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Small Samples, Big Science

Microscopes, blood analyzers, and Petri dishes are among the National Zoo’s front line of animal-care practices. Tools like these allow the Zoo’s pathology department to diagnose illnesses with little more than a drop of blood or a fecal sample.

By Cristina Santiestevan

Origami kangaroos and paper elephant mobiles hang from the ceiling. Chemical-resistant countertops and deep laboratory sinks are reminiscent of high school chemistry class. Three stainless steel refrigerators bear matching signs: “No Food.” Above a doorway hangs another sign: “From poop to blood: We do it all.”

Pathology lab sign: Froom poop to blood, we do it all.

This sign adorns the Zoo's pathologoy lab. (Mehgan Murphy/NZP)

The sign above the door is absolutely correct. The people who work here, in the Zoo’s pathology lab, are essential members of the animal-care team, although they rarely leave the lab or see an animal. Instead, they interact with the Zoo’s elephants, clouded leopards, and golden lion tamarins through blood, urine, and stool samples. They gauge an animal’s health from the laboratory, by counting blood cells, testing stool samples for pathogenic bacteria, and running countless other tests.

“It’s a lot of fun, because you learn all the time,” says Ann Bratthauer, a 27-year veteran of the Zoo’s pathology department. Bratthauer, Betty Ackerman, and Stephanie Otto are the lab’s three technologists. They handle samples from Rock Creek, Front Royal, and Smithsonian field researchers around the globe, and they conduct tests in the clinical pathology lab. The variety of patients is both a challenge and a delight. Some animals are still relatively unknown to science, and every blood test might bring a new discovery. “Everything we do is research,” says Bratthauer.

What’s Normal?

Almost every animal at the Zoo will have a sample analyzed in this lab. In total, approximately 5,000 individual cases will be handled by the clinical pathology lab this year. Each will be summarized on a detailed data sheet and filed for future reference. Data sheet number 2010-3599, for example, summarizes routine tests that were recently completed for a female Gila monster. Her test results reveal she’s in fine health.

These standard checks are a routine but important part of the monitoring animal health at the National Zoo. Parasite loads are tested at least once a year, and sometimes as often as once a month. Water samples are routinely checked for harmful bacteria. Blood and urine samples allow Zoo staff to track kidney and liver function. If a test comes back abnormal, the veterinary staff are alerted, and the animal receives medical care.

Most important, however, may be the countless tests that come back as normal. For many animals, researchers are just beginning to understand what typical really looks like. Healthy human blood cells, for example, should be round. Healthy camel blood cells, in contrast, are oval. Without knowing this, a researcher might incorrectly assume that a healthy camel is sick, all based on those oval blood cells.

The differences between animals extend beyond blood cell shape and include practically every measurement that might determine an animal’s health, such as blood cell size and count, proportions of red and white blood cells, and hemoglobin content. Each of these parameters in mammals can be measured using the Hemavet, an automated blood analyzer. But the automated results will mean little, unless the team has already determined what typical looks like for that particular species.

Snake

Puja Basu and Esther Langan trim preserved tissues. Mehgan Murphy/NZP)

“People doctors have no clue how hard it can be,” says Bratthauer, who clearly does not mind the challenge. “They have only one species to work with.” The sheer number of species is not the only challenge here. There is also the fact that the patients are all wild animals, and many species and individuals are not exactly cooperative with their caregivers.
For example, those clean urine sample cups that human doctors pass out to patients for every routine physical simply don’t work here. Instead, urine samples must be collected in whatever way possible, even if it means getting them off the floor. This means the lab technologists must be able to tell the difference between contamination in a urine sample, and an actual symptom. Pollen grains, for example, can look remarkably like parasites when suspended in a drop of urine.

Pollen-contaminated urine samples and oval-shaped blood cells are simply part of the equation for a zoo’s clinical pathology lab and the people who work there. “It doesn’t matter if it’s blood or feces or what have you,” says Bratthauer. “It’s our job to do it right.”

Learning From the Dead

The laboratory is only one part of the pathology facility. A short walk from the lab is a separate building that is the ultimate destination for every animal at the Zoo. This is where Tim Walsh, the Zoo’s head pathologist and self-described animal coroner, conducts necropsies—animal autopsies—on deceased animals.

“Every animal we have will eventually die,” says Walsh. Even though zoo animals tend to live longer than their wild counterparts, none of them will live forever. When they die, it is Walsh’s responsibility to determine why.

Whether it is a much-loved Zoo resident or an anonymous wild sparrow, every animal that is discovered dead on the Zoo’s grounds is given the same treatment: a thorough necropsy and, often, a microscopic investigation of the animal’s tissues and organs. This attention to detail is a necessity in a world where wild animals can transmit their diseases, parasites, and infections to the Zoo’s residents. Determining the cause of death for one animal may save the lives of countless more.

Every necropsy shares the same set of goals. First, the necropsy team attempts to learn why an animal died. In particular, the team looks for evidence of disease that might pose a threat to other Zoo animals, or might indicate a larger trend in an animal group or species. If the animal died of normal causes, the necropsy helps develop a better understanding of what normal looks like for that particular species. Finally, the team collects and saves samples that may be used in research projects, for educational programs, or as museum exhibits.

A complete necropsy also includes histopathology—the investigation of tissue or organ samples through a microscope. Some necropsies may also include tests for parasites, toxins, viruses, and more. “Most of our answers come from looking through a microscope,” says Walsh.

Thousands of microscopic samples can be preserved from a single two-millimeter slice of tissue or organ. Each sample may hold a clue that will ultimately further our understanding of these animals, leading to advancements in their care in zoos, and their conservation in the wild. “We try,” says Walsh, “to learn all we can from the animals, since many of them are so rare.”

Gaboon viper eye

Pathology lab slides (Mehgan Murphy/NZP)

Disease Discoveries

Samples collected from National Zoo necropsies support research and conservation projects around the world. “That’s a whole research project,” says Walsh, pointing at a container that holds the heart of a deceased male gorilla. Heart problems are not humans’ alone; in fact, they are the leading health problem with male gorillas in captivity and a key focus for chief veterinarian Suzan Murray. No one yet knows why. But studying preserved hearts may eventually help a researcher discover the cause for heart problems in male gorillas. Knowing the cause is the first step to developing a treatment.

“Any researcher can ask for samples,” explains Walsh. For example, the Zoo recently shipped several primate samples to a researcher who is investigating the prevalence of alcoholism—once thought to afflict only humans—in various primates that eat fermented berries. The results might help treat addiction in primates.

Over the years, the Zoo’s pathology department has contributed directly to several important discoveries. For example, when Kumari the elephant died in 1995, the Zoo’s pathology department diagnosed herpesvirus as the cause of death. This was the first documented case of elephant herpes, and it led to the discovery that herpes is a leading killer of elephants, both in zoos and in the wild. Today, the National Zoo is the leader in a worldwide effort to understand the cause and treatment of elephant herpes.

Additional discoveries supported by the Zoo’s pathologists include callitrichid hepatitis (a fatal disease transmitted by mice) in golden lion tamarins and chytrid fungus (a deadly skin fungus that has devastated amphibian populations around the world) in frogs and toads.

These discoveries guide animal-care practices around the world. They also provide essential insight for conserving wild populations of animals in their native habitat. If zoo pathologists are able to develop a successful treatment for elephant herpes by working with captive animals, their findings may eventually contribute to decisions and actions relating to wild elephant conservation.

Likewise, by studying chytrid fungus in zoo amphibians, pathologists advance our knowledge of the condition, and may ultimately help reverse the downward trend of most amphibian populations around the world.

“These animals are ambassadors for conservation and science,” says Walsh of the Zoo’s many residents. That means great benefits come from translating research and knowledge gleaned at the Zoo into helping the species in the wild. “We are not just trying to save an individual,” says Walsh. “We are trying to save a species.”

 

Freelance writer-photographer CRISTINA SANTIESTEVAN wrote about clouded leopards for the May-June 2009 issue of Smithsonian Zoogoer.

 

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Smithsonian Zoogoer 39(6) 2010. Copyright 2010 Friends of the National Zoo. All rights reserved.