(click on images to enlarge and show captions)

---

Beetlebellyeasts

Meredith Blackwell, Professor of Biological Sciences, LSU

The first life on Earth was microbial in nature. Since the beginning, microbes, including fungi, have prepared the way for larger and more morphologically complex organisms through their prowess in ecosystem processes, often in associations with other organisms. For example the lichens that we learn about in grade school really do change rock to soil, but what is often less clear to the unaided eye is that lichens are mixtures of two or more organisms living in close association. Lichens are comprised of a photosynthetic organism (photobiont) and fungus. Often there is a third organism in the association to fix essential nitrogen, and very recent reports indicate that other fungi may live in the association. Still other fungi facilitated the move by plants to dry land by increasing the efficiency of absorption of nutrients and water from the soil, a process that continues to allow plants to cover certain vast marginal habitats, including some tropical forests. The fabled lone pine tree can become a dead pine tree by the time it is a few years old, because without other root-associated fungi it does not survive. Intricate associations of such unrelated organisms are known as symbioses and they are common occurrences. In fact there are no organisms that are not symbiotic in some sense of the term. Even humans have many relationships with other organisms. For example there is overwhelming evidence that mitochondria derived from aerobic bacteria invaded our earliest nucleated relatives, settled in to respire within their cells, and were passed on from mother cell to cellular offspring; bacteria-derived mitochondria are so well melded within our cells that we never consider most of them, unless they fail to function – resulting in death, as in cyanide poisoning.

Humans mostly have an anthropocentric view of the world; in our lab, however, we actually are more interested in discovering symbiotic associations among so-called lower organisms, most notably those that occur between fungi and insects. Toward this end we collect, dissect, and culture the gut contents of insects, thus far mostly beetles in Panama and the southeastern USA. After the collecting is done the real work begins. Identification and determination of the taxonomic status and relationships of the yeasts requires DNA sequencing and knowledge of yeast physiology and morphology when each is subjected to 100 different physiological tests and morphological observations.

It is evident that whether we work in Panama or literally in our own back yards –the gut of many insects contain single celled fungi known as yeasts; furthermore, many of the yeasts were unknown previously. Among our first 650 yeast isolates, almost 200 were undescribed species, a number amounting to a 30% increase over the 700 previously known yeast species. In addition, entire new insect-associated yeast radiations have been discovered and specific beetle-gut yeast associations extend over the range of certain beetles. Bayesian analyses estimate that resampling of the same habitats would greatly increase the numbers of species from these specialized habitats.

At first it is exciting to discover a new organism, but when 200 of our first 650 isolates from the gut of beetles in 27 families are undescribed, the exhilaration becomes tedium. How can we think up 200 new fungal names, let along write a Latin description of each (as required by our nomenclature system)! So far we have used names based on those of endemic peoples of Panama and Native Americans in the southeastern US. Thus we have used Latinized names commemorating the Bókatá, Bribri, Térraba, Guaymí, Cuna, Talamanca, Chocó, and Cho for yeasts from Panama and Chickasaw, Yuchi, Choctaw, and Atakapa for southeastern USA yeasts. The names seem particularly appropriate because some of these people were acutely aware of their natural environments and even recognized symbiotic associations in agricultural situations. We also have named several yeasts for people associated with the Panama Canal. One species honors the many Chinese laborers who died during construction of the canal and whose graves we see on the way to our collecting site on an island in the artificial lake created when the locks were built. Another yeast is named for William Crawford Gorgas, who dried out the holy water fonts in the churches of Panama in order to eliminate deadly disease-carrying mosquitoes during the Panama Canal construction.

At this stage of our research while we are still thinking up yeast names, we also want to know more about the associations themselves. Are the new yeasts restricted to the gut habitat? Are particular yeasts always present in certain beetles? Are the geographical ranges of certain beetles and yeasts congruent? When a beetle dies do the yeasts die too? Why should yeasts live in such a habitat? Are changes in gut yeasts correlated with changes in nutritional mode of certain beetles? What else influences the associations between beetles and gut yeasts? How do the yeasts get to a new host beetle? How and when do young beetles acquire yeasts? Do yeasts really perform services for beetles? It is presumed that beetles benefit from enzymes, sex attractants, vitamins, and lipids produced by the yeasts. Or perhaps the beetles simply provide a cushy place of residence for yeasts to acquire the leftovers ingested by beetles as a sort of internal remora? We don’t know all the answers yet, but such an understanding of the associations we discover so readily is a principal objective of our work. We do have hints about the associations, such as the production of B vitamins by the yeasts that might benefit the insects that generally lack the ability to synthesize them. In another example yeasts that inhabit the gut of certain wood–ingesting beetles produce rare xylose-degrading enzymes that could be useful in breaking down these components of the wood for the beetle as well as the yeast.

Who does this work? Dr. Sung-Oui Suh, a postdoctoral researcher, oversees the day-to-day operation of the lab. At least ten undergraduate students and several graduate students have worked on this project. The students are well-trained in culture and other techniques by Dr. Suh, the yeast expert of the lab. In addition to collecting insects and isolating and culturing the yeasts, the students learn to perform a wide variety of sophisticated molecular techniques independently, including DNA sequencing. Other people are involved in the research, and much of our work has been done in collaboration with Dr. Joseph V. McHugh, Department of Entomology, University of Georgia, who is the beetle expert. A new collaboration with Dr. James B. Nardi, Department of Entomology, University of Illinois, has allowed us to begin to investigate the yeast/insect gut interface using electron microscopy. In addition we are discovering other kinds of microbial gut inhabitants with not only microscopic techniques, but also by use of gene cloning techniques.

This past year has been good for our research because people are beginning to appreciate it. Not only has the lab been heavily involved in the editing of a new volume on insects and fungi [F. E. Vega and M.Blackwell, Eds. 2005. Insect-Fungal Associations: Ecology and Evolution.Oxford University Press, NY. 333 p] that includes two chapters on work done in the lab, but a recent article published in a mycological journal [Suh, S.-O., J. V. McHugh, D. Pollock, and M. Blackwell. 2005.Massive biodiversity yeasts from the gut of basidiocarp-feeding beetles. Mycological Research 109:261-265] will be discussed in the “News and Views” section of the journal Nature. Other encouraging news was the request of a federal funding agency to use pictures from our web site to accompany a short discussion of our work in an in-house presentation on their funded research.

As we begin the first field season of a new grant, our collecting site is not yet certain. We want to return to the Smithsonian Tropical Research Institute at Barro Colorado Island, Panama http://www.stri.org/english/visit_us/barro_colorado/index.php , but the cultures we would work so hard to establish might be in vain if we cannot get an importation permit from the USDA, something that is increasingly difficult to obtain in a timely fashion. In fact no longer can fungal cultures be hand carried home with us from a foreign country, but they must be shipped to a USDA approved facility, causing delays that promote contamination by other faster growing fungi. After Panama, permits allowing, we intend to compare the beetles and their yeasts from Panama with those we hope to be able to collect later in South Africa.

The National Science Foundation requires that there be broader impacts of our study that include training of future scientists and the contribution of our research products to natural history museums, culture collections, and DNA databases where everyone can have free access to them. Our students have been successful, and recent lab alumni have gone from LSU to graduate schools at Duke, Auburn, and Oxford Universities. At least four others have gone to medical school. Of the current crop of students nearing the end of their undergraduate careers, one is graduate school bound and another has been accepted into medical school. We also are expected to present our results at scientific meetings and to publish in peer-reviewed scholarly journals. Additionally, some of our data are available rapidly on the Internet, which we are doing by developing an interactive database http://nt.ars-grin.gov/taxadescriptions/keys/YeastsIndex.cfm . The growing database contains information on 170 yeasts that were collected in association with the beetles. Other research products include about 800 DNA sequences from gut yeast cultures and cloned sequences from the gut of beetles that have been deposited in GenBank, a public data base of DNA and protein sequences http://www.ncbi.nlm.nih.gov/Entrez/ and other data. Preserved specimens of insects, mushrooms from which the insects were collected, and yeast cultures have been deposited in several natural history and culture collections. The work has provided habitat photographs http://www.ent.ua.edu/personnel/faculty/strand/erotylids.html done by Joe McHugh for picture keys of beetles of Barro Colorado Island; we also have collaborated on the Tree of Life web project http://tolweb.org/tree/ that seeks to provide a site where all groups of organisms can be discussed. You can see more about our work and find links to photographs from collecting expeditions at our lab web site http://lsb380.plbio.lsu.edu/beetlebellyfolder/beetlebelly.

-- Meredith Blackwell

** BACK **