course, that will mandate, you know, depending on the size of the animals, we have multiple size. From 10 litres all the way to 70 litres, depending on the size of the fish.
In some cases, you know, the owners will have containers already prepared, you know, so we can use, you know, their containers to anaesthetize the animal. But important, you know, we have to have a manner to measure, of course, the anaesthetic, you know, so that we provide the right type of anaesthesia. This type of nets, the sock nets, you know, that have an impermeable side, you know, and then, of course, a permeable one, are pretty important, you know, to avoid lesions, you know, in the skin or in the fins of, of a larger size fish, you know, for example, we need to use those, you know, for, koi.
And it's important, you know, because as we move them, you know, from their container, we can drain all of the water and then just move, you know, the the koi itself. Little wagons, you know, transport wagons like this, you know, to basically allows you to carry all of the material, portable microscope, of course, critical, so that we can evaluate the skin scrapes, skill clips, even blood smears, you know, if, if we were inclined to do so in the field and provide, you know, that evaluation initially. And then, of course, you know, box, you know, so that we can transport our equipment.
In some cases, you know, we can do some, surgical methods, you know, in the field, you know, we carry liquid nitrogen, since we see a lot of, chromatoporromas in some of our patients. And, these allow us, you know, of course, you know, to, provide, you know, some of these services, you know, in the field while the fish is under anaesthesia. There's different types, you know, that can be used.
And again, you know, this will depend on, legislation and, the ease of acquiring some of these equipment, you know, in your different countries. But, but again, you know, certainly something that we commonly see and has been particularly useful, you know, for us to provide some of these, again, you know, surgical. Methods, you know, in the field.
And then, you know, a, a portable ultrasound, of course, you know, we understand, you know, there might be economic complications with this, you know, but it has improved, you know, tremendously. And, now we have, you know, very good. And, again, you know, I wouldn't say cheap, you know, but they are certainly, more comfortable, like, you know, with prices, you know, in different places that you can use and then provide, you know, these, ultra sonography on site.
You know, again, particularly with some of these. Animals, you know, when they have like egg binding, or they have, you know, a elloic distension, you know, or even, if they have, you know, exoalmia, like, you know, using some of these diagnostic methods, you know, in the field, you know, could be of a very, very, very suitable basically for those situations. OK.
And then water quality kids, you know, if you get into this field, you know, as I mentioned before, you have to patients. You have the water and you have the fish. And it's very important that you understand the value of the water, quality, OK?
You need to understand how to measure it, you know, the pros and cons, you know, there's all of this equipment, you know, you can purchase, online, of course, you know, there's pros and cons for each one of them. The strips, you know, are typically the cheaper ones, you know, but again, are a little bit less reliable. You have to really be aware of the expiration dates, you know, for this equipment.
Because, of course, you know, if you're charging, you know, service, you know, for this, you want it to be accurate. So the strips, you know, again, you know, they have some use. I will not say that they're useless, you know, but are some, you know, that we might recommend, you know, the client, you know, to have, you know, so that they can monitor some of the parameters.
Even better, this type of, titration type kits, you know, you know, they cost, you know, about 30 to 40 bucks, here in the US, you know, a little bit cheaper, you know, more reliable data. And of course, you know, the more, you know, professional systems, you know, like these ones, you know, we use. Different ones, you know, that have been recommended, you know, I will not promote any brand, you know, but, you know, if you're interested, you know, in, in hearing what options, you know, we use, you know, email me, and, and I can, provide, you know, some of the answers.
But, other colleagues, you know, might use different ones. My point in here is that if you are providing, you know, evaluation for fish patients, you know, you also have to be able to evaluate the water quality. All right.
So now we start dealing with the evaluation on site, you know. So these, for example, are videos, you know, that different clients have shared with me. So that you can start assessing, you know, these, behaviour, these clinical signs, you know, fish and then start formulating a differential diagnosis.
So, again, all of these are different presentations that might start indicating different things. We're going to play the first video and see what you what you think, OK? So you see the goldfish, you know, as swimming.
He's having a hard time swimming. You can see the effort of the fins moving, you know, and he's not able to maintain position in the water, and it just presents with negative buoyancy. OK?
So, that's the first case, you know, in the second one, we have tilapia fingerlings, some of them again, lying in the bottom, some of them in the side, but some of them like twirling or circling around, again, abnormal presentation. So which samples would you prioritise? OK.
Think about it. In the first case, you know, of the goldfish, you know, this abnormal swimming, you know, that omic distension that is presented. You know, really, really suggests to me, you know, that there is issues with a swim bladder.
Something is happening, you know, to that swim bladder that is not allowing it, you know, to maintain position. In this specific case, it was due to polycystic kidney disease, you know, so there is a lesion. In the kidney that is just generates generating displacement of that swim bladder is not able to allow it, you know, to maintain position in the water.
It is completely compressed, you know, and of course, then the fish basically has to sink into the bottom. That excessive muscle muscular movement and the fin movement is the fish, you know, just trying to compensate, you know, and maintain position in the water, but of course it's exhausted and and then just presents this negative buoyancy. In the case of the tilapia fingerlings, you know, what it really suggests to me is something at the level of the brain.
OK. Fish are lethargic, you know, fish are in their sizes, and then fish are presenting that really abnormal swimming. In this case, you know, particularly, it was a bacterial infection that has gone into the brain, it's causing a meningoencephalitis.
And of course, you know, that is causing neurological clinical signs, you know, as the ones that we observed. OK? So, again, The value of these videos is that start allowing you to formulate, you know, some of these differentials.
In some cases, as I mentioned, you know, the patient will come to you, in the ideal world, you know, the patient will come, you know, with your client, you know, and then you will be able to evaluate it, but the reality of it is that sometimes, you know, they have to be shipped, you know, so that you can evaluate them. So, particularly again at the point of a population medicine scenario. You will receive, you know, samples, you know, that again, you know, should allow you to start formulating a good diagnosis.
It is very, very important that you educate your client as to what type of samples to send. The best samples to evaluate are moribund fish. So again, you know, those that haven't died but are presenting typical clinical signs of disease.
If those animals can be collected, they could be euthanized and shipped. They can be shipped in on water sometimes, but again, you know, that might be a little bit more complicated and not possible in some scenarios, you know, if they have to be shipped, and they cannot be shipped in on water, they should not be frozen. They should be shipped in individual bags, you know, and chilled, you know, basically to refrigerator temperatures.
Those samples need to be processed as soon as possible. Again, critical that they do not send dead fish. It is easier to catch a dead fish than a live fish, you know.
So if given the option, every single client in the world is going to send you a dead animal. OK? Do not accept it.
Don't lose your time, you know, working with them. I understand that sometimes, you know, it's going to be the only sample that you can evaluate and then you have to make a decision if it's worth it or not. But know that again, you know, the fish patient becomes out ofly very fast.
There are changes, you know, histologically that will occur really fast. There will be opportunistic bacteria that will take over, not only from the gut of the animal, but also outside, you know, the gills in the skin, you know, again, all of these, is going to have to be very, very, very well understood, you know, because again, Even you are isolate, you know, organisms, you know, from internal organs, you have to have in your mind that it could be completely incidental and due to the, less than ideal sample that was submitted for diagnosis. And additionally, a number of parasites, you know, that might be very easy to diagnose, you know, in a fresh animal.
Will leave the dead host, you know, or they will die themselves, you know, so, they might complicate, you know, even more the analysis. If you are receiving the, the samples, the client is shipping them, you have to take time, you know, to educate them also on how to send those samples. And again, there's different regulations in different parts of the world as to how, you know, samples, you know, can be submitted, you know, from, from animals.
But, you know, some very, very general recommendations pad the sample container to avoid breakage or spillage, you know, sometimes, you know, we triple back them. Position the samples upright, you know, particularly if they're receiving, you know, with water. Use pack material to hold samples in place, you know, and again, a submission form, you know, needs to be received, you know, in sealed plastic, additionally, by email.
So, for example, this is a sample, a very poor sample that I received, you know, a while back. So you can see, again, you know, I asked, you know, for a close container, they send like, you know, these laminated syrophones. That, of course, you know, you know, didn't really hold any material.
I, I asked, you know, for ice packs, you know, they put a bag and fill it with regular ice that melted very fast, leaked, and then, you know, provided, you know, again, the breakage, you know, in the carton box, etc. Versus this. So basically, this is what I wanted, you know, nicely sealed, Styrofoam cooler, a little bit again, .
Lighter for shipping, ice packs, and then weld label and triple back, you know, material where the sample then, you know, can be easily submitted. OK? So, educating your client, you know, will be important.
The number of fish to evaluate, of course, is gonna be also critical. . In some cases, you know, the ideal scenario for me, if you're talking about a population level type scenario, even in ornamentals, you know, in food fish, I typically ask, you know, between 5 to 10 clinical affected animals.
Depending on the size of the animals, of course, you know, less might be needed, you know, if they're like larger size fish, you know, you can collect multiple samples, you know, from the same animal for histology, for microbiology, etc. In some cases, you know, if you're dealing with guppies, for example, or you're you're dealing with tetras or you're dealing with zebra fish, you know, very tiny fish or with fry of any species, you know, they will be very tiny. So then you have to dedicate, you know, an entire fish, you know, for histological evaluation and then another sample for virology, and another sample for microbiology.
So then you have to ask, you know, for more animals. There are some guidelines, you know, for diagnosis of disease that the Blue Book for the American Fisheries Society is going through some Revisions, you know, so stay tuned because this might provide, you know, very valuable, information, you know, in terms of, fit for purpose, you know, diagnostic methods, you know, recommendations. Again, at the end, all of it is guidelines, and you have to obey, regional, as well as national regulations as to what test is acceptable, etc.
Etc. But just to put you an example, you know, if you're dealing with a clinical infections, you know, and you are testing a group of animals, for example, for interstate or international movement, there might be some guidance as to the number of animals to evaluate. The WOHA, what was previously known as the OIE, so the World organisation of Animal Health, also has, you know, guidelines, you know, for moving international samples, you know, against selected organisms.
And again, you know, In the United States we have the blue book. Currently, for example, this is one of the recommendations that is given. So depending on the lot size that you're moving and depending on the the prevalence, you know, that is suspected, then the number of animals, you know, that you will need to evaluate to provide a 95% confidence, you know, that at least you will be able to detect one infected animal in the sample.
OK? So again, these guidelines exist, you know, from different regulatory bodies in different regions. So, again, the veterinarian needs to be familiarised, you know, with what to sample and and how to process those samples.
OK, sedation and anaesthesia, of course, you know, is gonna be critical, you know, for a number of our patients, you know, these. These patients, you know, will become really stressed, you know, adjust with, handling, you know, so reducing these stressors, you know, is gonna be. Critical you know for the way that they respond to treatment.
And, and of course, you know, with, the welfare of the animals, you know, something that it is absolutely critical that you are fully aware, your clients are aware with it and will expect the utmost, you know, respect, you know, from the veterinarians. So familiarising yourself, you know, with different drugs and products that allow, safe, you know, sedation and anaesthesia is going to be important. Probably the most common one is, MS222, methylsulfonate.
Also known as trica, in this case, you know, this product again is commercially available, is pretty safe, you know, it's one that, has been investigated for decades. And it is, fairly safe, you know, for a number of species. However, different fish species have different susceptibility to this drug.
One important key point, you know, is that it is an acid. So it has to be buffer. You know, with bicarbonate, we use in this case, baking soda at a ratio of 1 to 1 or 1 to 2.
So basically MS2 to sodium bicarbonate, OK? Depending on the alkalinity of the water, you might have to go higher with the bicarbonate or not. But again, different species have different susceptibility.
For example, salmonids are very susceptible to this, whereas cichlids, you know, for example, are fairly resistant. Cypri it, yo, goldfish are fairly resistant to it. So depending if you are looking for a light you know sedation or anaesthesia, and of course, depending on the plane of the anaesthesia, you will need to add less or more of this product, OK?
So, so it's again, you know, important to familiarise yourself. If you don't know what concentration to use, look in the literature or contact colleagues, you know, that might have work, you know, with the fish species that you're going to be working with, to provide, you know, some recommendations. OK?
For uhOI, for example, you know, if I am sedating them, you know, I use anything from 40 to 50. But if I need anaesthesia because I am collecting gill samples or blood, I will use, you know, 120 to 150 milligrammes per litre to knock that fish, in a safe manner that I can then, of course, you know, collect a suitable, sample for diagnosis. OK.
In other countries, Eugenol, Io Eugenol and club oil are commonly used. Again, different countries, you know, have different, approvals, you know, for different drugs. Acquiescinol, for example, is commonly used in Latin America, but it's one that is still not approved inoofish by the FDA here in the US, you know, so again, you have to familiarise as to legislation.
In your different countries. But again, these products, you know, that are, you know, coming out from compounds of club oil are, are also frequently used in fish medicine. And there's many, many others, drugs, you know, that have been used.
Some of them, you know, again, you know, fairly available in, small animal practises, you know, as well as in, mixed animal practises, you know. So, the literature will tell you like, you know, a little bit more of those. But these are the two more commons.
What are we looking for in sedation? So look at this patient, you know, this is a koi with about 60 milligrammes per litre of buffer MS 2. So, let's play the video again.
You will see that the fish is still percolating, is maintaining position, you know, in the water, but it is not swimming around the top, you know, with, with a lot of excitement. If we increase the concentration, then we see the fish losing position, still or percolating well, and at this point is where the veterinarian starts making the call as to if it's safe to start manipulating this fish, because at this point it is where I want to start collecting samples, conduct a full physical. I.
Yeah. OK, I hope you were able to hear that, you know, but again, see, I am manipulating the fish. The fish is still percolating, and also at this point, I can also use a Doppler to take a heart rate of my animal.
Clinically, I do not take, you know, again, this heart rate, you know, very frequently, unless my patient is not uperculating. I typically use that operculoppercular rate. A lot more often than the heart rate, you know, to assess, you know, how deep my patient is, .
As I conduct, you know, the different procedures that I'm conducting, but Doppler is also potentially used for heart rate measurement. You can conduct at this point, you know, a full physical evaluation, you know, where you can look for growth changes on the animal, a little bit more in depth, excess mucus production, you know, you can see skin lesions, you know, for example, in this tilapia. They lost the scale.
So by definition, because the scales come from the dermis, by loss of the scales, you have an ulcer, a celomic distension, you can see nodules, you know, in this case, you know, parasites in the, coal tail of this animal. And in this rainbow trout, you know, then you can start seeing, you know, The gas bubble. So all of these are bubbles of air.
So in this case, gas bubbles is in the tail of this animal. So again, as you conduct, you know, in during anaesthesia, you can start looking a little bit more in depth on the different growth changes of these animals while you conduct your physical exam. You can collect measurements as well.
There's different measurements, you know, that can be collected. We typically collect total length. So from the snout all the way to the most caudal point, you know, of the caudal fin.
Girth, basically particularly useful if you're going to do multiple evaluations of the animal. And of course, the weight that allows, you know, that, you know, if we need to do specifical treatments, you know, injection of antimicrobials, you know, or anti-inflammatories, etc. To collect, you know, the weight of this animal, also allows us to monitor if the animal is losing weight or gaining weight as we do multiple evaluations of the same animal.
The skin scrape, OK, I'm gonna pause in here the video for a second and I'm gonna go over what is gonna happen in here. So what's happening in here, this is a method you can collect, you know, from a live animal. I like to sample two sides, you know, in the fish.
One of them is going to be beneath the pectoral fin because there is less dragging of water in there. So some parasites like to hang in there. What I'm gonna collect in here is a sample anterior to posterior, so basically following the same.
The same, Plain as the scales because again, I want to avoid getting scales into my slide because as I mentioned before, if you get scales, then you are generating an ulcer by definition. You want to sample the mucus. What you're sampling in here is just the mucus of the animal, OK?
And another important point, you know, is that because you are evaluating the external surface of this animal, the mucus, what you can also do is collect a small clip of the fin, very important that you talk to your client to make sure that they are OK with you collecting a. Fin clip because sometimes, you know, particularly in ornamentals or in show fish, they will not want you to do this, you know. But in some cases, you know, for example, if dealing with culinary disease, you know, the thin clip, it could be very, very informative.
And at this point, you can evaluate at the same time as you do the, skin scrape. So I would collect, you know, a drop of water from the same container that the fish is in. You do not want to use ethanol, you do not want to use sterile water.
You want to use the water from the same system that fish came in, a little drop of water in the slide, you can put again a small amount of that thin clip and then collect, you know, that mucus sample. So now we're gonna see the video step by step. So again, collecting anterior to posterior, and I can collect two different edges of that cover slip.
And I'm gonna put on top, there you go, you see the fin clip. You wanna make sure that that air pocket is no longer there. So in some cases you have to put a little bit more water.
So that by capillary action, you know, you don't have, again, you know, any pocket of air between your cover slit and your slack. And then you're ready to evaluate this under the microscope. There's few bacteria that can be diagnosed, you know, at this method, but flubacterium colonary.
One of the causative agents of colonna disease could be diagnosed this way because the bacteria is very long, is very thin, but produces these haystacks, you know, this accumulation of bacteria that basically you can very easily see under the microscope. At that point, also, you can collect blood samples again and in here, although, you know, the vast culture of the animals might change the species to species, you know, you can collect, you know, typically from the coral vein, you know, in this diagram from Robert 212, you can see again, you know, the causal vessel. Most people will call it the coal vein.
You really don't know if you're taken from arterial or venous blood vessel because they run together. So, again, it really depends on how critical you want to be of that, you know, and just call it, you know, the caudal vessel. But, in here is the position, so you want to go, ventral.
Usually you want to go beneath the scales first and then in a 45 degree angle, touch the vertebra, because those vessels run ventral to the vertebra, then you will pull out in a little bit the needle, generate a small negative pressure, and then reposition the needle until you see a flash of blood, and then you just want to hold in position. So these blood samples could be used for clinical chemistry, for microbiology, for serology, etc. What is important is that there's missing reference ranges, you know, for a lot of the different species, you know, so again, You be, you need to be aware of what value of diagnosis, you know, you are going to get, you know, from these samples.
Don't do it, just to do it, because again, you know, that interpretation is equally as important. So let's go through the video again for that sample collection. So again, beneath the scale, relocate, you know, The vessel, touch bone, and then generate a negative pressure and collect your sample.
How much blood to collect, you know, from the fish? Typically, 0.5 to 1% of their body weight is what's recommended.
But, of course, if you are dealing with a sick animal, one that you suspect is already anaemic because you see the gills very pale, you know, probably collect, you know, in the lower range. OK? Then the gill biopsy, another very important method.
So let's go over what's happening here. So again, drop of water from the same system the fish came in. We're going to leave the operculum and I'm gonna assess again, of course, the gills, and then I'm gonna collect, you know, from the most apical portion of the gills a small sample.
But this sample has to be, again, in some cases, less is more. Why? Because you don't want to get You know, a lot of these deal samples to the point, you know, that when you put your cover slip, it's not gonna allow to observe individual primary lamella.
You want to see those individual primary lamella lining up in the slide. OK? So let's see the video.
So a drop of water. I lift the opercula, assess the gills, and then from the apical portion, I'm going to collect the sample. In some cases, you know, even that same sample could be used, you know, for submission for microbiological evaluation.
For example, there's a number of viruses that have tropism to the gills in KOI, so coy herpes virus, carpinema virus. So I can use that same sample and submit for microbiological evaluation via molecular diagnosis. So again, individual lamella, put, you know, the cover slip, you know, and that allows me to observe individual primary lamella under the microscope.
So, when we evaluate them, we can see a number of different pathology. In this case, a very high infestation of parasites, you know, I know by the morphology of these parasites, you know, that they're monogeneans likely in the genus Dactylogyrus. But again, you know, I can see a number of bacterial diseases, you know, so for example, epitheliocystis that is caused by a chlamydia-like, infection, or monogeneans, you know, in the gills, as well as other protoso and parasites like, for example, Ioterus Multiphillis, cotia, etc.
Etc. Then we might have to euthanize animals, you know, and again, you know, learning how to euthanize them, you know, is absolutely critical for veterinarians, you know, as well, again, not only because of welfare, you know, but for a good sample collection, OK? Depending on the type of sample that you need.
To collect, you know, a different methods of euthanasia might be needed. OK. There are guidelines by ABMA on fish patients, you know, but again, you know, with over 30,000 species of fish, some of them might work better than others, you know, for some fish species.
For example, one way, one recommended guideline is the. Overdose of Tricaine of MS222. That will work out fine with like zebrafish, you know, it might work out great, you know, for salmonids.
However, for cyprinids like goldfish and koi, it doesn't work as well, and they can still recover even when 5 times the dose used for anaesthesia is used. OK? So, for that.
Koi and the goldfish, you know, some other recommendations is while in deep anaesthesia, they can be used like the brain or basically exanguination. OK? Depending again, you know, of, what type of patient you're dealing with, you know, you will not want to do some of these methods, you know, with the client, for example, having a 2025 year old coy.
In front of them, you know, you know, and, and piece their brain, you know, might be very shocking for them. So, you gotta choose, you know, and there has been a lot of research onto better methods, you know, for euthanasia euthanasia in the icing patient, OK? This is one example again, you know, of a pe thing, you know, of a smaller size fish, you know, with a needle.
But in some cases, you know, even severing, like, you know, the spine, you know, while again, you know, has gone through that very deep anaesthesia might be needing. For example, if you need to sample the brain. If you want to sample the brain for microbiology or for histology, you cannot pick that animal because if not, your diagnostic sample will be gone.
OK. Other methods that have been investigated, you know, for example, you know, in collaboration with Doctor Brost, you know, colleagues, you know, at at Davis, you know, we have been using, basically the coal vessels, you know, to investigate CT contrast, you know, but this same method, you know. Be then used, you know, to, inject, for example, potassium chloride intravascularly during deep anaesthesia of an animal, in this case, in a sturgeon.
And what you're going to hear and see there is basically the heartbeat of the sturgeon in anaesthesia. When we provide, you know, the potassium chloride intravascularly and you will see in the ultrasound, you know, as well as with the Doppler, that heart rate basically disappearing very fast, OK? So you gotta be accurate, you know, but I think this is a great, great method and we have used this, you know, also in Cypriites in Koi, for example, a lot less shocking for the clients, of course, you know, if they're present during this procedure.
OK? So. Yeah Well Yeah I OK, so you say like, you know, a very successful method again, very fast, within seconds, that heart rate is completely gone.
So, again, further research is, of course, needed, you know, but a great, you know, potential for this method. Then again, as I mentioned, you know, veterinarians should be familiarised, you know, with how to perform a full necropsy of these patients. Again, as with other animals, having everything ready prior to initiating the necropsy will be absolutely critical.
At this point, you know, of course, you know, the anatomy of the animals, you know, that you may see might be different. In this case, you know, in here, a lid, the tilapia, where I recommend doing different incisions. One of them, opercular incision to remove the operculum.
The second one is going to be a ventral incision, and try to go anterior to the vent, you know why? Because if you start inserting the scissors right into the annals, you know, then the problem is going to be that you're gonna rupture intestinal tract, you know, and that will be a source of contamination internally. So a lot of times, you know, we need to get aseptic, you know, samples from internal organs, you know, and that will prevent that from happening.
So, again, do it anterior to the vent, generate, you know, that ventral incision. I usually go all the way to the ventral portion of the opercular cavity. Then a lateral incision that is going to be an inverted L all the way to the dorsal portion of that opercular cavity, and then a cranial incision, you know, if I sample the brain.
So what you start seeing, you know, is this window in this case, in the tilapia that allow me, again, not only to recognise, you know, the different organs in situ, but then also to start, you know, collecting samples. Even if I'm not familiar with the species itself that I'm opening. So let's say that I'm opening another.
By familiarising myself with the tilapia, at least, you know, I will have a better idea on what is what, OK? Of course, there's differences interspecies, you know, and different genus of fish, but again, you know, generally, I always do these necrosis with the fish in right lateral recumbency so that I can maintain again this picture of in situ organs, you know, in the fish in my mind. This is an example of what the oi will look like.
So you'll see like, you know, of course, you know, obvious evidence, you know, differences. The posterior kidney is a lot more like, you know, concentrated in one single side. This is a physostomofa, you know, and this is the two lobes of the swim bladder.
The liver and the intestine are a mixed, you know, with that, you know, so a little bit complicated to, really distinguish, you know, one from the other in some cases. The spleen in here, very, very, easily observed, the heart, and then, of course, you know, the gills. In the salmonid, again, you know, different, anatomic features, you know, in here the presence of a pyloric zika.
But again, you know, the spleen, you know, is the observed, you know, again, posterior kidney in the, dorsum of that sillamic cavity, the liver, and the heart. In, food production animals, of course, you know, it's gonna be critical to evaluate also the muscle. So in here, you can do like different incisions, you know, in different I usually sample in the axial, the pacial muscles, so throughout, you know, the dorsum of the, of the animal.
Because, of course, you know, there are some lesions, you know, that can occur, some infections that can occur at the site, and, those are, the edible sites, you know, so at the production level, critical to evaluate the muscle. Then you go into the internal examination again, familiarise yourself, you know, with The organs in situ is very, very important. First thing to collect, you know, will be samples for microbiology, OK?
You want to collect this as soon as possible to avoid contamination. From there on, you know, there's a number of things, you know, that you can do. Again, you can collect samples for histology.
You can do squash mounds of different tissues, you know, or you can see in some cases in cystic parasites, for example. And at the end, removal of the visceral block and then opening that GI tract, you know, where, again, you know, you can see the presence of or evaluate, you know, the presence of not only, you know, intestinal content in order to evaluate if the fish has been eating or not, but also, you know, parasites, you know, that are very, very common in certain scenarios. And then at the end, of course, evaluating the brain or other tissues of interest like the heart, etc.
That you might have. OK? So for that evaluation, again, you know, these are just gross findings that we have had, you know, in different cases.
In this case, a porcupine fish where you can see the amount of fluid in the selloic cavity, so very clear ascites in that same animal. The liver is, highly, highly infested, you know, with different cestodes, you know, of course, you know, the liver is similar as in mammals, involving in the production of different proteins, I, globulin, etc. They are important, you know, to maintain.
You know, hydrostatic pressure, etc. Osmotic pressure, you know, in the animal and preventing of these societies. So if the liver is affected, it's very common to see the presence of these societies.
In, in here in this cichlid, you know this tilapia, you can see primary lymphoid organs and secondary lymphoid organs. So in here the head kidney and the spleen. Remember that fish do not have, or most hemopoiesis occur at the level of the head kidney.
They do not have a bone marrow basically, as for hemoposis, they have, you know, the head kidney, and they do not have lymph nodes, you know, so they have other secondary lymphoid organs such as the spleen and the posterior kidney. So, in this case, you know, it is an infectious disease known as Franciellosis, so it's caused by Francilla orientalis. And what it causes You know is this megaspleno and renomegaly.
So in here, this kidney, presenting this renomegaly, probably like, you know, 50 times its size, and covered with these white nodules, you know, a similar presentation in here with the spleen with the white nodules. Here a very enlarged gallbladder, the heart, and then the liver. Then finally, in this sturgeon.
This disease, you know, is colloquially known as fluid belly because of the amount, of course, of, of fluid is actually caused by the infection of a, of a fungus or hyyy, known as Veronia boriosa, which again, you know, also targets the liver and ends up causing again, you know, this very nasty, nasty, fluid accumulation. Other examples, swim bladder. This is the swim bladder, normal swim bladder of a sturgeon, how it should look like, and then a swim bladder of an infected sturgeon with streptococcus in, in which, you know, there's cause, you know, this aerocystitis, you know, very nasty infection, you know, ulceration of the swim.
Bladder. And again, you know, this is caused by systemic infection of streptococcus in in white sturgeon, in tilapia, we see bilateral exothalmia, in this case, you know, could have been from Aromonas, could have been Francia, could have been streptococcus, you know, so, very common presentation to see. Typically on a systemic bacterial infections, you know, in tilapia.
And in here, in a porcupine fish, similar presentation, so you can see, unilateral exothalmia, the eye is, completely, let's call it Popeye, you know, commonly. But in this case, you know, it wasn't a bacterial infection, but a, parasite infestation. In this case, metaroema.
So a nematode basically causing this infestation, and then, the, the inflammatory process, you know, in the eye causing the exothalmia. OK. Sample collection, as I mentioned to you, might be different, you know, in the icing patient, you know, and, it's important, you know, to organise it.
So, initially, we will sample for external parasitology. I keep putting here mycology as well when we do the skin scrap and guilt clips. Later on, in some cases, you know, we might collect samples for bacteriology, virology, and mycology, for example, if we suspect, you know, bacterial infections, you know, in the skin or in the gills.
However, interpretation of these samples, you know, is challenging. Why? Because these tissues are exposed to a number of microorganisms normally.
So it is very easy if there is any type of Lesion or stress, you know, to organisms that are opportunistic, that might be even part of the normal flora of the animal to go ahead and cause an infection. So, interpretation of those external samples is is difficult. Blood can be collected and again, processed, as I mentioned before, you know, for a number of downstream diagnostic methods.
Internal bacteriology, so now internal organs will be sampled. The most common organs to sample in here will be, again, as I mentioned before, those secondary and primary lymphoid organs in fascia. So the posterior kidney, the spleen, and the head kidney.
But of course, in different presentations, you know, the liver or the heart or the brain might be the best tissue to collect. And then at the end, that internal parasitology evaluating, you know, the, the digestive tract, you know, it's important. So this is the type of lesions, you know, external lesions that might complicate your life, OK?
It is difficult to interpret and without the use, you know, of selective media, might be really hard, you know, to really get, to pinpoint, you know, the theology that is causing this. So in This is a oi, you can see this very large ulcer. You can see again, you know, the advancing lesion, you can see this inflammatory processing here.
So if I have to sample these guys, I will not get a sample from the middle of the lesion. I will go into the advancing lesion. I will flush it, typically with saline or PBS and then I will go ahead and take a sample, typically either underneath the the scales or we do a small incision and then try to collect, you know, from, below that lesion.
OK? And again, you know, if I have selective media, I will go ahead and utilise it. Internally, as I mentioned before, the posterior kidney will be the best sample to start with.
Why? Once, because it is protected retrosomically. So typically the swim bladder will provide some protection.
So if there is some contamination, as you are doing your necropsy, typically the posterior kidney is a little bit a safer place to go. It is a filtering organ. So again, you know, blood flows, you know, through the posterior kid.
Knee and then it is a secondary lymphoid organ. So similar as lymph nodes, there is a lot of antigen presentation occurring at this site. It's an immunological reach site.
So it is, for those reasons, you know, the first, sample or the first tissue that we sample. Spleen is a great one also to collect, you know, it's really too easy to localise, you know, but again, the caveat of the spleen is that it is also easier to contaminate as you are doing those incisions during the necropsy. Brain, I love collecting brain, particularly in small fish or fish that are, autolithic.
I mentioned before, avoid sampling dead animals, you know, but if that's the only material that you have and you suspect of infectious agent, I will collect the brain for sure. Why? Because again, you know, it is protected, of course, you know, the brain blood barrier, you know, provides, you know, some protection, you know, again, you know, the fact, you know, that the brain is, is, somewhat a a hard tissue to contaminate, right?
But there is a number of different etiologies that will target the brain, you know, from viruses, you know, of course, you know, to bacteria. So, in asepticus, you know, and particularly when they start displaying that abnormal swimming behaviour, you know, the brain will be a fantastic place to sample. So, in cases of lactococcosis, for example, or weillosis, streptococcosis, the brain is, is a great place, you know, to, to target.
OK. So in this example, the right, the brain, you know, that has been evaluated in this case was a Chinook salmon that had a flabby virus infection. So you can see growth changes in here in this brain due to fly virus infection.
And in here, tilapia, look at the posterior kidney. This is a tilapia infected with Francilla. So again, you know, it's a dematose, you know, that that that kidney with white nodules.
So you at this point, I will get a sterile, clean, you know, scalpel blade. I will cut through here and then go with the swab and collect, you know, my sample right from the posterior kidney. OK, virology, this is an art, absolutely.
There are right cell lines, you know, that need to be used in the right conditions, you know, for, recovering viruses, you know, particularly again, you know, if you're interested in cell culture. There's a number of cell lines, you know, available, you know, and again, they use different media, etc. So you need to really talk, you know, to, diagnostic lab, you know, that is familiar with these, methods, you know, to be successful with this.
But for sure, there is procedures, you know, for different size of fish, different tissues that might be recommended to go ahead and sample for virology. And then in other methods, you know, of course, you know, histopathology, for parasitology, for electron microscopy, and for molecular diagnosis, they're similar as to what we typically do in other animals. So if you're submitting samples for histopathological analysis, neutral buffer formalin is the most common one.
No more than 1 centimetre thick, so that, a good preservation occurs. What this means is that a small size fish, you know, you typically process holes. So I removed the operculum and size, you know, the, the sloic cavity and then, and then fix hole.
And other diagnostic methods, you know, that have been, of course, been done more and done in veterinary practises, such as, imaging, are of, of course, of great value for a number of different diseases, you know, in ornamentals, you know, for example, in oi, these omic distentions are very common. There's different etiologies for this could be egg bound, you know, could be tumours, like in this case, you know, so you can see the swim bladder. Here, and all of this is a selomic tumour, polycystic kidney disease, etc.
Etc. But even for non-ath associated, you know, issues, for example, you know, sex in the animals. So you can do this with endoscopy or you can do this, you know, with ultrasound in certain animals, you know.
So, again, you know, that distinction between male and female might be important at the level of production as well. And, it can be done, you know, with ultrasonography. The most basic one and typically the cheapest one is radiography.
Where you can, of course, you know, gain a lot of information, you know, we use this one in a lot of times, you know, when we have, again, you know, abnormal buoyancy, you know, either positive or negative buoyancy, particularly to observe, you know, the swim bladder. In this case, in a oy, you can see again, there are two lobes of the swim bladder and the intestinal tract, you know, this is from Nick and Erin's, veterinary clinic Exotic Animal publication from a while back. Ultrasonography, again, you know, a highly valuable.
You can now do this, you know, in field or in the hospital, in this case, in a coI. We are sampling now for the kidney, which sits, you know, between the two lobes of the swim bladder, very clear in here. So again, you know, a swim bladder, swim bladder, and right in here, this is the kidney.
And this is a kidney with polycystic kidney disease. So when you start seeing is a very, very clear amount of of cystic structures, you know, instead of the kidney. Computer tomography, these are my two great colleagues, you know, Doctor Kelsey Brost, you know, and Doctor Brittany Stevens, you know, that they really pioneered this method, you know, in COI.
So basically, utilising methods, you know, to evaluate, you know, by CT using contrast, you know, in, in, in the in the patients. So we have been using A lot of these methods, you know, to evaluate who prior to surgery. So, again, you know, you can use aopametol, you know, as, as a, as contrast iodine in type products, you know, where we inject the animal in the causal vessels and then take CT.
Analysis, you know, at 5 minutes or at 10 minutes, you know, where you can see really nice distribution of the product. And again, get a very, very, good sense of the anatomy of the animal. So, these are, videos, of course, you know, form, you know, with a CT really nicely showing, you know, that internal anatomy and the distribution of this product, you know, of the vascular system in the oi, following, you know, CT analysis.
You can see here the posterior kidney. Cattle vessels and of course the vertebrate etc. Teleoscopy, this is Dr.
Stevens, you know, how we published this work in AGVR a while back, you know, the position of the endoscope. And in this case, you know, I'm evaluating the heart, so you can see, again, you know, through the endoscope, you know, the heartbeat, you know, and again, you know, this allow evaluation of internal organs. Of course, again, you know, familiarising yourself with the anatomy of the animal will be key for all of these methods.
So with that, a lot still to cover, I hope, you know, that it is useful and I hope, you know, that you are encouraged, you know, to get into this beautiful field. There's a number of great organisations where you can get more information. WAFMA, the American Association of Fish Veterinarians, and the American Fisheries Society official section are just some to name them.
And of course, you know, great meetings, you know, to attend worldwide. That can provide, you know, CE options, you know, for veterinarians. So, again, you know, my email is Soto [email protected].
You can search me at UC Davis, you know, if you have any, any, any questions, send me an email, happy to, entertain any questions and I hope to see you in person or hear from you virtually at some point. Take care.