The true cave paintings at Lascaux have been closed to the public because of microbial damage. By measuring light absorption of biofilms in the catacombs of San Callisto, researchers have found that blue lights (top left) should inhibit the growth of the cyanobacteria (bottom left) at the heart of the colony. Credit: P. ALBERTANO

Every year some 300,000 people visit the world-famous Lascaux cave in southern France to see its prehistoric paintings. But those who are not forewarned are liable to be disappointed: the cave they enter is only a replica, created 200 metres away from the original. The real cave was closed to the public in 1963, after it was found that a combination of carbon dioxide from visitors' breath and microbes on the rock surface were devouring the artwork. Lascaux is not alone. One after another, the doors to subterranean cultural treasures across Europe have been shut.

Such drastic measures have long seemed to be the only way to preserve these sites, as cleaning the walls with disinfectants can seriously damage the delicate paintings. But Lascaux and places like it could eventually be reopened as a result of a European research programme aimed at understanding the ecology of the paint-eating microbes. As it turns out, it is much more effective to be subtle. Where harsh chemicals have failed, rays of blue or green light are succeeding.

Lascaux closed when patches of algae and moss began to spread across the walls, the result of local climate changes caused by the intense flow of visitors and strong illumination. Even back then, the cave attracted more than 1,000 people per day, all curious to see the ‘Sistine Chapel of its time’ with its 17,000-year-old paintings. Once sealed, the cave's climate and its art returned to their original state. Today the air and paintings are monitored by computer-controlled sensors.

Under attack

Over the years, archaeologists and microbiologists have discovered a huge variety of microbes attacking subterranean monuments. Despite the nutrient-poor conditions, colonies of algae, mosses, bacteria and fungi all find ways to survive. Conservators have tried using fungicides, bactericides and quicklime to control them, but with limited success. “To throw chemicals crudely at the problem is not ideal because they can cause their own damage,” says Patrizia Albertano, a biologist at the University of Rome ‘Tor Vergata’ and coordinator of the European research programme.

But gentler microbicides that target specific types of organism can also be ineffective because they just shift the balance to other species within the colony. Unlike the simple layer of mould that sometimes grows in a bathroom, the colonies that invade newly opened subterranean sites develop into complex communities of interdependent species. The only way to tackle these effectively without harsh treatments is to research the exact constituents. “We need to understand the biology of the biodiversity as a whole and work out how best to control growth and damage,” says Albertano. “This isn't easy as the range of microbes is so great.”

Albertano's team, made up of ten groups from six European countries, has focused on the cause of most of the damage: biofilms, complex mats of interacting microorganisms. The scientists selected three study sites where the ambient conditions are typical of many famous archaeological remains in southern Europe. Two are in Rome — the catacombs of Domitilla and San Callisto with their third-century frescoes, which are among the earliest known Christian paintings. The other is the Cave of Bats in Zuheros, southern Spain, which contains remarkable geological formations and Palaeolithic rock paintings.

Eaten away

In the Roman catacombs, the researchers were faced with extensive patches of green on the frescoes. They found that the dominant organisms in these biofilms were cyanobacteria. Through their ability to photosynthesize, the cyanobacteria were supporting the growth of a great variety of bacteria and fungi, thus accelerating the spread of the biofilm. The acids produced by the other members of the colony were causing ‘biocorrosion’, a normal phenomenon along rocky coastlines. “But in this case,” says Albertano, “it was destroying the frescoes.”

Considered to be the ancestors of plants, cyanobacteria depend on sunlight. Yet they can get by in places with poor light, and are found in very dimly lit caves. When their underground dwellings are flooded with artificial light, the bacteria begin to grow wildly. The same is true of algae — which the team found in the Cave of Bats — and mosses. Such photosynthetic species are the anchors for the entire biofilm, and the by-products of their metabolism feed all manner of microbes in the colony.

Light, the team realized, might be the biofilms' Achilles' heel. To find out, the researchers used a spectroradiometer, which identifies the wavelengths of light that a surface absorbs and reflects. The cyanobacteria in the catacombs absorbed light from the full visible spectrum except for a narrow band in the blue region. In theory, if the catacombs were lit with only that blue light, the microbes should stop growing.

Albertano's team tested the idea in a chamber of San Callisto called the Cubiculum of the Ocean. Bulbs emitting the correct wavelength were hard to find, but the group eventually got some from a company that supplies lights to nightclubs. Under the blue light, the biofilm's growth rate has decreased noticeably (P. Albertano et al. in Molecular Biology and Cultural Heritage 151–162; Balkema, Lisse, 2003). But Albertano says she is reserving judgement on the long-term effectiveness of the technique until sometime this year.

The same approach was taken at the Cave of Bats, where a species of green algae is the anchor for the biofilm. Like all green plants, their colour is a reflection — literally — of the wavelengths of light they cannot use. So the cave is now bathed in pure green light. “We certainly see that green algae and also mosses grow more slowly under green light and eventually die,” says Mariona Hernández-Mariné, a pharmacologist at the University of Barcelona and a member of the team.

Do visitors mind having to squint through monochromatic light? Apparently not, according to a questionnaire given out by the team at San Callisto. Most tourists said that they preferred the coloured light to the alternative of reducing the number of visitors.

Community service

But monochromatic light won't work at all sites. In some caves, biofilms trap nutrients carried down from the surface by seeping groundwater. And in caves that have both cyanobacterial and moss-fed biofilms, neither blue nor green light is a total solution. In such places, it may be possible to target chemical agents essential to biofilm growth.

One possibility is a group of compounds called siderophores, which are used by bacteria to absorb iron, a nutrient they need to produce essential enzymes. The researchers reason that the judicious application of siderophores produced in the lab could soak up iron before the bacteria can make use of it, and so inhibit their growth. Such an approach is being tested by VTT Biotechnology near Helsinki, Finland, as a way to reduce salmonella infections in chicken houses.

Another option is to interfere with communication between bacterial cells. Bacteria can grow and divide individually without doing too much damage to the rocks. They only form destructive biofilms when their population density reaches a certain threshold. The cells detect the surrounding population by sensing the concentration of secreted molecules called AHLs, or N-acylhomoserine lactones. A chemical method that blocks or destroys AHLs could prevent a biofilm from forming at all.

Such chemical methods are still under development and have yet to be tried in the caves. But if they work, they could be combined with pure light to allow archaeological sites currently sealed off for protection to reopen to the public. “These biocleaning procedures may develop into a valuable alternative to bactericides and fungicides, and would be less dangerous for the health of researchers and visitors,” Albertano says.

Will visitors get a second chance to see the original Lascaux cave? The San Callisto catacombs have remained open to the public, and the Cubiculum remains illuminated by pure blue light. Conservators at Lascaux declined to answer Nature's enquiries about the possibility of using the method there. But perhaps as blue light and biocleansers begin to prove their merit at other underground monuments, tourists will get their chance to see the real version of this ancient site.

→ http://www2.bio.uniroma2.it/lab/algae/CATS.htm