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Table of Contents:
Keywords: Roseate tern, Sterna dougallii, microsatellite marker, population genetics, sex ratio, conservation biology |
 Roseates at Bird Island |
DNA technology is a topic that is regularly covered by the media. Molecular techniques are at the center of the debate about cloning, and stem cell research and DNA evidence is often used in rape and murder trials. There is another important field where molecular biology is commonplace -- although rarely mentioned -- conservation biology. Conservation biologists use molecular applications to genetically identify individuals, to define and trace pedigrees, to enhance breeding programs for critically endangered species, and to determine the source of poached animals and invasive plants. Impressive examples of conservation enhanced with molecular knowledge abound.
Using molecular technology, in 1997, I began studying an endangered seabird, the Roseate Terns (Sterna dougallii), for my dissertation research at the University of Massachusetts Boston. Roseates are easily distinguished from other tern species by a few key features: graceful, long streamers at the outermost points of the tail, a beak more black than orange, and a beautiful, rose color to the breast early in the breeding season. They are a cosmopolitan species, found in the United States from Maine to Long Island, and also in the Caribbean, the west coast of Europe and Africa, the Indian Ocean, and Australia. Although it sounds as if they are everywhere, Roseate Tern populations are small and quite fragmented.
Roseate Terns nest on small, sandy and rocky islands just off the coast and, occasionally, on peninsular beaches. Due to their precarious breeding habitat, and because they were hunted historically for plumage and eggs, Roseates have suffered several severe bottlenecks over the past 100 years. They were first listed as an endangered species in the United States in 1987, and have been quite slow to regain population size. There are probably several reasons for the species’ slow recovery. One major reason is habitat loss, both to humans and to nesting gulls. Another increasingly important threat is predation by land-based avian predators such as owls, falcons and Black Crowned Night Herons. Last summer, at one colony in Connecticut, Black Crowned Night Herons reduced fledging success of Roseate Terns by about 90% (Spendelow, 2002, personal communication). This is a major problem that has catastrophic potential since birds are known to avoid returning to breed in areas where they have had previous nesting failures.
Most recently, in April 2003, a large oil spill in Buzzards Bay has posed a serious threat. This spill occurred near two large breeding colonies of Roseate and Common Terns when the birds were prospecting for nest sites. Ram Island has recently been recolonized by Roseate Terns; however, it had to be cleaned extensively before the birds could be allowed to nest. This meant days and days of harassing the birds to keep them from landing and nesting on the oiled beach. At Bird Island, terns were coming to their nests with oil on their plumage that they were preening (and presumably eating) from their feathers. Studies are now underway to assess the impact of this spill on the terns led by Dr. Ian Nisbet. [See also, Harris article, The Seabird Ecological Assessment Network (SEANET): A Citizen Science Initiative for Marine Ecosystem Health] Additional losses or degradation of breeding habitat and predator problems could mean the end for these beautiful birds in New England.
The North Atlantic population of Roseate Terns evidences an interesting phenomenon -- there are too many females. Approximately 56% of breeding adults are female, which is another factor contributing to the problem of slow population recovery. Because males and females are sexually monomorphic (they look alike), it is not immediately apparent who is nesting with whom. The sex ratio bias, however, was noted as early as the late 1800’s when “super normal clutches” (SNCs) were first observed (Mackay, 1897). This phenomenon is thought to be increasing over time (Nisbet and Hatch, 1999). Because there are more females than males at the breeding grounds, some females cannot have a male mate of their own. To resolve this dilemma, females act together in tending a nest and attempting to rear chicks on their own. 
Female Roseates typically lay one or two eggs, but SNCs contain three, four, or even five eggs, indicating that two, three, or even as many as four females lay eggs in a single nest, sometimes with a male, and sometimes not. The eggs in SNC nests are often fertile. However, Roseates usually are unable to properly incubate more than one or two eggs, so SNC nests have lower hatching success than a male/female tended nest. Female groups encounter another problem, as well. Even when one or two chicks hatch, the female groups have difficulty feeding the chicks enough to prepare them to fledge the nest. Fledging success is considerably lower in these SNCs than in normal nests. Therefore, the female-biased sex ratio creates a further problem to Roseate recovery because many nests are not producing chicks that are able to leave the nest and eventually return to breed on their own.
The ability to determine the sex of an organism is one example of how molecular techniques have revolutionized conservation biology. Researchers all over the world are currently employing this technology. I have a genetic marker (Sabo et al. 1994) that allows me to identify the sex of each bird. This is essential in my research not only to determine the sex of adults (I cannot tell by just looking), but also the sex of chicks that have just hatched. Even in species that are sexually dimorphic (males and females look different), typically, one cannot determine the sex of chicks just by looking at them until they are much older. By trapping the adults on the nest, I can take a blood sample of each one back to the lab to determine how many females and males are tending the nest. This marker does not indicate WHICH females have actually laid eggs in the nest, just those that are tending it. I have been able to confirm the existence of multi-female associations at nests with two eggs as well as SNCs with three, four, or five eggs.
I have also been able to determine when the female-bias first occurs in the Roseate Terns’ life cycle. I spent the summer of 1997 at one breeding colony, Bird Island in Buzzards Bay, MA, where I trapped and sampled adults tending nests, and took blood samples from the chicks in those nests. I wanted to find out (1) if the eggs that are laid contain more female embryos than male, (2) if more female chicks hatched than male (higher male embryo mortality), and (3) if female chicks had a higher survival rate to fledging than male chicks. The study at Bird Island uncovered some interesting information (Szczys et al. 2000). We estimated that 53% of eggs laid are female, and that the female bias increases during the time of parental care so that 56% of fledging chicks are female. At the Bird Island colony, we found that there is no difference in mortality rate between males and females during the time of parental care. Instead, mortality depends on the order in which the eggs are laid in the nest. The second of two chicks in a nest has lower fledging success than the first, especially when eggs are laid later in the season. In addition, more of those second eggs are male, which means that fewer males fledge the nest (Nisbet and Szczys, 2001).
While we have some very interesting results on when the sex ratio bias manifests in the North Atlantic population, we still do not have an explanation as to why we see these biases; on the surface it does not seem to be an adaptive strategy. This is work in progress; in my preliminary investigation of sex ratio bias at another colony on Falkner Island, CT, I have not found the same evidence of sex bias at hatching, even though there are many nests with multi-female associations tending them. Similarly, in a small study conducted in Australia, there was no evidence of a sex ratio bias at breeding (Hatch and Szczys, 2000).
The second part of my research is focused on determining the genetic mating system of the Roseate Tern. A Roseate Tern cannot raise chicks alone. 
There must be a pair tending the nest because of the high demands of fishing and nearly constant incubation. Females that tend nests together often lay fertile eggs, which means that some other female’s male mate is fertilizing them. There are many questions we can ask about a genetic mating system that is cryptic and not always in line with the obvious social mating system. If males are cheating on their female partners, are the females also cheating by soliciting extra-pair fertilizations (EPFs) while their mates are away? Do these EPFs result in offspring? Who are the males that are fertilizing the eggs in the multi-female association nests; is it just one male or many? Are the female pairs sisters, mothers and daughters, or completely unrelated?
In order to investigate these questions, I have again employed techniques in molecular biology, but this time I have developed a different kind of genetic marker. Microsatellite markers allow for individual identification of plants and animals. They work in a way that is very similar to the DNA fingerprints that one often hears about being used in criminal justice. Microsatellite markers serve as a genetic signature that uniquely identifies an individual.
I have collected blood samples from approximately 200 families (3-4 individuals each) of Roseate Terns at the two colonies in Massachusetts and Connecticut. After extracting the DNA from the birds’ blood, I use the microsatellite markers to identify the parents at each nest, and determine if there are any young being raised in that nest that are not the biological offspring of (typically) the social father. Sometimes, females lay eggs in the nests of others (conspecific brood parasitism), but this is uncommon, we believe, in most birds other than obligate brood parasites such as the cowbird. However, an offspring may be found that is related to its mother but not to its social father. My study is still in the data collection process, and I haven’t yet analyzed how many extra-pair offspring there are in Roseate Tern nests. An interesting fact in avian mating systems is the huge range in the rate of EPFs that is found across all bird species. Some birds, such as the Acorn Woodpecker show 0% of young in nests as being unrelated to their social father; others, such as the Reed Bunting, show that up to 55% of young are unrelated to their social father (Griffith et al. 2002). My data will allow me to address questions about the evolution of avian mating systems, to see where within the range of EPF rates the Roseate Terns fall, and with any luck, it will also shed more light on the problem of the sex ratio bias in the North Atlantic population.
One more application of genetic data is perhaps the most important to improving the management and recovery of the Roseate Tern. Looking at individual genetic profiles at a population level, we can determine if the birds are actually one genetically distinct population or several smaller populations within the North Atlantic. Terns are quite philopatric, meaning that they generally return to breed at the colony from which they hatched. This could mean that each colony is genetically distinct. Saving genetically diverse sub-populations is essential for the future health of the species. We know from banding data that some birds do, indeed, show up at other colonies during the breeding season. However, until the genetic data comes in, we can’t know if they are contributing their genes to the next generation; nor can we determine if we have genetically distinct subpopulations. This type of gene flow information is a critical part of making appropriate decisions about breeding site conservation/restoration and predator intervention, and ultimately, the recovery of this endangered species.
The application of molecular techniques to conservation biology has been revolutionary. Avian behaviors that were held in esteem as exemplary mating systems in the animal world have been exposed as false. Thirty years ago, birds were believed to be the model of morality, strictly monogamous (male/female) pairs working in harmony to raise young. At that time, no one could have anticipated such an array of interesting covert behaviors: females working together to tend nests, cuckolded partners raising not only their own young but a few of the neighbors’ offspring as well. Determining actual gene flow, rather than just physical movement, between groups and defining the source population of an individual are imperative to conserving genetic diversity and eliminating illegal animal trading or harvesting. These processes were extremely time-consuming and expensive, if not nearly impossible, before the widespread use of molecular techniques in ecology. Hopefully, future innovations in biotechnology will continue to supply powerful tools that can further enhance conservation efforts.
Colin Hughes introduced me to the “bird world” and helped me realize the strength of molecular ecology in conservation biology. I am indebted to him for microsatellite development expertise and for lots of support and encouragement. Ian Nisbet introduced me to Roseate Terns and has allowed me to work with him on Bird Island where he has been studying terns for more than thirty years with great dedication. Richard Kesseli is my academic advisor at the University of Massachusetts Boston where he has provided funding and lab facilities, and shares an interest in conservation biology using molecular techniques. Jeff Spendelow is my collaborator at the Falkner Island, CT colony. He generously helped me trap and sample families during two breeding seasons at the colony he has been studying for over 20 years. My research is not possible without the help of these people and others.
All photographs copyright by Patricia Szczys and cannot be reproduced without permission.
Griffith, S.C. Owens, I.P.F., and Thuman, K.A., 2002. Extra pair paternity in birds: a review of interspecific variation and adaptive function. Molecular Ecology 22: 2195-2212.
Hatch, J.J. and Szczys, P., 2000. Lack of evidence for female-female pairs among Roseate Terns, Sterna dougallii, in Western Australia contrasts with North Atlantic. Emu 100: 152-155.
Mackay, G.H., 1897. The terns of Muskeget Island, Massachusetts, Part III. Auk 14: 278-284.
Nisbet, I.C.T., and Hatch, J.J., 1999. Consequences of a female-biased sex ratio in a socially monogamous bird: female-female pairs in the Roseate Tern, Sterna dougallii. Ibis 141: 307-320.
Nisbet, I.C.T., and Szczys, P., 2001. Sex does not affect early growth or survival in chicks of the Roseate Tern. Waterbirds 24: 45-49.
Sabo, T.J., Kesseli, R., Halverson, J.L., Nisbet, I.C.T., and Hatch, J.J., 1994. PCR-based method for sexing Roseate Terns (Sterna dougalli). Auk 111: 1023-1027
Spendelow, Jeffery, USGS Patuxent Wildlife Research Center, 2002. Personal communication.
Szczys, P., Nisbet, I.C.T., Hatch, J.J., and Kesseli, R., 2000. Sex ratio bias at hatching and fledging in the Roseate Tern. Condor 103: 385-389.
The views and opinions expressed in all articles that appear in "Conservation Perspectives" are those of the authors and do not necessarily reflect those of NESCB.