Unlocking the Past

by Martin Jones

  At the time of writing, this has not been taken further, but work to do so is under way in Oxford. Across Europe, there are many burials coinciding with the Justinian Plague that would be suitable to test. The most interesting goal would be to establish when and where the bacterium prevailed in prehistory. It may have been something relatively new in classical times, a consequence of the sheer scale of trading links in the ancient world, bringing so many together from distant regions into large classical towns. On the other hand, in an agrarian world in which clustered permanent settlements were the norm, these pandemics may always have been a possibility whenever some external factor shifted the sensitive balances between micro-predator and human prey. On the basis of ancient tree-ring evidence, Mike Bailley has argued that the climatic perturbations that might encourage pandemics are scattered through prehistory. DNA may now begin to provide an answer.

  the white death

  The plague bacterium has come to and gone from the Old World population according to a long-term dynamic, one which we have yet fully to comprehend. Only when the search for Yersinia pestis has reached back into prehistory will we be able to do so. Some other diseases by their very nature seem to fit into a long-term trajectory of the human past.

  The ancient Egyptians described a disease that was almost certainly tuberculosis, as did Hippocrates, the Greek forefather of medicine. Tuberculosis, sometimes known as the ‘white death’, is a disease the spread of which would be directly stimulated by the central features of Childe’s agricultural and urban revolutions. Farming brought the lives of humans and animals close together, and cattle are a likely source of the disease. Close proximity between townspeople further assists the bacterium responsible for this illness. In historical times, up until the twentieth century, tuberculosis has been a faithful and fearful companion of the progress of industrialization and urbanization around the world. Infirmaries for the sufferers of ‘consumption’ became as familiar a feature of nineteenth-century cities as their smoke-filled skies.

  The disease gives the impression of being an artefact of the Neolithic deviation from nature. Particular strains of the very hardy tuberculosis bacterium are passed easily between humans and cattle, particularly if the blood or the milk of those animals is consumed. As the relationship between humans and domesticated cattle has grown increasingly intimate, so has tuberculosis flourished. This much can be gleaned from recent medical history, and by records showing how, in recent centuries, the spread of tuberculosis accompanied the spread of western agriculture across the world. It has not simply been a marker of the proximity of humans and cattle, but has also flourished where humans themselves live extremely close to each other, especially where overcrowding has been accompanied by malnutrition and poor hygiene. While industrialization and urbanization moved steadily forward in the eighteenth and nineteenth centuries, tuberculosis reached near-epidemic proportions, and was the leading cause of death in the western world. Infirmaries were built to house the sick, but sometimes the crowding and poor living conditions in these places of refuge further encouraged the spread of this poorly understood disease. Late eighteenth-century records of one such infirmary in Newcastle, in the north of England, show that one in ten of the patients admitted suffered from tuberculosis. By the time the patients within that infirmary expired, the prevalence of the disease had almost trebled. The sailors, pitmen, labourers, soldiers and industrial workers who were admitted to the infirmary in great numbers met with the overcrowding and malnutrition upon which the disease relied.

  With the early development of agriculture came the shrinkage of the food chain to a small number of domesticated species, all concentrated together with the human population in compact, permanent settlements. It has been seen as the ecological precondition for population growth and complex society. It also seemed to be the ecological precondition for diseases such as tuberculosis, and for the widespread suffering they inflicted. This progress of tuberculosis through the ages, running in parallel with the progress of agriculture itself, can be followed in the archaeological record through the skeletal deformations suffered by some of the afflicted. One particular pattern of bone deformation was very characteristic. An eighteenth-century physician, Sir Percivall Pott, was the first to describe the pattern of eroded spinal discs and collapsed vertebrae that gave its sufferer an angular spine and a hunched back. The condition is still known as Pott’s disease. Similar bone deformations, which may have resulted from tuberculosis, have been found scattered through the archaeological record, dispersed along the path of agricultural and urbanized societies.

  One of the first Neolithic farmers to open up fields near to what is now Heidelberg in Germany suffered from the disease. By the Bronze Age, deformed bones are accompanied by pictorial representations of hunchbacked sufferers of Pott’s disease, and by the beginnings of the historical period, skeletons with the characteristically collapsed spine are found as far to the east as Japan. This spread of evidence mirrors the close relationship between tuberculosis and settled life, farming and the husbandry of domesticated cattle. When Europeans transformed human settlement in the New World, the virulence of tuberculosis became global. If, however, we look back beyond the European discovery of America, we become aware that this is not the complete story. Something else was going on, prior to that major phase of globalization. When Columbus’s ships reached America five centuries ago, they may well have brought a range of diseases, but this mass killer seemed already to be on the loose.

  Once again our information comes from the fringes of the Atacama desert in Chile, where the particular environmental conditions have led to the remarkable preservation of a group of mummified individuals of the pre-Columbian Chinchorro and Inca societies. One of these individuals was a young girl in her early teens, laid to rest 1,000 years ago in a simple shirt and sandals. One aspect of her garments revealed something significant about her posture. A solid sash and belt had been made to support her lower back, and when her bones were studied, it was clear why. They were as porous as a piece of Swiss cheese. She must have been in great pain, a pain alleviated by chewing coca leaves. She still had a few of these narcotic leaves lodged in her cheek cavity when she died.

  This young girl was one of several people found as bodies or skeletons in South America who have the skeletal symptoms of tuberculosis, but she presented something of an enigma. The cattle, and the Old World farmers who we had assumed carried the disease, had yet to arrive in the New World. They would not do so until several centuries later. Her radiocarbon date was AD 1040 ± 70, four to five centuries before Columbus’s journey. Other New World bodies take the disease even further back, to as far as 800 BC. If the supposed link between Old World animal husbandry and tuberculosis holds, how could tuberculosis have reached pre-Columbian America in this way? One major limitation of these data is that none of the bone deformities is totally specific to tuberculosis. It could have been argued that the New World deformities had some other cause–that is, until the possibility of detecting the bacterium itself arose.

  Of all the disease organisms that might be tackled using ancient DNA analysis, the Mycobacterium genus, which includes the species responsible for tuberculosis, has received the greatest attention. A diagnostic stretch of DNA sequence has been identified, a repeat of an insertion occurring several times across the genome and labelled IS6110. By 1993, Mark Spigelman had published a successful amplification of M.tuberculosis DNA in a variety of archaeological skeletal remains. They came from a wide variety of contexts, including Byzantine Turkey, mediaeval England and seventeenth- to eighteenth-century Scotland. At around the same time, Bernardo Arriaza’s team had begun looking for disease among their rich assemblage of Chinchorro and Andean mummies. They tested for tuberculosis DNA within the vertebrae of the young teenage girl whose suffering from Pott’s disease was so clear. She too gave a positive PCR product for the bacterium. Further to the north, in the Osmore Valley in Peru, another group had succeeded in amplifying tuberculosis DNA f
rom the lesion in a mummified woman’s lung. The case was strengthening for pre-contact tuberculosis in the New World.

  Perhaps the link between the Old World Neolithic revolution and tuberculosis was not quite as clear as it at first seemed. Instead, it is more likely that tuberculosis arrived in the New World with the first colonizers from Beringia. We now know they brought domesticated dogs with them, but certainly not cattle, which had yet to be domesticated and in very different parts of the Old World. It need not have been the humans themselves who brought tuberculosis across, but instead some other mammal, such as buffalo, elk, moose or deer, the populations of which were also expanding into America. Either way, it must have passed between wild animals and their human predators, rather than depending upon the Neolithic deviation and the artificial intimacy of a permanent farming settlement.

  When amplifying the DNA, not from the host organism itself but from a separate microbial species that may have infected it, we have to be doubly careful about contamination. Some close relatives of the tuberculosis bacterium are free-living species within the soil. Furthermore, there may well be Mycobacterium relatives that are now extinct. The immense detective power of the polymerase chain reaction could be deluding us here–we could be making too much of a tiny number of DNA molecules. Those molecules might even belong to a close relative of tuberculosis rather than to tuberculosis itself, and perhaps we needed some other route to this ancient disease.

  from the inner nucleus to the outer coat

  The bacterium at the heart of tuberculosis is not simply a package of DNA, but has a fairly complex cell wall designed to defend itself on the perilous journey between and within hosts. The thick wall that guards against desiccation and chemical attack is built of sugars, proteins and long fatty acid molecules packed together. The fatty acids are quite distinctive. Whereas the more familiar fatty acids are built around a single linear carbon chain, these ‘mycolic acids’ have a complex, branched structure that varies from one type of bacterium to another. They are, in other words, markers of the particular bacteria present. If any of these bacteria leave their DNA behind in the bones, then there is a fair chance they will do the same with their lipid coats. As we saw in the previous chapter, lipid residues can be extremely persistent in archaeological materials, thanks in large part to the inability of the soil water to dissolve them and wash them away.

  At Newcastle, Angela Gernaey embarked on a search for these characteristically branched my colic acids in ancient bone. She had the good fortune to be around when the burial ground attached to the Newcastle Infirmary described above was being excavated, in advance of a new building development. The excavation was a telling experience. So many of the sailors, pit-men and industrial workers who spent their final days here were clearly disposed of, when dead, hurriedly and without much ceremony. They were stacked up, without rows or plots, in a crowded grave space, sometimes in a coffin, sometimes not. Certain of these skeletons had the physical appearance, the collapsed spine or bone lesions, that we associate with tuberculosis. Those were, however, just the tip of a much larger iceberg of infection, as Gernaey was to discover.

  She selected a rib bone from each individual from a sample of twenty-one. One had suffered from a collapsed spine, but the others showed no skeletal sign of infection. The sampled rib bones were ground up and their lipids dissolved and slightly modified in order that they could be assayed using a technique called High Performance Liquid Chromatography (HPLC) that sorts molecules according to their mobility through a carefully designed liquid medium. Gernaey built an HPLC fingerprint for each of the twenty-one ribs, and then a separate ‘control’ fingerprint from a clinical sample of modern tuberculosis, and an extensive series of samples of soil was taken from immediately adjacent to the excavated ribs. The different fingerprints were compared, and the result was clear. The soil samples completely lacked any of the lipid peaks displayed in the control sample. The same was true of sixteen of the rib bones. For the other five, Gernaey found a clear match between the ancient samples and the modern. Just as around one in four patients is recorded as having died of tuberculosis, the rib bones of one in four of the buried skeletons have retained part of the disease bacterium’s lipid coat. The lipid has survived in great enough quantities to be detected centuries later.

  DNA, lipids and the different types of physical bone lesions are each telling us something slightly different about the disease within these ancient skeletons. The DNA indicates the presence of the disease organism itself, in however small a quantity–the potential of infection. Ancient DNA alone cannot itself demonstrate the onset of disease. The amplified sequences from a small number of bacterial cells will appear similar to the amplified sequence from a heavy infestation. The lipids, however, are not amplified and, judging from the Newcastle Infirmary, are detectable when the bacterium is rife within the bone. Neither the presence of the bacterium nor even intense infection need lead to bone disorders. Indeed, the impaired breathing from damaged lungs can end life before any change to the bones has occurred. Lesions within them indicate the disease within a living person, coping with increasing disablement but staying alive for a while at least. In the St Helen’s cemetery in York, only one of almost 1,200 bodies studied had visible signs of tuberculosis in his or her bones. If we were able to return to the bodies armed with a range of molecular analyses we could no doubt place that individual in the wider context of a larger population experiencing the disease at its many different stages.

  Mark Spigelman and Angela Gernaey brought their two approaches together via a trace of mineral deposited within an otherwise healthy looking skeleton. Be’er Sheva University had been excavating a Byzantine church in the Negev Desert in Israel when they encountered a 1,400 year old burial. As they examined the skeleton, specifically its ribs, they noticed a limy residue, corresponding to the place the pleural cavity had been. Mark Spigelman recognized this as a fragment of calcified lung pleura, which would suggest that the buried individual had been infected with tuberculosis. No one had ever examined such material for biomolecular evidence, and Spigelman was taken by the idea of bringing together DNA and lipid approaches to test his hypothesis. The calcified fragment came up positive on both counts. It had retained both the DNA and the mycolipids specifically associated with Mycobacterium tuberculosis.

  enemies or friends?

  Micro-organisms have played a major role in human prehistory as our most significant predators. As agriculture has substantially raised the human population so, from time to time, have our microbial predators felled it. Yet not all microbes are as bad for us as the title of this chapter might suggest. The surfaces of our bodies, and in particular our mouths and guts, are full of microscopic wildlife without which our health would become very poor. They aid in digestion, protection against unwelcome microbes, and the provision of some nutrients. The flora of our gut is as much a cultural artefact as that of our back garden, and largely a consequence of what has been fed into it. Different culinary traditions in different parts of the world have tended to generate regional differences in the human gut flora. Having caught a glimpse of the inhabitants of the saliva applied in some manner to the Marchioness of Vasca’s left arm, Franco Rollo realized that he could probe yet further. He could examine these ancient floras, and get some sense of how they had evolved through time.

  He started probing into another mummified body, that of a young woman from the ancient Inca capital at Cuzco in Peru. The body had withstood the centuries with many internal organs intact. The stomach, lungs, intestines, liver and heart were all recognizable. Rollo’s team took samples from the muscle of the heart and from the large intestine, and prepared them for amplification of a short section of the 16S rRNA gene. After various modifications of their method, they came up with a protocol that worked, and generated a series of DNA amplification products that could be cloned, sequenced, and matched to a wide variety of contemporary gut bacteria.

  The commonest types of bacteria encountered were of
the genus Clostridium. This is a widespread genus of rod-shaped bacteria, found in soil and water as well as in the gut. Some species lead to diseases such as botulism and tetanus, but the species within the Cuzco woman were largely benign members of her gut flora. This dominance of Clostridium was probably not, however, a fair reflection of the wide range of microbes that would have originally colonized her gut, but one sign of a level of persistence in certain microbes that is truly remarkable.

  life in old fossils?

  Clostridium species today display a considerable resistance to heat, desiccation, toxic chemicals and detergents. This is because they are among the bacteria able to form spores, one of the most resilient resting stages known in the living world. At the heart of these spores is a condensed package of the bacteria’s DNA, bound together with short protein molecules. This condensed package is enclosed by several layers of protective coating, which shield their precious cargo from heat, light and chemical damage. Not much is known about how long these tiny capsules of genetic information can last, but the bias in the Cuzco woman’s intestine towards this genus is very likely a consequence of the survival powers of these spores. That power to survive could, however, take us very much further back than the ancient Inca empire, perhaps to the oldest fossils probed in the quest for ancient DNA.

  The first three years of the 1990s was a heady period in that quest. The oldest dates rolled back millions of years at a time, from the Magnolia leaf (12-17 million years) and the insects in amber (20-30 million years) to the weevil in amber (130 million years). At California Polytechnic State University, Raul Cano was one of the scientists keeping the world abreast of these amazing discoveries. He was as keen as others to get his work published fast, but in late 1991 he encountered something so unusual he decided to remain silent on the matter until it had been checked and double-checked. What he and his student Monica Borucki had noticed concerned an insect in amber from which they were hoping to amplify DNA. The insect’s gut contained what looked like bacterial spores. Having sterilized the amber and everything around these spores, they transferred them to a sterile nutrient solution. They then witnessed something that is still hard to believe. A single-cell organism, trapped in amber for 30 million years, seemed to be coming back to life and growing.