Allies and Enemies: How the World Depends on Bacteria (29 page)

chapter 7 · climate, bacteria, and a barrel of oil

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In 1996, NASA added fuel to the “bacteria on Mars” fire when it

announced that a meteorite that had crashed near Antarctica 13,000

years ago held traces of bacterial growth. Scientists named the meteorite ALH 84001 and recovered it in 1984. By the early 1990s,

NASA scientists believed it had blasted off from Mars and traveled

interplanetary space for 16 billion years. Astrobiologists, meanwhile,

focused on tiny wormlike structures embedded in the rock that

resembled fossilized microbes. Analysis of the structures’ elements suggested that the structures were more of biological origin than geological. With prior discovery of ancient rivers and seas on Mars’ surface, science seemed to hold circumstantial evidence of water and life on the Red Planet.

The analysis of Mars’ atmosphere has also provided evidence of

methane. Considering that Earth’s methane is almost 95 percent of

biological origin, the presence of this gas on Mars has been viewed by

some astrobiologists as another point in favor of life on Mars. Earth’s atmospheric methane on a volume basis is 1,750 parts per billion, but that of Mars is only 10 parts per billion. No one knows why the dis—

parity exists between the two planets or if any of Mars’ methane came

from living things.

Another research team examined the meteorite’s mineral content

and found magnetite crystals similar to the magnetosomes in Earth’s

Aquaspirllium magnetotacticum. Dennis Bazylinsky of the University of Nevada-Las Vegas has been studying magnetotactic bacteria for more than 20 years. When reviewing the meteorite data, he found the magnetic crystals in the meteorite to be identical to the crystals made by Earth’s magnetic bacteria. Again, scientists held circumstantial evidence in their hands, but the task of comparing Earth’s magnetic bacteria to extraterrestrial crystals would not be easy. Very few cultures of magnetic bacteria exist in laboratories worldwide. New strains are known to exist in nature but they live in difficult-to-reach marine sediments.

Naysayers to life on Mars have pointed out that methane and

inorganic structures may resemble conditions on Earth but can also

be explained in nonbiological terms, which is true. Microbiologists have questioned the meteorite’s “worm holes” because they are much smaller than the smallest Earth bacteria and thus unlikely to contain

all the molecules needed for life. Of course, those scientists are

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speaking about Earth life, which may be a bit egocentric considering

the size of the universe. By 2000, however, most astrobiologists concluded that the worm holes were probably fossilized debris, organic debris perhaps, but not microbial.

Conclusive studies on nanobacteria on Earth may recharge the

life-on-Mars debate. Finnish researcher Olavi Kajander discovered nanobacteria in 1988, but the majority of microbiologists rejected the idea of their existence. (Notice how every new discovery mentioned

in this book endured a period of denial?) More than a decade of study

on nanobacteria suggested that these microbes played a pathogenic

role in arterial and kidney calcification.

By 2005, literature had accumulated on Nanobacterium sanguineum, a gram-negative motile species with a calcium-coated outer shell. The bacterium measures only 20 to 200 nm across, but it is big enough to contain 16S rRNA. Studies on N. sanguineum have followed a similar path as studies in the Golden Age of Microbiology: The medical importance of the microbe has superseded environmental studies. But nanobiology will very soon be part of the growing sci—

ence in interplanetary biology.

Shaping the planet

The Earth’s biosphere consists of millions of ecosystems. When

ecosystems interrelate, they form large ecosystem communities. The

Earth thus has grassland, rainforest, polar communities, and so on.

Although members interact at boundaries called edges, such as the

interface between marine and shore life, many of Earth’s communities remain separated by distance. Migrating herds and birds connect some communities but not all of them at once. Only bacteria connect

all of Earth’s communities by the constant recycling of nutrients through soil, the oceans, and the atmosphere.

No one needs a degree in microbiology to find these microbes all

around and performing their life-giving activities. In the country, notice the lichen growing on rocks, dead leaves decomposing underfoot, and the glimpses of color when a lake ripples. If you live in the city, bacteria live all around. Biofilms coat storm drains and metal-metabolizing bacteria weather bridges and buildings. Soot in the air chapter 7 · climate, bacteria, and a barrel of oil

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carries bacteria from block to block. It is easier to detect the invisible universe than it is to find places having no bacteria.

You may never look at your surroundings the same way again, and

that is a good thing. Appreciating bacteria is the best way to acknowl—

edge the larger community of Earth. When I was in college in the 1970s, I realized microbiology is a hard subject. It encompasses the basics of cell biology, covers chemical and biochemical reactions, touches on the Earth sciences, and is intimately connected to genetics. Microbiology recruits only scientists willing to study organisms they cannot see. But it is impossible to delve deeper into the microbial world without seeing that bacteria run this planet. Humans reap the benefits of bacterial actions when they discard garbage, avoid infection (remember the skin’s good bacteria), and simply breathe.

Bacteria should not be synonymous with disease. Making cheese

out of milk also seems to sell these microbes short. Because of bacteria, our lives are richer, healthier, and more hopeful. Hopeful because no matter what predicament humanity puts itself in, there is a very good chance that a bacterium somewhere can solve the problem.

Stop worrying about germs and start appreciating bacteria. Few

pathogenic bacteria exist that cannot be stopped by simply washing

hands, preparing food properly, and steering clear of others who are

obviously sick. As for the good bacteria that fill the environment, we

need not nurture them because they grow just fine without any help

from humans. In the process, bacteria supply us with the nutrients we

need. Bacteria shape the planet and they also shape us.

For safety’s sake, thinking of bacteria as occasional enemies as well as constant allies helps maintain your health. In the bigger picture, however, bacteria are your best friends. They welcomed humanity into their home tens of thousands of years ago, and they will stay with you to the end. Bacteria work behind the scenes to protect us,

feed us, and decompose our wastes. I cannot think of a better ally than bacteria.

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Epilogue: How microbiologists

grow bacteria

Spending a career working with invisible objects can stretch a person’s patience. Bacteria demand that a microbiologist wait several hours, overnight, or even several days before multiplying to high numbers. Mycobacterium takes three weeks to reach numbers high enough to study the organism. Finally, a person cannot call herself a microbiologist without mastering the art of aseptic technique. The technique truly is an art because no two bacterial cultures behave exactly the same way, and the avenues for contamination seem limit—

less. The standard practices described here avoid some of the pitfalls

that students new to microbiology see when growing bacteria.

Microbiology samples may be patient specimens (blood, sputum,

stool, and so on), foods, consumer products, soil, drinking water, untreated surface waters, or wastewater. Microbiologists usually take samples of 100 milliliters liquid or 10 grams of a solid to a laboratory for “processing,” which is the series of steps needed to determine if bacteria are in the sample, how many, and what kinds.

Microbiology employs two aids for working with the huge numbers common in this field. First, microbiologists dilute samples containing millions or billions of cells in a technique called serial dilution. Second, the microbiologist converts these large numbers to logarithms.

Serial dilution

A sample containing more than a million bacteria per milliliter or gram is too concentrated with cells for scientists to study a species 165

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and draw meaningful conclusions. Rather than juggle numbers of this

magnitude, microbiologists dilute each sample sequentially to reduce

the cell concentration to between 30 and 300 cells per milliliter.

The method called serial dilution consists of a set of tubes each

containing 9.0 milliliters of sterile buffer (water with a small amount of salts to maintain a constant pH range). By taking one milliliter of the sample and adding it to one of the 9-milliliter tubes, the microbiologist has made a one-to-ten dilution, or 1:10. With this step, a sample containing three million cells per milliliter now contains 300,000. A milliliter of this new dilution transferred to another sterile 9-milliliter volume lowers the concentration to 30,000 cells per milliliter. The microbiologist continues diluting each new dilution until arriving at what he assumes is a much lower concentration than the original sample. This is tricky because the process is done on supposition. When microbiologists receive a sample from a patient, food, or the environment, they have no idea if the sample contains millions of bacteria or a few. Serial dilution helps span the range of possible concentrations to determine the actual concentration of cells in a sample.

Following the serial dilution, the microbiologist has a set of tubes

before him, each tube containing one-tenth as many cells as the preceding tube. The next step involves inoculating agar plates with small volumes, called aliquots, from each dilution. The microbiologist might take a 0.1-milliliter aliquot from each dilution and put this amount onto individual sterile agar plates. For example, 0.1 milliliter of the 1:10 dilution goes onto a plate (microbiologists usually include duplicate or triplicate plates for each dilution), 0.1 milliliter of the 1:100 dilution does the same, and so on. After all of these transfers have been completed, the microbiologist has a set of inoculated plates each containing a subsample (the aliquot) from the 1:10, 1:100, 1:1,000, 1:10,000, and 1:100,000 dilutions.

Next, the microbiologist spreads each of the aliquots over the agar

surface to spread out whatever bacteria may be there—remember,

they are invisible. This spreading step requires a sterile glass or plastic rod about seven inches long with a bend at one end about an inch from the end of the rod. Visualize a hockey stick shape. These spreaders are in fact called “hockey sticks” by microbiologists. When the aliquot has been spread as a thin transparent film over the agar surface, the agar is epilogue · how microbiologists grow bacteria

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called a spread plate. Each plate comes with a cover, which now goes

onto the inoculated spread plate.

The scientist puts the entire stack of spread plates into an incubator

set at a favorable temperature. Although a stack of agar plates in an

incubator seems an obvious space-saving arrangement, this innovation

of German bacteriologist J. R. Petri in 1887 changed microbiology. The

stackable, compact Petri dishes enabled microbiologists to study more

replicates and a wider variety of microbes than in previous experiments.

Most bacteria recovered from temperate environments grow at

body temperature, so incubators can be set to about 98.6°F (37°C)

for the incubation step. Many foodborne contaminants and almost all

pathogens and native flora prefer this temperature. Soil and water microbes and some foodborne psychrophiles grow better at lower

temperatures.

Incubation lasts overnight, a day or two, or several days to weeks,

depending on the bacterium. After incubation of the plates, the microbiologist sees visible colonies, usually no bigger than one-eighth of an inch in diameter, each containing millions of bacteria.

Counting bacteria

A colony of bacteria growing on agar contains identical cells that have all descended from a single ancestor cell. When a microbiologist inoculates agar, individual bacteria disperse in the medium. During incubation, each cell from the inoculum doubles in number every half hour or so, depending on species, until they form the visible mass of cells

known as a colony. Microbiologists call the colonies CFUs for colony-forming units and count them either manually under a magnifying glass or electronically by scanning the agar plate with a laser beam.