Frederick H. Sheldon, PhD - US grants

The Louisiana State University Museum of Natural Science (LSUMNS) Collection of Genetic Resources is probably the largest and most active collection of wild vertebrate frozen tissues in the world. It contains nearly 40,000 specimens and over 100,000 samples of frozen tissues and extracts (e.g., DNA, proteins, and antisera). In the last five years, the Collection has given more than 2,000 samples of frozen tissues to researchers form 10 countries. During the same period, more than 80 publications and theses relying on LSUMNS tissues and extracts have been produced. In the years to come, demands for tissues and tissue extracts are expected to rise substantially as more and more researchers turn to comparative molecular methods and as the LSUMNS Collection grows and becomes better known as a repository and source of genetic materials. Improvements in molecular technology and methodology will cause an increase in specimen requests form systematists and population geneticists (currently the Collection s main users), and will lead to greater interest in the Collection form nontraditi onal sources -- ecologists, behaviorists, wildlife managers, epidemiologists, and other comparative molecular biologists. Already, the Collection is feeling an impact particularly from bird ecologists and behaviorists interested in an historical (phylogenetic) perspective from which to interpret their copious life- history data. The LSUMNS Collection must meet this expected increase in demand. The Collection is growing rapidly as a result of the Museum s aggressive program of collection and soliciting tissues and tissue extracts. This program is intended not only to increase holdings but also to salvage and centralize tissues and genetic extracts that might otherwise be scattered and unavailable to the research community. This project will support repairing, replacing, and adding new ultracold freezers and their alarms and backup systems. It will also assist in storing and in preparing and mailing of frozen specimens to researchers at other institutions.

A cophylogenetic analysis of avian hosts and their parasites: toucans
(Aves: Piciformes) and chewing lice (Insecta: Pthiraptera)

I am using nuclear and mitochondrial DNA sequences to reconstruct the phylogenetic history of an avian host group (Ramphastos toucans) and two of its parasitic chewing louse genera (Austrophilopterus and Menecanthus), which differ in life history characteristics. The object of this study is to determine how parasite groups with different ecological, behavioral and morphological characteristics respond to identical geographical and habitat perturbations (diversification and speciation of the host group). I will compare host and parasite histories by superimposing the phylogenies of the parasites and their hosts to determine whether or not parasite lineages with differences in ecology, behavior, and morphology respond similarly to speciation events in their hosts. If differences in the life history of the chewing lice are important, the different parasite lineages should vary in their degrees of cospeciation (parallel speciation). However, if
parasite lineages respond to host isolation events similarly, then one would expect the phylogenetic trees of the two parasite lineages to be congruent.

Insights gained via this simple system should shed light on patterns of speciation in more complex systems of free-living organisms. Phylogenetic studies can answer important questions about speciation. However, one problem faced by phylogeneticists is that, when studying free-living organisms, they often lack nformation about the history of the study organism's habitats and geographic distribution. Thus, a useful
first step in phylogenetic studies is to reconstruct this history. Cophylogenetic studies of hosts and their parasites do this by superimposing the parasite phylogeny onto the phylogeny of the host. Speciation events in the host group are analogous to biogeographic vicariant events (e.g., when geographic barriers islolate opulations leading to speciation). Speciation, or lack thereof, in the parasites represents a response to geographic and ecological changes caused by host speciation events (vicariant events). By comparing parasites with different ecological, morphological, and behavioral characteristics we will begin to understand how these different life history characteristics affect the process of speciation.

Early Bird is a large-scale, cooperative effort among five institutions in the U.S., one in Scotland and two in Australia to determine the evolutionary relationships among all major groups of birds. The project will make these relationships known to the research community and the public, and make it possible to use these relationships as a comparative framework with which to organize and understand the vast amount of information already available on avian ecology, evolution, physiology, and behavior. The project will generate large amounts of DNA sequence data for all major avian lineages from a series of carefully selected genes. These data will be integrated with existing and new morphological and fossil data to bring all relevant evidence to bear on the problem. Extensive analysis of the evidence will be conducted with the rapidly expanding suite of computational tools available for the inference of relationships, divergence times, and evolutionary patterns. The data will be made accessible to other researchers in an online database with tools for analysis and export, and to the general public through a website with background information on the importance of evolutionary relationships, progress reports on the project, and interactive educational tools with which interested persons can explore the data themselves.
The immediate results of this project will be to provide a detailed, comprehensive and robust estimate of the "family tree" of avian relationships, and to facilitate the use of that tree to organize and interpret other information about birds. The impact of this project on science and society will be far-reaching. Birds are among the most prominent and engaging creatures in most ecosystems worldwide. They have been the subject of an extraordinary number and diversity of scientific studies that figure largely in our understanding of the natural world and humanity's place in it. Their position high in many food chains together with their great mobility makes them sensitive indicators of environmental quality, and monitoring of bird populations is widely used to set conservation and management priorities. Their powers of flight, physical beauty, and captivating behaviors amaze and inspire us, and birds provide tremendous amounts of recreation for serious hunters and birders, as well as millions of backyard birdwatchers each year. All of these human interactions with birds will be enriched by a better understanding of avian evolutionary history and genetic diversity, which Early Bird will provide.

A grant has been awarded to Dr. Frederick Sheldon and Mr. Christopher Witt at Louisiana State University to study the evolutionary origins of the exceptionally high species diversity of birds found in the Neotropics. Although biologists have known for centuries that the global distribution of biodiversity is skewed towards the tropics, little is understood about the phylogenetic relationships of tropical organisms and the historical events that may have led to their extensive diversification. This study will use museum collections to make DNA comparisons among every available species and population of two families of Neotropical birds, the puffbirds and jacamars. Trees of evolutionary lineages will be reconstructed using analyses that incorporate models of molecular evolution into measurements of DNA similarity. Patterns of new species formation and genetic differentiation will be compared among related groups that share common distributions. In this way, evolutionary trees will be used to infer the ages of evolutionary lineages and to identify the geological and climatic factors that have contributed to modern day diversity at all taxonomic levels. The results will have implications for the ongoing debate regarding relative contributions of long-term geological stability versus recent climatic tumult to the disparity in diversity between temperate and tropical bird faunas.
This study will involve collection of DNA data from many related organisms across a broad geographic region. The results will have two primary benefits: (1) inference of evolutionary causes of species diversification; and (2) identification of conservation priorities. Current classifications almost certainly do not adequately represent true genetic diversity in tropical organisms. Genetic studies such as this one are needed to identify cryptic or overlooked species, to recognize areas that harbor significant genetic diversity, and to suggest strategies for protecting both the products and the process of evolution.

The Louisiana State University Museum of Natural Science Collection of Genetic Resources is one of the largest and most active collections of wild vertebrate frozen tissues in the world. It contains more than 100,000 samples of frozen tissues and tissue extracts (e.g., DNA, proteins, and antisera) from over 52,000 individual vertebrate specimens. The Collection is housed in 13 ultracold freezers, but needs for its protection to be moved to liquid nitrogen (LN) storage. Ultracold freezers have a minimum temperature of -80oC, whereas liquid nitrogen has a temperature of -150oC. The colder temperature would preserve the tissues more effectively, and LN storage would protect the specimens from damage caused by electric power outages, which are not uncommon in Louisiana. In addition, many of the herpetological samples in the Collection require substantial curatorial attention, including transfer from old containers to Nunc tubes, database updating, and cross referencing with voucher specimens. This grant would fund the transfer of herpetological specimens from ultracold freezer storage to LN over the next three years. It would also fund the upgrade of the herpetological collection in the process. In the future, the rest of the Collection would be moved to LN storage. Specifically, the project would entail purchasing two LN freezers and associated equipment, which would be capable of holding about 60,000 samples, and employing a herpetology graduate student as a curatorial assistant to oversee the move. The curatorial assistant would review the packaging and data of each of our ca. 15,000 herpetological specimens while moving the samples to LN freezers.

The project would have several benefits for the Collection and the research community. The herpetological specimens would be better protected, and their data would be verified, making them more useful as specimens. The graduate curatorial assistant performing the work would gain training in the curation of frozen genetic collections, a skill that will become increasingly important as the number of these collections grows. The research community will benefit from improved curation and care of the Collection in general. From 1995 through 2000, the Collection accessioned 15,817 tissue specimens from 22 countries and provided 285 grants consisting of 3,908 tissues to researchers in 11 countries, plus the USA. Since 1995, at least 240 theses, papers, and books containing results based on comparisons of the Collection's tissues and extracts were published. The contribution of the Collection to the productivity of the research community will grow directly as a result of the grant.

This award provides support for acquisition of a modern capillary DNA sequencing instrument capable of determining nearly 200,000 bases a day at 98.5% accuracy, and also capable of high throughput fragment analysis including that of microsatellites and single nucleotide polymorphisms. The instrument will be used in a number of ongoing projects, including an NSF-funded Tree of Life sequencing project aimed at producing a molecular phylogeny of all birds. The instrument will be used. In addition to its role in research, the instrument will be used for training at the high school, undergraduate, graduate, and post-doctoral levels. The instrument will replace an existing slab-gel instrument that has been heavily used for studies of population genetics and evolutionary genomics of avian systems, co-evolutionary genetics of mammals and their parasitic lice, and evolutionary genetics of reptiles and amphibians. The new instrument will provide a several-fold increase in the sequencing capacity that can be brought to bear on these and other projects at the Museum of Natural History.

Darwin's finches of the Galápagos Islands stand as one of the most impressive examples of evolution. After colonizing the islands, a single ancestral finch species eventually diversified into 13 species of different sizes, bill shapes, and ecologies. The suboscine passerine birds exhibit a similar history, but their diversification was not limited to a small set of islands; instead they are found throughout South America, Central America, North America, and parts of the Old World. There are now 1,300 species that differ dramatically in body shape and size, feeding behavior, and nest architecture. The primary goal of this project is to use DNA sequences to reconstruct the evolutionary history of these species, and to use this model system to investigate why the tropics are so biologically diverse.

A vital component of this collaborative project between U.S. and Brazilian scientists is the research training it will provide. With fieldwork planned in Argentina, Chile, Costa Rica, Malaysia, Venezuela, and Viet Nam, young scientists from a diversity of countries will be exposed not only to a diversity of natural environments but will also interact and forge relationships with their international peers. By working together to reconstruct perhaps the most spectacular evolutionary radiations of birds in the world, the researchers will learn how evolution works on a continental scale while training a new generation of scientists.

The causes of geographical gradients in biodiversity are not well understood, and remain a major scientific challenge. Many physical and biotic factors may influence the evolutionary diversification and species distribution dynamics that result in these gradients. This study examines an elevational gradient of tropical birds in Asia, where functional, genetic, and phylogenetic diversity of organisms are woefully understudied and underlying patterns provide a strong system for testing hypotheses about biodiversity along elevational gradients. The proposed research evaluates both traditional (physiological tolerance, competition, opposing boundaries) and novel (taxon age, adult survival, life history trade-offs) factors that vary along elevational gradients and integrates them with dispersal, genetic structure, and phylogenetic history.

The research partners will share samples and data, and collaborate in cross-disciplinary training of students, in pursuing tests of the proposed hypotheses. This integration will lead to a new mechanistic understanding and conceptual framework for the influence of physical climate gradients and biotic interactions on species distributions and the evolution of diversity. The project will provide extensive opportunities for the training of undergraduate and graduate students, including a unique cross-cultural life experience as well as hands-on educational and field experience. Public outreach includes work with high school students and teachers, museum exhibits, and development of an educational video on tropical bird nesting behaviors. Finally, this project has critical conservation ramifications, as tropical Asian forests are being lost at the fastest rate of any tropical forests in the world, and this research will provide critical information on possible climate change impacts that are of grave concern to extinction risks.