Annie PAGE_KARJIAN 03/29/17 Marine WIldllife Rehabilitation

Thank you Megan for the introduction and thank
you guys, all for joining me this afternoon just to talk a little bit about wildlife rehabilitation,
which is one of my favorite topics. So I’m gonna start the talk this afternoon, talking
about just, like, an overview of wildlife rehabilitation, some of the principles and
goals underlying wildlife rehabilitation efforts and then I’m gonna tell you a little bit about
my research. Actually a good portion of my research is based on rehabilitating animals,
especially sea turtles, and so I’m gonna tell you about that work. So some definitions for
you: Rehabilitation means to restore to customary activity through therapy and education and
that usually includes treatment, which means to give medical aid to. One of the main concepts
of rehabilitation is that the focus is on the individual animal or human rather than
the population at large. There’s a lot of underlying ethics surrounding wildlife rehabilitation
that I think are important to consider. When we look at animals and animal welfare
and animal treatment, animal medicine, a lotta times it comes down to decisions that are
made by the animal’s owners and with our pets we know who the owners are, farmers we know
who the owners are but who owns wildlife? So legally wildlife is owned by all of us.
It’s owned by the public trust and that’s in the United States so we all have a stake
in animal welfare when it comes from wildlife and I think most people would agree that we
have a moral imperative to humanely care for a wild animal in distress as their owners
and especially when the distress is due to human activities. So I’ll just give you some
examples. Of some horrible things that have happened to the wildlife, this is a harbor
seal who has outgrown his packing strap entanglement. This is an oiled sea turtle. This is from the Deep Water Horizon oil spill
in the Gulf. Here is a Florida manatee who succumbed to respiratory distress because
of harmful algal blooms that are ultimately due to human activities. This is a sea turtle
in rehabilitation at the Houston Zoo who swallowed some fishing hooks. Here’s a California sea
lion who some nasty person shot with a bow-and-arrow probably because either for fun or because
the animal was raiding his fishing lines or nets. This is an American bald eagle that
has lead toxicosis from scavenging on an animal carcass that was killed with lead bullets
and this is a patient that was at the International Bird Rescue Center in California. This pelican they think a fisherman probably
– he was probably – again raiding the line or the net and the fisherman got mad,
caught up the bird and cut his pouch with a knife, which is a horrible thing to do but
actually this is a good story and this one, too. These animals were rehabilitated and
released so just some examples. So when we talk about the principles of wildlife rehab,
programs to rescue and rehabilitate sick animals are founded upon two general principles: Altruistic
assistance to the sick or injured and conservation of wild populations and, in the process, we
gain medical and scientific knowledge and provide a valuable interface wild-animal welfare
and the public. The ultimate goal of wildlife rehabilitation is a successful transition
and return of an individual animal back to its natural habitat in a fit and healthy state
to reproduce and perform all natural functions and behaviors benefiting the environment and
the species. So I think that’s important to remember that the outcome is release. Conservation:
So the Second Principle of Wildlife Rehab is Conservation, as I just mentioned. Conservation
is a realistic motive for reintroduction programs for threatened species so, yes, reintroduction
programs for threatened species such as the rhino. For robust populations however conservation
is not a defensible argument for rehabilitating animals because reintroducing a few or even
a few hundred of these animals will be of little benefit to most stocks and even less
benefit to the population as a whole. So when we talk about rehab, a lot of people wanna
focus on the good things but there are also a lot of downsides to rehab that a lot of
biologists will argue. First of all it’s extremely expensive. There are several recent studies
indicating that rehabilitation of a single animal can cost over $100,000.00, depending
on the species and the duration of rehabilitation, and this cost may be justified for endangered
species who with populations that are imperiled, however the cost is very difficult to justify
for non-threatened animals. Also many biologists would argue that rehab facilities divert money
away from programs based on wild population conservation and habitat protection. Rehabilitation success rates vary by species
but they tend to be quite low. For example the sea-turtle rehabilitation success rate
in Florida between 1986 and 2004 was 36.8 percent and when we look at – when we say
success that means the animal was released. In another study in California 14 percent
of 35 rehabilitated dolphins over a 25-year period were released so generally low success
rates. There’s few long-term, follow-up data on rehabilitating animals. The data is logistically
difficult to obtain and maintain. It’s expensive and it’s difficult to track animals after
you’ve released them. That’s the bottom line. Some people argue that wildlife rehabilitation
has no benefit to the population unless the animal reaches sexual maturity and reproduces.
Minimally the animal should have an annual survival expectation equal to the age class
of that population and anything less than that is arguably a postponement of the animal’s
fate by the number of days that it was in a rehabilitation facility. Another concept
is that rehabilitated animals may be returned back to health but it doesn’t necessarily
mean that they’re releasable. The animal has – in order to be released a wild animal
has to be able to form all of its natural functions, including movement, maintaining
stable body weight by normal foraging behaviors, have normal blood health, have no indication
of infectious diseases and not be habituated to or dependent upon humans for survival.
So there’s some pretty tall criteria and the fact is that a lot of rehabilitated wildlife
species end up as permanent captives. There’s also a risk of pathogen spread so when you
bring an animal into captivity, there’s a risk to the humans who are interfacing with
that animal, being exposed to zoonotic diseases, and there’s also a risk when you reintroduce
that animal into their wild population that they may have picked up some diseases during
captivity that they would then take back to their population so those are the downsides. There are a couple other ethical dilemmas
that underlie wildlife rehabilitation. There’s a lot of research that’s been done on these
concepts so the main question is that “Do we invariably act in the best of the animal
or are our best interests or our Number 1 focus is on, like, our altruism and that it
makes us feel good to rehabilitate the animals?” Also the question is “Is it humane to rehabilitate
a non-releasable wild animal?” And also “How can we ensure a released animal will survive
in the wild or contribute to the population?” Like I said you need long-term follow-up data
that can be expensive to do and difficult to maintain. There’s also a rehab bias and I just thought
this is worth mentioning. We tend to focus on species that we have a special affinity
towards, there’s this cute-and-cuddly bias. We like adorable animals and who doesn’t?
There’s also a flagship species bias where we tend to gravitate towards charismatic megafauna
and kind of ignore the less charismatic species like snakes and things like that that people
may not relate to as easily. So the truth about wildlife is that humane, rapid euthanasia
is often the best option for injured or sick wildlife. Euthanasia stems from the Greek word euthanatos,
which means “good or easy death”, and the key to ensuring the welfare of individual
wildlife casualties is a proven process of triage that enables rapid euthanasia of animals
that are considered unlikely to be released back into the wild. Success in this sense
may be judged in terms of preventing unnecessary suffering whereby a high level of euthanasia
could be considered successful. Once a decision is made to bring the animal into rehabilitation,
success is then defined as bringing an animal back to health and releasing it into its natural
environment and benefiting from this process, in terms of public education and scientific
knowledge. But, either way, the triage process should be done quickly, animal-side and the
decision should be made quickly if you are going to euthanize. So again the key to successful
rehab is a well-defined triage criteria, which is supported by research, effective, appropriate
intake criteria, which is often species-specific. You need highly-trained personnel and lots
of infrastructure including facilities, funds and flexibility and usually a lot of those
things. So those are just some of the overall concepts
about rehab and now I’m gonna talk about my favorite part about rehab, which is access.
So an important benefit of animals cared for in rehabilitation is their role as sentinels
of ecosystem health because rehabilitating wild animals are so accessible, they can help
contribute to the developing reservoir of information on care and medicine that we can
then apply to free-ranging wildlife needing help. And this health knowledge is crucial
for wildlife conservation and also veterinarians gain skills and knowledge from rehabilitating
animals that enable them to play a vital role in conservation and also, in turn, to rehabilitate
animals in the future better. So some of the things that we learn from access to rehabilitating
wild animals is information on physiology, life history and nutrition, behavior, diseases,
toxicology and medicine, including treatment and diagnostics. Wildlife rehabilitation centers
can be a valuable resource for the surveillance and monitoring of pathogen activity and wildlife,
which, in free-ranging animals can be cost-prohibitive and logistically challenging. So although
there are some inherent biases in this approach, obviously, it can still provide valuable information
to assist in management and conservation decision-making, including approximations of disease prevalence
in the general population, species variation in disease susceptibility, clinical syndromes,
standardization of diagnostic protocols, development of medical approaches, therapeutics and outcomes
associated with various pathogens. And we can use this information to make these population-wide
inferences that we can apply to general wildlife populations and that we can also then, in
turn, apply back to rehabilitating wildlife to inform and improve future efforts. All
right so that’s just my introduction and now I’m gonna tell you a little bit about my research.
For the past seven years I’ve been working on figuring out this disease that affects
sea turtles, called fibropapillomatosis or FP and since you live here in Florida you
may be familiar with it. It’s a debilitating, infectious, neoplastic disease that affects
all sea-turtle species but has reached epizootic status in green turtles, meaning it’s an epidemic
in many green sea-turtle populations, across-the-world. It’s been reported in all oceans in many countries.
It tends to mainly affect juvenile green turtles after they migrate back to their coastal habitats
so usually around four to seven years old. It can affect any — they can have tumors
on any part of their body, including internal organs and it can be fatal in some cases.
There’s a virus associated with these tumors called Chelonid herpesvirus 5. It’s the presumed
etiologic agent of FP. Here’s the virus right here. I say presumed because like many herpesviruses
this virus is not able to be isolated in cell culture, however the tumors have been proven
to be transmissible via cell-free tumor extract, meaning that a researcher cut off a tumor,
ground it up, passed it through a virus-sized filter, injected it into naïve turtles and
they grew tumors. This virus is consistently associated with FP tumors via multiple molecular
techniques. The herpesviruses group into alpha, beta and gamma herpesvirinae and this one
is here. It groups with the alphaherpesviruses. Phylogenetic studies show that the viruses
coevolved with its sea-turtle hosts meaning that the recent epizootic or epidemic is probably
not related to the virus but probably something related more to the host, which is the turtle,
or its environment. Other factors that have been associated with
FP most significantly is the nearshore shallow-water embayments so they tend to get tumors when
they move from offshore to inshore. It’s also been associated with pollution, eutophication,
harmful algal blooms and invasive forage species, like macroaglae. It’s been associated with
warm-water temperatures, which is not that surprising because a lot of herpesviruses
are temperature-sensitive and also with host immunosuppression. So the reason that I’m
interested in FP is because when we talk about emerging infectious diseases these days generally
there’s this, like, a triad that involves the pathogen, the host and the environment
so usually there’s a bit of environmental degradation or contamination or both. There’s
also host immunosuppression and then you have a pathogen and when these three things meet,
we see diseases that may not have been a problem 100 years ago. Again you have environmental contamination
or degradation, including exposure to this type of environment. Then the host is immunosuppressed
probably related to the environment so they have decreased cell counts, decreased cell
function and then they have an increased susceptibility to pathogens that normally wouldn’t affect
them in that way and, in some cases, you get neoplasia, which is cancer. So that’s what
we’re seeing with this disease and you can see that, again, it fits in really nicely
with this paradigm so we have the host, which is immunosuppressed probably due to a degraded
environment and then we have a pathogen, which historically didn’t cause disease and now
it does. In rehabilitating turtles FP is a huge problem. It’s infectious. It’s horrific
for an individual animal. It tends to necessitate special husbandry conditions, including quarantine,
like stringent quarantine. The tumors can influence clinical progression,
extend rehabilitation duration and complicate prognoses and here are some examples. This
turtle came in. I think she was hit by a boat. This is at the Georgia Sea Turtle Center and
she was in rehab for over a year and she was doing fine, getting ready to be released.
She was quite robust and then the staff noticed a bulge in her plastron. They did an MRI and
it turns out she had a huge internal tumor and she was euthanized. This is another turtle who came in with a
boat-strike wound on the carapace and was progressing quite nicely and then developed
these tumors within the boat-strike lesion and the turtle was euthanized. So I got interested
in around 2010 during an externship. When I was in vet school at the Georgia Sea Turtle
Center I had some patients. Some of these turtles I just showed you were some of my
patients and I became really interested in this disease and so I started out my journey
of this research doing a retrospective case-series analysis of all of the turtles that had or
developed FP tumors at the Georgia Sea Turtle Center since it opened. So that’s over a five-year
period and we looked at the occurrence, clinical presentation and outcomes of disease. There
were 25 turtles overall, which was – represented 12.6 percent of all the green turtle patients
at the Georgia Sea Turtle Center, which is not insignificant. In sea turtles we age them
by looking at their size. Generally it’s called the straight carapace length so the bigger
they are, the older they are and you can see that it’s pretty consistent that the juvenile
turtles, these smaller animals are the ones who are most affected by the disease. In the study we also a modified tumor scoring
system, based on tumor number, size, location and severity so, like, how aggressive the
tumors are and the tumor-scoring system ranges from zero, which is unaffected, 1 mildly,
2 moderately and 3 severely affected . We found that 48 percent of these turtles develop
tumors over one week after entering rehab and that ten of them did so during the warmer
months so again there’s a temperature component to this, to the tumor outbreak and also probably
– so they developed tumors after they entered rehab so they were either infected when they
entered rehab or they became infected in rehab and what I think is most likely is that they
were already infected and the stress of whatever brought them into rehab and the stress of
being in captivity led to them developing tumors. FP tumors were most often first observed
in the inguinal regions here so we can advise rehabilitators to focus on these areas, looking
for new lesions and we came up with this estimation that overall about 10 percent of rehabilitating
green turtles may develop FP within 90 to 120 days of rehab, which hadn’t been described
before. We found that ocular tumors, these eye tumors, were observed in about two fifths
of the tumor turtles and turtles with ocular tumors were over 8 times less likely to survive
rehab than tumor turtles without ocular tumors so basically having tumors in your eyes is
bad [laughs] and we also found that 10 out of 25 of the turtles had these – only had
these – plaques, these flat plaques, which histologically and molecularly were consistent
with FP but these turtles were significantly more likely to survive rehab and over 50 percent
of them had spontaneous tumor regression whereas none of the other turtles that had the other
types of tumors had spontaneous tumor regression. So if you have flat, plaque-like tumors it’s
good. Laser surgery is the treatment of choice for tumor removal and we found in this study
that the number of surgeries is not significantly related to case outcomes so a turtle could
have up to 6 tumor-removal surgeries and it wouldn’t necessarily affect the outcome of
the case. So here’s a good example. This turtle had
severe FP. This is a patient at the Marathon Sea Turtle Hospital. You can see she was blind.
This is a pretty nasty case so here she is having her tumor-removal surgery and then
this is the same animal. So just because they look horrible doesn’t
necessarily mean that they’re gonna have a bad outcome. I think that’s important to note.
However we also found in that study that 38.5 percent of turtles that had their tumors removed
experienced tumor regrowth within 36 days. So here’s a turtle. These are the little FP
lesions just developing in the inguinal regions. This is the day that they were removed right
after surgery and then 15 days later you can see the tumors are back and they’re even worse
than before and this fits in with what we know about herpes biology, right? So herpes
is for life as we all know [laughs] and just because you remove the tumors you’re not removing
the virus and so that’s why it’s so important to bring the health of the turtle up as well
as removing the tumors because the turtles, their immune system has to be able to combat
the virus as well. Because of this study – this little study
– we came up with a lot of recommendations for rehabilitators, including – for green
turtles – trying to release them within 90 days, if at all possible, to avoid tumor
development in captivity because once they develop tumors then you’re in for the long
haul. The animal has to be able to survive in its current condition, like I told you
guys before. But we also came up with the recommendation that mild, stable, cutaneous
FP that’s not progressing and the animal’s otherwise healthy, eating on its own, healthy
blood values, they can still be candidates for release and this wasn’t recommended before
but because of what we know about herpesvirus we know that they’re not gonna get rid of
that disease and the most important thing is that the turtle overall is healthy so not
– just because they have tumors doesn’t necessarily mean they can’t be released. We
recommend that they’re released where they’re found to avoid introduction of a new disease
into a naïve population, water temperature over 18 degrees preferred and, if possible,
it’s important to track the patients just like I told you before. We did this same thing
for loggerhead sea turtles in the southeastern United States. I looked at 3 rehab facilitates. Just basically, briefly, we found that FP
is not really a problem for loggerheads. They do get tumors but they’re typically incidental
or mild. They are usually not that aggressive. Interestingly they get these oral tumors that
green turtles, in this area, don’t really get. They get a lot of ocular tumors but they’re
usually mild and exposure to green turtles is a risk factor for loggerheads in captivity.
So we recommend that green turtles are kept quarantined away from all other species and
then green turtles with FP are kept in an even higher level of quarantine away from
all other green turtles as well. However we did find in this study that for individual
loggerheads, FP can be a huge problem just kind of like how diseases in people could
be. So we had this one patient who had severe, severe FP. She was really sick. This is from her intestinal
tract. She was euthanized and – but she – we actually learned a lot about this turtle
and there – so far there have been three studies and I’m involved in a fourth just
on this one turtle alone so she provided us with a lot of information even though she
died. So ongoing research: What I’m doing now to follow up with this is I’m doing it
again. I’m doing a case series analysis but on a much bigger scale and including all species,
not just green sea turtles, involving the Marathon Turtle Hospital, the Georgia Sea
Turtle Center, the Whitney Lab, the Brevard Zoo and the Clearwater Marine Aquarium, hopefully.
And we think we’ll have hundreds of cases in this study so hopefully we’ll get a lot
more of this type of information that can then be passed on to rehabilitators. And the
questions that we’re asking are “Does FP Tumor Score accurately predict rehabilitation outcome?
What percent of surgically-removed tumors regrow? In what timeframe do the regrowths
occur?” and “Is there a seasonality?”, so again looking at temperature. So changing
gears I did a follow-up study of rehabilitating green sea turtles using molecular biology
to try to get a better idea of the herpesvirus itself. For this study I developed validated
and optimized PCR assay. PCR stands for Polymerase Chain Reaction.
It’s just a way for us to detect and quantify pathogen DNA. In this method we can provide
absolute quantification of viral DNA. The assay is extremely specific. It’s very sensitive
and it’s repeatable and robust. And using this assay I evaluated the relationship between
Chelonid herpesvirus DNA loads and clinical disease in rehabilitating turtles. So we looked
at 351 samples taken from 67 turtles, 23 with FP tumors and 44 without FP tumors. We analyzed their blood, their plasma, cloacal
and oral epithelial swabs, feces, urine and skin biopsies and this is the data that we
got, we found. So red is turtles with FP and blue is turtles without FP and then these
are the sample types where we got some results. If the sample types aren’t mentioned that’s
because none of them were positive. Interestingly we found viral DNA in urine samples, which
has never been described before. We found viral DNA in blood, which has never been described
before for this disease and cloacal swabs, again, a novel finding. And we found that
in a lot of skin biopsies so, which is something that we had found with previous studies but
it just supported the data. And you can see that turtles with FP had a lot more samples
that were positive but it was pretty interesting that we had these asymptomatic animals that
also had positive samples so basically telling us we have asymptomatic infections in some
green turtles. When we look at the level of agreement between the different sample types
taken from an individual turtle I thought it was really interesting that there was 100-percent
agreement between blood and urine taken from the same turtle. We know that urine is a filtrate
of the blood so this points to the potential that urine is a method of transmission of
this virus. The fact that we found it in normal skin in
this and another study of free-ranging turtles points to normal skin being a potential route
of transmission as well. And from non-tumored animals as well as from tumored animals. I
followed up this study by taking extensive samples from 10 green sea turtles at necropsy,
5 of them had tumors. Five did not. We took samples of all of their major organs, their
neurological tissues, including optic nerves, nerves innervating their front flippers and
nerves innervating their hind flippers, skin, tumors, blood and urine. These are some of
the animals just to give you a picture and this is a tumor that was on the kidney of
one of the animals. This is the tumor on cut surface. You could
see the kidney was still trying to make urine and we took samples of this fluid and did
PCR on that as well for viral DNA. Interesting results: We found again the urine samples
were positive for viral DNA and then this was really surprising or to me it was surprising
that a lot of the kidney samples were positive for viral DNA and also some of the nerve samples
were positive for viral DNA, which has never been described before. We know that alphaherpesviruses
tend to go latent within nerve cells and so this pointed to a potential site of latency,
which has not been described for this virus. So there’s a lot of stuff that we don’t know
about this virus and I think by understanding the basic biology of the virus then we can
understand more about the disease process. So this study gave me more questions than
answers but basically made me wonder if the kidneys are a site of persistent viral infection
probably related to the presence of the viral DNA in urine, pointing to a renal urinary
route of viral excretion. Also the presence of the viral DNA in nerves points to again
potentially a site of latency of this virus, which is typical of this type of virus. And
the sites where we found virus-positive nerves were nerve cells that innervated parts of
the body that had tumors so we found an ocular nerve, an optic nerve that was positive for
the virus in a turtle that had a tumor in that eye. We found brachial plexus nerves that were
positive for the virus in turtles that had tumors in those flippers and so it supports
that idea. So when we put all this information together we can kind of get a sense of this
Chelonid herpesvirus infection cycle whereby the virus – this is our little virus – enters
the skin, enters the host. We’re not sure of all the transmission routes at this point.
The virus undergoes primary replication and then disseminates throughout the body via
blood and/or nerves probably to the kidneys and potentially other sites of persistent
secondary infection. The virus probably establishes latency maybe in nerves, maybe in other things. The virus is then shed. We know the virus
is shed through the skin and through tumors and maybe through the urine as well. And then
to follow up on this research now I’m doing a study with the University of Florida and
the Brevard Zoo where we’re looking at abnormal kidney samples from green sea turtles so you
can see this top turtle, this is a normal kidney sample here and then this is in situ
or in the turtle. And then this is an abnormal kidney we have generalized severe fibrosis
tract through medullary regions and this lesion is extremely common in green – in juvenile
green – turtles in Florida to the point where it has been called normal because it’s
so common and we’re wondering if maybe it’s due to herpesvirus infection. And so for this
study what we’re doing is again molecular biology paired with histology to see if maybe
this – these types of lesions – test positive for the Chelonid herpesvirus and maybe that’s
why we’re seeing those lesions even though these turtles don’t have tumors. And throughout
this research we’ve found a lot of subclinical disease carriers so you guys may not realize
it but subclinical disease carriers are really important within the epidemiology of a disease.
So the fact that we have animals that are infected but asymptomatic points to an environmental
and/or host factor that plays a role in tumor development, like I’ve said before. It also
suggests that this virus is endemic in some populations. These animals serve as a reservoir
so you can see in this diagram these are the subclinical infected animals. They serve as a reservoir that can lead to
infections in susceptible animals just as clinical animals can serve as a source of
infection and so they can be a source of viral dissemination within a population and this,
actually, probably explains why there was one study where they looked at the exposure
– viral exposure – of this virus in green turtles in Florida and found really high percentages
of exposed turtles in populations that had zero tumor prevalence. So how did they become
infected if none of the turtles have tumors? So this is a likely explanation for that.
To follow up on that study right now I’m doing a study on loggerhead and green sea turtles
in North Carolina. Historically FP has not been a problem for sea turtles in North Carolina,
however it’s becoming more and more prevalent, either because the water is getting warmer
or the sea turtles are transmitting it northward or both. We’re not sure but we’re just doing a survey
of free-ranging turtles, working with NOAA to get samples from them and also rehabilitating
turtles at the Karen Beasley Rehab Facility to see if we can identify the prevalence of
this virus in asymptomatic turtles or subclinical turtles. I’m doing another study – so in
all these pictures you may have realized or you may have noticed that the tumors can take
lots of different types of appearances. They can be these flat plaques. They can be sessile
and smooth. They can papillomatous. This is the most common presentation in Florida
or they can be polypoid and smooth like grapes and, in another study, they noticed that there
are four different variants of this virus in Florida. There’s A – where’s B – B,
C and D. These have only been reported in loggerheads and A, B, and C have been reported
in greens and so it got me to wondering are these viral variants related to tumor morphology
so is the – does the – variant identity predict what the tumor looks like and therefore
its virulence within the host? So we’re doing a study now with the Marathon Sea Turtle Hospital
where we’re looking at using molecular biology to look at virus variant identity in loggerheads
and greens, again, and statistically evaluating those relationships between morphology or
tumor morphology, species and variant and looking at both free-ranging and rehabilitating
turtles and comparing the results. So just to summarize, overall, I think I tried to
make it clear that wildlife rehabilitation has both benefits and drawbacks. It can have
conservation benefits in cases of threatened or endangered populations but the most important
concept to me is that access to wildlife provides an excellent avenue for disease research and
education. And I just wanna thank all of my collaborators. I work with a lot of people, certainly could
not have done any of this stuff alone, including the rehab facilities and University of Georgia
and Harbor Branch and I’ll take any questions and also this is my email address if you have
other questions or wanna email me, have ideas you wanna share, please feel free. So thank
you very much.

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