Tetracycline

Ancient Nubians and Egyptians had a way with antibiotics.

Some twenty years ago, Debra Martin placed a bit of bone from a mummy under a microscope and discovered that a person who lived in Nubia (northern Sudan) during the fourth century A.D. had apparently ingested tetracycline, a broad-spectrum antibiotic that entered the arsenal of modern medicine only in the 1950s. Finding a pair of designer sunglasses on the mummy would hardly have been more startling. And the discovery was purely serendipitous.

Today Martin is a professor of anthropology at Hampshire College in Amherst, but at the time she was a graduate student in biological anthropology at the University of Massachusetts. As part of her training, she was visiting a research laboratory at Henry Ford Hospital in Detroit, Michigan, to learn techniques for making thin sections of bones from archaeological finds. Normally she would have relied on a standard microscope, and the tetracycline would have gone undetected. But because the standard microscope was unavailable, another researcher suggested Martin try one that used ultraviolet light.

At one specific wavelength, ultraviolet light causes tetracycline to fluoresce with a unique yellow-greenish color. In the lab, researchers under the direction of Harold Frost were using tetracycline to measure the rate of bone formation. Tetracycline tends to bind with calcium and phosphorus, which make up more than 80 percent of the mineral portion of mature bone. (Patients who are taking the drug are advised not to drink milk or take antacids containing calcium, since the tetracycline will bind to the calcium and lose its antibiotic effectiveness.) Any tetracycline circulating in the body may bind with calcium that is being deposited in the bone, "labeling" (tagging) the bone with its indelible signature. In the laboratory study, people who were scheduled to have bone removed during biopsy or amputation were asked to take tetracycline at intervals before the surgery. Bone deposits formed during this period could then be identified and measured.

When Martin returned to the University of Massachusetts, where I was then teaching, she told me of her discovery, and we began to explore several issues: Was this really tetracycline? If so, was it incorporated into the bone during the subject's lifetime 1,600 years ago, or could it have been produced by organisms that invaded the remains after death? If it was ingested by ancient Nubians in their food or medications, what was its source?

That we really were dealing with tetracycline was demonstrated by James Boothe, a chemist who had worked on the initial commercial applications of the antibiotic for American Cyanamid. He was able to extract it from our Nubian bone and show that it could still kill bacteria. More recently, Mark Nelson at Paratek Pharmaceuticals has been determining its precise molecular structure (there is actually a whole family of tetracyclines in nature).

Evidence that the tetracycline was incorporated during the lifetime of the Nubian mummy came from its osteons, which are microscopic cylindrical building blocks of cortical bone (such as the outer layers of bone shafts). In response to physical stresses, bone tissue undergoes a continual process of fine-tuning. Bone cells called osteoclasts break down small amounts of bone mineral, which other cells, called osteoblasts, then replace. The result is the formation of new osteons. It takes about four months for any one osteon to become fully mineralized, and tetracycline may be incorporated during the process. When we examined bone from the Nubian mummy, we found that some osteons had layers of mineral containing tetracycline alternating with layers without tetracycline. Such a pattern could have developed only during life, not if the tetracycline was somehow introduced later; it indicated that while these particular osteons were forming, the individual was ingesting tetracycline intermittently. In most of the osteons we examined in the mummy, however, we found that tetracycline was present in all the layers, suggesting that during the four months it took for these osteons to mineralize, this individual had continuously ingested the antibiotic.

To determine the extent of tetracycline use by ancient Nubians, three undergraduate researchers in our lab at Emory University--Kristi Kohlbacher, Jennifer Cook, and Kristy Collins--painstakingly sampled thousands of osteons from our original mummy and from seventy-seven other Nubian and Egyptian remains dating from about the same era. All but four of the seventy-eight individuals showed some degree of exposure to tetracycline, and no significant differences by age or sex were evident. Even the remains of two of the three infants contained tetracycline, showing that it was passed to them in their mothers' milk.

Following the publication of these findings in the 1980s, other researchers began to report evidence of tetracycline in African prehistory. Physical anthropologist Megan Cook (then at the University of Toronto) and her colleagues, for example, found that the mummified remains of all twenty-five individuals recovered from Dakhla Oasis in Egypt, dating from the Roman period (A.D. 400-500), showed tetracycline labeling. The patterns were consistent with doses occurring at two- to three-week intervals. And Ann Grauer and I have recently reported evidence of tetracycline in bone from a Jordanian site that dates from the second century B.C. through the fourth century A.D.

But none of this told us why the antibiotic was showing up in the ancient bones. In nature, tetracycline is produced by streptomycetes, moldlike bacteria commonly found in soils. These slow-growing cells do not do so well in the wet, acidic soils where most bacteria flourish, but they have the edge in hot, dry, and neutral-to-alkaline environments. Ten-year-old spores survive in dry sand and are easily cultured.

Initially we thought that during famine or drought, the ancient Nubians and Egyptians might have been forced to eat moldy grain. (Even one or two grams of tetracycline consumed by humans in a single day will produce fluorescence in bone.) The warm, dry, alkaline environment of storage bins made of mud could have been an ideal environment for streptomycetes. But we learned that when they are growing well, streptomycetes actually produce little tetracycline. Given the degree of tetracycline labeling in the Nubian and Egyptian remains, we had to consider other possibilities. The key turned out to be beer, known as bosa in much of present-day Africa.

Searching through both ancient and later texts, Everett Bassett, Margaret Keith, and other members of our team realized that in the region's grain processing, there was an important link between bread baking and beer brewing. Egyptian art also shows baking and brewing in constant association. In fact, baked bread is an essential part of the traditional beer recipe still used today by villagers who live along the Nile.

The beer produced in ancient times, according to Barry Kemp, author of Ancient Egypt: Anatomy of a Civilization, was quite different from the modern commercial product: "It was probably an opaque liquid looking like a gruel or soup, not necessarily very alcoholic but highly nutritious. Its prominence in the Egyptian diet reflects its food value as much as the mildly pleasurable sensation that went with drinking it." University of Cambridge archaeologist Delwen Samuel and his colleagues from the British brewery Scottish and New-castle have undertaken extensive research on brewing and baking in ancient Egypt. They analyzed the remains of food left in tombs as offerings and the residues of beer and crumbs of bread encrusted on pottery shards and vessels. They even examined floor sweepings from tombs and living areas.

Successful brewing depends on the use of a grain that provides enough sugar for fermentation. In modern recipes, grain is made to germinate and is then heated and dried to halt the process. Known as malting, this procedure releases the enzyme diastase, which converts the starches in grain to maltose sugar. The malt is then boiled, strained, and incubated with yeast. In the traditional Egyptian method, bread dough is set out to capture airborne yeast. (Other traditional recipes actually add bosa that was held back from previous batches for this purpose, since the liquid contains yeast.) When baked, the bread forms a crust but is removed from the oven before the center has had a chance to cook, allowing the yeast to grow in the warm, slightly cooked dough. The partially baked bread is then broken up and added to a broth of malted grain to make the beer.

We theorized that airborne streptomycete spores were captured in the ancient brewers' dough during its exposure to the air and that the streptomycetes then produced tetracycline while the yeast grew in the partially baked bread. To investigate brewing's capacity to give rise to tetracycline, Daniel Popowich and Brennan Posner, undergraduates at Emory University, added streptomycetes during two experiments with the traditional process. In the first, they added a small colony of streptomycetes to the just-baked bread; in the second, they added the streptomycetes to the mixture of malted grain and bread. The second technique was the more successful and produced significant amounts of tetracycline.

The fermenting brews of ancient times, we concluded, provided the somewhat harsh environment in which the streptomycetes were stimulated to yield tetracycline in quantity. Nowadays, companies that make pharmaceuticals deliberately control and limit certain nutrients as a way of forcing streptomycetes to make tetracycline.

Given that the ancient Nubians and Egyptians were getting doses of tetracycline, another question is whether this afforded them any medical benefits. In Food: The Girl of Osiris, William J. Darby and coauthors provide archaeological, historical, and ethnographic accounts of beer's use as a mouthwash to treat the gums, as an enema, as a vaginal douche, as a dressing for wounds, and as a fumigant to treat diseases of the anus (the dried remains of grains used in brewing are burned to produce a therapeutic smoke). This shows that even in the distant past, Egyptians and their neighbors appreciated beer's medicinal qualities.

Today tetracycline remains the drug of choice in the treatment of both acne and gingival disease. Researchers studying gum disease originally assumed that the tetracycline worked because of its antibiotic qualities. But tetracycline also appears to inhibit collagenase, an enzyme that breaks down collagen. There has been a concerted effort to produce chemically modified tetracyclines (CMTs) that have this effect but not the antibiotic qualities. In addition, both tetracycline and CMTs have proved to be very effective in inhibiting matrix matallproteinases, enzymes involved in a number of bone and connective-tissue diseases, such as rheumatoid arthritis, osteoarthritis, periodontal disease, osteoporosis, and even cardiovascular disease. The ingestion of tetracycline may thus have had real medical benefits for ancient Nubians and Egyptians.

As we enter the new millennium, many people are concerned that our own use and abuse of antibiotics in medicine, agriculture, and even manufactured products has been encouraging the rise of antibiotic-resistant bacteria. When we reported the discovery of tetracycline in ancient bones in the journal Science, we wondered whether, owing to long-term ingestion of the antibiotic, the Nubian and Egyptian populations might have suffered an increase in disease caused by resistant bacteria. To test this, we have examined the bones in our sample for signs of periosteal reactions--roughened surfaces that form as a result of bone infection. We have found no evidence that infections became more intense during the centuries represented by the bones, as would be expected if more resistant bacteria had evolved. But during our own lifetimes, 1,600 years later, many of us may well fall victim to bacteria that are resistant to all the known antibiotics. If we do, our bones will reveal this to archaeologists of the future.

George J. Armelagos is a professor of anthropology at Emory University in Atlanta.

COPYRIGHT 2000 American Museum of Natural History
COPYRIGHT 2003 Gale Group