Despite 300 years of peering through microscopes, growing bacteria in culture, and screening soil, air, and water for new microbial species, scientists have clearly overlooked much of life on earth. Thanks to powerful new research tools, bacteriologists are discovering that the living world is suddenly far bigger and more complex than they imagined even a decade ago. The finding is similar in magnitude, perhaps, to Dutch microscopist Antonie van Leeuwenhoek’s first glimpse of microbes-he called them “animalcules”-cavorting beneath his crude glass lenses.

Because bacteria were essentially undetectable unless they could be grown in culture, most species-which exist in hard-to-replicate environments-have been, in a sense, off limits. “Imagine if our entire understanding of biology were based on visits to zoos,” says Norman R. Pace, a biologist at University of California, Berkeley. “That is analogous to our situation with respect to understanding the microbial world.”

Melvin Simon, chair of the department of biology at the California Institute of Technology, estimates that “about 99 percent of the bacteria out there can’t yet be grown” in laboratory culture dishes. That’s because scientists often can’t determine the precise combination of conditions-including oxygen, temperature, and light-that microbes need and because it’s difficult to simulate these requirements all at once to make them happy. About 6,000 species of microorganisms have been formally described. And until now, he says, the rest have remained “invisible” to us.

The key to gaining a first glimpse at this new world was in learning how to isolate, amplify, and study an organism’s genetic material directly from the environment without culturing the organism itself. By using molecular instruments that selectively pick out a bacterium’s highly specific genes from a broth of DNA, scientists can now quickly identify new organisms, explore their properties, and determine their kinship to organisms that are already known.

Prominent among the new molecular tools is the gene-amplifying technique called PCR, the polymerase chain reaction. PCR allows specific genes to be isolated and copied endlessly, so enough become available for detailed study, such as of a gene’s DNA sequence. And it’s in the sequence-actually the differences in sequences from one organism to another-that the microbes’ identities can be seen.

Along with PCR, the new tools include rapid sequencing technology, quick computer processing of gene-sequence data, and increasingly accurate electrophoresis technology, which electrically separates genes in a gel or fluid according to size.

One of the major analytical targets of such work is the ubiquitous RNA (ribonucleic acid) in ribosomes, the little rounded bodies that living cells use as protein-building devices. All cells have ribosomes, and because the RNA they contain differs slightly from one species to the next, it serves as a reliable marker-sort of a molecular dog tag-that can be used for identifying, classifying, and analyzing each new microbe.

The surprising finding is that everywhere researchers apply these new techniques, they are encountering strange new bacteria, in abundances and diversities that no one really anticipated. For example, as Paul Dunlap, an associate scientist at the Woods Hole Oceanographic Institution in Massachusetts, explains in the Woods Hole publication Oceanus, the sea’s microbial life is so incredibly abundant that in some cases organisms occur “in extraordinarily high numbers, up to about 100 billion per milliliter” of seawater. In fact, organisms less than 2 microns in diameter found living in the sea are so abundant they account for 95 percent of the mass of all forms of life in the world’s oceans.

Sedimentary soils, even the ancient mud deposited on the deep-sea floor, are also vastly alive with microorganisms. Recent drilling into the floor of the Sea of Japan, for example, brought up samples-from layers of sediment buried 500 meters deep-containing millions of organisms in every gram of soil.

New Realms

The findings suggest that even a tiny sip of water, or a small sample of soil, carries enough bacteria-most of them previously unknown-to keep squads of researchers busy for years. “There are new genes, new enzymes, and new functions we don’t know anything about,” Simon notes, and the data are piling up too fast to be analyzed in detail. “They (the microbes) even have some different kinds of cell membranes,” suggesting that whole new realms of biology are there to be explored.

Since each organism has its own genes, characteristic biochemistry, structure, and behavior, the industrial potential is huge. Each species harbors between 1,000 and 5,000 genes, and for the most part their biochemical properties are still unknown. So it is likely that new enzymes, foods, drugs, chemicals, materials, and processes will be discovered. Bacteria that can live in the hostile environment of hot springs, for example, have yielded special enzymes that perform beautifully at high temperature, which could be of use in chemical processing. Also, Pace says, some microorganisms make substances that adhere to surfaces in water that is almost boiling, which could be valuable in certain manufacturing operations.

In evolutionary terms, Simon notes, it should come as no surprise that so many different kinds of bacteria exist, even in places where they once seemed unlikely. After all, “for 3 billion years the world was populated by bacteria” and probably very little else. So these single-cell organisms have had far more time to evolve than most other creatures on earth.