What killed the harbor seals?

My time as a Ph.D. student in the Vega-Thurber lab has come to an end. Completing my dissertation was a very exciting and relieving moment in my life.  To add to this excitement, just a few days after my latest paper from my dissertation was published in the Journal PeerJ. This paper was an extension of the work that I talked about on a previous blog.

 

Celebrating the completion of my P.h.D

Celebrating the completion of my P.h.D

In my last paper, I looked at the bacteria and viruses from a group of seven young harbor seals that died from some unknown brain disease. When veterinarians performed a necropsy (the animal version of an autopsy), it appeared that these animals were infected with viruses. Although, when we used high-throughput sequencing (sequencing of large amounts of DNA or RNA) to identify a virus we did not find any. The lack of viruses in my results was very surprising as well as disheartening. See I wanted my research to revolve around characterizing viruses, but given that, I could not find any viruses this inhibited my grand research plans.

This situation is very common in research; our hypotheses are not always correct and we often get negative results. It’s one of the aspects that makes the work of a scientist challenging, but it also keeps us thinking and generating new ideas.

My next step was to refocus my question and approach. If a virus did not kill this group of animals something else did. Now I had a new question to answer. What killed the harbor seals? To answer this, I examined the genes of the seven harbor seals. I used a sequencing technique that is known as transcriptomics or RNA-sequencing. This technique is a high-throughput sequencing approach looking at all the genes in a sample. In my case, I was examining the genes in each of the seven seals that died from the mysterious brain disease.

I then analyzed the genes of the harbor seals using computer programs, statistics, and biological principles (bioinformatics). To my surprise, all these animals had high levels of genes related to fatty acid metabolism. Unexpected results are also a common occurrence in research. I initially believed that these animals died from a virus, but my results were suggesting that they likely died from a metabolic disease!

What does high levels of fatty acids in the brain signify?  Generally, fatty acids are important for building cells and providing energy to cells. Fatty acid gene activity typically takes place in fat and the liver cells, and large production of these gene types may indicate a metabolic disease. Things that trigger high fatty acid production includes poor nutrition intake or exposure to toxins. So, we think that these animals either did not absorb proper nutrients or encountered a toxin that caused high fatty acid metabolism production in their brains, which may have led to their death.

So now that we know that fatty acid metabolism dysfunction can occur in the brains of marine mammals we can begin to monitor these genes in other marine mammals. We can look for the same fatty acid genes that we found in our study in other animals. This will help us better understand this disease and hopefully prevent the deaths of other animals. Interestingly, when I began this research question, I envisioned the final results of my study very differently. I didn’t expect to examine marine mammal genes and metabolic pathways, but research has a mind of its own and leads us to unexpected and exciting findings.

What does a marine biologist do in a landlocked country? Study elephants…

On my first post I declared that I would write about marine mammals, but things have changed since then and now I hope you also welcome posts about the microbiology of elephants. To make the transition easier I will show you a picture of a baby elephant.

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Baby Asian elephant at the Elephant Breeding Center in Chitwan

Why I am studying elephants?

I was lucky enough to be given an NSF GRIP award. The goal of GRIP is to partner students with federal agencies. A great opportunity for me since one of my goals has been to work at the Smithsonian. I looked for researchers at the Smithsonian that were taking GRIP students and working with viruses. The lab that peaked my interest was at the Smithsonian National Zoo and works on elephant endotheliotrophic herpes virus (EEHV), a herpesvirus that was detected as fatal only in 1995. Since then about eight different types of EEHV have been detected, but EEHV1 is the most pathogenic. This virus is specifically deleterious to young elephants and can progress to death within 1-7 days after symptoms arise. It has mostly been found in Asian elephants, but there have been some accounts of EEHV African elephant infections.

The project

My project sounded pretty amazing. I would go to the Smithsonian to get some molecular biology training and then fly to Nepal, live for 4 weeks in Chitwan National Park, and check elephants for EEHV; thus saving the baby elephants!!! To sample elephants I will take trunk wash samples since its one of the most effective ways to test elephants for this virus. Basically, a veterinarian pours some saline (salt and water) down an elephants trunk and the mahout instructs the elephant to blow it back out into a container.

Reality

After a two week training at the Smithsonian, I packed up my molecular and fieldwork bags and landed in Kathmandu, Nepal. My plan was to collect samples the week I landed. Two and half weeks later and I just collected my first set of trunk wash samples. I declared them the most beautiful thing I’ve seen in Nepal. Not that Nepal is not a beautiful country (e.g. picture below), just that the hurdles to get these trunk washes illuminated my tubes of samples with a magical essence.

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Rhino making its way to the Rapti river in Chitwan

The struggle

In addition to working with the Smithsonian, I also wanted to develop my fieldwork skills. So why not attempt fieldwork on a topic I just learned, on field techniques I am not trained to do, in a country where I don’t speak the language, and where electricity is sparse. Needless to say many problems occurred, from permits, to miscommunication about the support available to me in the field, and socioeconomic problems.

While I have been able to manage many of the problems because of the help of local Chitwan people and the support from NTNC, the socioeconomic aspect of this experience has been the most difficult and thought provoking. I think I used the word “socioeconomic” in almost all the proposals I wrote on this project, but I had a limited understanding on how my project could affect the economy or society in Chitwan’s elephant industry. I thought my research would provide a free healthcare service to the elephants of Nepal and people would be mostly enthusiastic about my study, right? Wrong.

I was advised to work with elephants that are privately owned, which are mostly used for tourism. While I have not spoken with all the owners, I have talked to some of the major members in the committee and the consensus from the ~45 elephant owners has been that my study would be a burden to them both economically and socially. Why do they think this, you ask? Their hesitance comes from a previous study conducted to survey elephants with Tuberculosis (TB). A bacterial disease that infects many mammals including humans and elephants and is transmitted through the air.

I was told that elephants that were diagnosed with TB could no longer partake in tourist activities (to avoid transmitting TB to people) and were not allowed into the forest (to avoid transmitting TB to the wildlife) until the elephants were treated for TB. Now it is seen as an economically poor decision to have scientist look for a pathogen in their elephants, because they risk losing money from tourism and are forced to treat their elephants. These consequences were especially troubling, because besides the diagnosis their animals looked otherwise healthy. Also, a positively diagnosed animal may be seen in the community as inferior and may decrease the animal’s value.

My solution 

The elephant owners protested that if a diagnosis of an illness is made then the scientist must also provide a solution and treatment. However, I can not provide these resources. Instead I tried to provide education about EEHV to elephant owners. For instance, TB is not like EEHV and should not affect the owners economically. EEHV has not been shown to be transmissible to humans or other animals, so tourism should not be affected. If an adult elephant was diagnosed with EEHV they do not need to be treated, but it is important to know that they can transfer the virus to a calf.

Although, mostly (and I mean mostly) all the elephant owners are skeptical about my research there was enough of them that supported my study; hence the picture of my beautiful sample collection.

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Elephant trunk wash samples.

Ever wonder what’s inside of the brain of a harbor seal?

As promised I am here to provide some new information about marine mammals. I know you’ve waited too long. This time I will be telling you about my very own research that was just published in PLOS ONE!!! This is my first, first author publication so there was a steep learning curve on data analysis, graphing, and the publication process. In future post I plan on taking the reader on the journey of my first publication. Especially the struggles, since the public often doesn’t hear about that side of science.

For this blog I will  give a quick synopsis of my paper for those who just really want to know what I found inside the brains of Pacific harbor seals.  Although, before I get to the nitty gritty of my story I like to tell you a bit about my study subjects, the harbor seals. I mostly worked with pups (< 1 month) and weaned (1 -12 months) harbor seals, which tend to be born between February and April. Their mothers wean them for about 3-4 weeks and soon after they begin to catch small fish and shrimp. During these early stages of life, they encounter many dangerous situations for example, being preyed, starvation, and disease.

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Popeye a Pacific harbor seal not related to this study. Photo by me

I am particularly interested in the diseases of young harbor seals, since understanding this may increase their survival rates during this fragile stage of their lives. About 44% of marine mammal diseases remain a mystery. That means that many harbor seals die and we have no idea what’s killing them. So, I wanted to help discover possible culprits that cause marine mammal diseases.

I started my search with the brain tissue of 14 harbor seals that were found sick on the California coast. I first looked into the viral world and I found viruses in four of the animals from the family Herpesviridae, which have previously been found in harbor seal brains in European and North American waters. A virus from this family (Phocine herpesvirus-1,  PhV-1) are particularly harmful for young seals since they have an undeveloped immune system.

However, the bacteria side of the story proved to be a bit more interesting. One of our significant discoveries was the presence of Burkholderia along with a high amount of Burkholderia genes that are known to cause disease. Burkholderia is an interesting genus of bacteria that can be zoonotic (passed from animals to humans), but it is also ubiquitous and can be harmless. While this is not the first time someone has looked for bacteria in the brains of marine mammals and found Burkholderia, it is the first time that this bacteria was found in harbor seals in the USA. So where else were Burkholderia found in the brains of marine mammals? Well in Southeast Asia, this bacteria was found in an aquarium, which caused the death of marine mammals.

Our other interesting finding on bacteria comes from Coxiella burnetiiSimilar to Burkholderia, it was found at a high abundance with high amounts of disease causing genes. Unlike, Burkholderia it was not found in all our harbor seals, but only in three of our animals. Meaning this pathogen may be less common in harbor seal populations compared to Burkholderia. 

C. burnetii is a known pathogen that needs to replicate within a cell (obligate intracellular pathogen). It is best known for causing Q fever, but in marine mammals it causes inflammation of the placenta (placentitis) and it has never been found in the brain of harbor seals. We think since these harbor seals were young animalsthe placenta may be a source of C. burnetii infection for pups, but this is just a hypothesis that needs some testing!

So what exactly did we learn from all this? Well our study adds to our knowledge about the distribution of Burkholderia in marine mammals and like in Southeast Asia it may also be causing the death of harbor seals in the USA. Also, now we know that C.burnetii  can infect the brains of harbor seals and we should investigate the source of this infection. Finally, we can now begin to monitor for these bacteria in the brains of these animals as possible sources of infections.

Will someone please think about the marine mammals?!

Marine mammals get a lot of attention in pop science because of their charismatic nature, but since our lab is mainly focused on coral reefs, marine mammals can sometimes be overlooked!

Hi, my name is Stephanie, and I am the one member in the Vega Thurber lab that has decided to study the microbiology of marine mammals. So, I like to explore the marine mammal side of things. For instance, Ryan is now diving in the Red Sea at KAUST, sampling and assessing coral diversity while surrounded by (but ignoring) frolicking dolphins. Through Ryan’s dolphin watch reports, I became curious of what other marine mammals Ryan may ignore in the Red Sea.

It’s me Stephanie

Doing a little research, it is easy to discover that the Red Sea is home to many marine mammals, but I was mostly surprised that it was home to the dugong, which roams throughout the Indo-West Pacific Ocean. The dugong belongs to the same order as the manatee, but has been the only member in its family, Dugongidae, since the Steller’s sea cow was hunted to extinction in the 18th century. The dugong itself is not listed as an endangered species, but is considered vulnerable. Unlike corals, dugongs have a tendency to swim around, which makes population counts difficult. One new way to solve this problem is by utilizing unmanned aerial vehicles, but this technology is still work in progress.

Dugongs vs. Manatees. Encyclopædia Britannica Online. Retrieved 18 March, 2015, from http://global.britannica.com/EBchecked/media/57538/Features-of-dugongs-and-manatees-compared

In spite of our inability to count all living dugongs, scientist can still use fancy math models to predict the dangers these animals may encounter. The dugong like many marine mammals, including the most endangered marine mammal (the vaquita, a porpoise), is threatened by overfishing. Initially this sounded a bit counterintuitive to me since dugongs are mainly herbivores, but will snack on the occasional jellyfish or some delicious shellfish. Instead, overfishing affects dugongs because it leads to the destruction of seagrass beds, which is where dugongs like to swim and eat. Dugongs are not alone in the plight that is overfishing. Overfishing causes ecological, social, and economic problems. One way to help this problem is by purchasing sustainable seafood, which is made easier by using an app by Seafood Watch.

Since I am a microbiologist I would like to end this post with some microbiology. Strangely, there are few studies that investigate the microbiome of many marine mammals, but it turns out that there is a study on the dugong gut! I know very exciting! Gut microbiomes can be studied by examining fresh feces, thus in this study scientist collected feces from wild and captive dugongs and extracted the DNA. Using DGGE techniques, they concluded that captive and wild dugongs have different bacteria communities. With captive dugongs having fewer bacteria types, which can be considered unhealthy.

I hope you guys enjoyed this marine mammal post and expect some more in the future.