Prokaryote classification and diversity

Different groups of prokaryotes. Evolutionary relationships of bacteria and archaea.

Extremophiles.
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Key points:

 The two prokaryote domains, Bacteria and Archaea, split from each other early in the evolution
of life.
 Bacteria are very diverse, ranging from disease-causing pathogens to beneficial
photosynthesizers and symbionts.
 Archaea are also diverse, but none are pathogenic and many live in extreme environments.
 A DNA sequencing approach called metagenomics lets scientists identify new species of
bacteria and archaea, including ones that can't be cultured.

Introduction

Prokaryotes, which include both bacteria and archaea, are found almost everywhere – in every
ecosystem, on every surface of our homes, and inside of our bodies! Some live in environments
too extreme for other organisms, such as hot vents on the ocean floor.
_Image credit: "Black smoker in Atlantic ocean," by P. Rona (public domain)._
Although they are found all around us, prokaryotes can be hard to detect, count, and classify. The
prokaryotic species we know of today are a tiny fraction of all prokaryotic species thought to
exist.^11start superscript, 1, end superscript In fact, the very idea of a ―species‖ becomes
complicated in the world of prokaryotes!
In this article, we'll first look at major groups of prokaryotes. Then, we'll explore why it’s often
tricky to identify and classify them. Finally, we'll see how DNA sequencing methods are helping
us get a better picture of the prokaryotes around us.
A prokaryote "family tree"

For a long time, all prokaryotes were classified into a single domain (the largest taxonomic
grouping).
However, work by microbiologist Carl Woese in the 1970s showed that prokaryotes are divided
into two distinct lineages, or lines of descent: Archaea and Bacteria. Today, these groups are
considered to form two out of three domains of life. The third domain (Eukarya) includes all
eukaryotes, such as plants, animals, and fungi.^22start superscript, 2, end superscript
[How did Woese figure this out?]
^3start superscript, 3, end superscript

This phylogeny (evolutionary tree) depicts the evolutionary relationships between the three
domains of life: Eukarya, Archaea, and Bacteria. The two prokaryotic domains (Archaea and
Bacteria) each comprise several smaller taxonomic groupings. Within the Archaea are the
euryarchaeotes, crenarchaeotes, nanoarchaeotes, and korarchaeotes. Within the Bacteria are
proteobacteria, chlamydias, spirochetes, cyanobacteria, and gram-positive bacteria.
Image credit: "Structure of prokaryotes: Figure 3," by OpenStax College, Biology (CC BY 3.0).
Since splitting off from one another millions of years ago, both Bacteria and Archaea have split
off into many groups and species.

Bacteria

Domain Bacteria contains 555 major groups: proteobacteria, chlamydias, spirochetes,

cyanobacteria, and gram-positive bacteria.
The proteobacteria are subdivided into five groups, alpha through epsilon. Species in these
groups have a wide range of lifestyles. Some are symbiotic with plants, others live in hot vents
deep under the sea, and others yet cause human diseases, such as stomach ulcers (Helicobacter
pylori) and food poisoning (Salmonella).
 Characteristics of the five phyla of bacteria are described. The first phylum described is
proteobacteria, which includes five classes, alpha, beta, gamma, delta and epsilon. Most species
of Alpha Proteobacteria are photoautotrophic but some are symbionts of plants and animals, and
others are pathogens. Eukaryotic mitochondria are thought be derived from bacteria in this group.
Representative species include Rhizobium, a nitrogen-fixing endosymbiont associated with the
roots of legumes, and Rickettsia, obligate intracellular parasite that causes typhus and Rocky
Mountain Spotted Fever (but not rickets, which is caused by Vitamin D deficiency). A
micrograph shows rod-shaped Rickettsia rickettsii inside a much larger eukaryotic cell.
Beta Proteobacteria is a diverse group of bacteria. Some species play an important role in the
nitrogen cycle. Representative species include Nitrosomas, which oxidize ammonia into nitrate,
and Spirillum minus, which causes rat bite fever. A micrograph of spiral-shaped Spirillum minus
is shown.
Gamma Proteobacteria include many are beneficial symbionts that populate the human gut, as
well as familiar human pathogens. Some species from this subgroup oxidize sulfur compounds.
Representative species include Escherichia coli, normally beneficial microbe of the human gut,
but some strains cause disease; Salmonella, certain strains of which cause food poisoning, and
typhoid fever; Yersinia pestis–the causative agent of Bubonic plague; Psuedomonas aeruganosa–
causes lung infections; Vibrio cholera, the causative agent of cholera, and Chromatium–sulfur
producing bacteria bacteria that oxidize sulfur, producing H2S. Micrograph shows rod-shaped
Vibrio cholera, which are about 1 micron long.
Some species of delta Proteobacteria generate a spore-forming fruiting body in adverse
conditions. Others reduce sulfate and sulfur. Representative species include Myxobacteria, which
generate spore-forming fruiting bodies in adverse conditions and Desulfovibrio vulgaris, an
aneorobic, sulfur-reducing bacterium. Micrograph shows a bent rod-shaped Desulfovibrio
vulgaris bacterium with a long flagellum.
Epsilon Proteobacteria includes many species that inhabit the digestive tract of animals as
symbionts or pathogens. Bacteria from this group have been found in deep-sea hydrothermal
vents and cold seep habitats.
The next phylum described is chlamydias. All members of this group are obligate intracellular
parasites of animal cells. Cells walls lack peptidoglycan. Micrograph shows a pap smear of cells
infected with Chlamydia trachomatis. Chlamydia infection is the most common sexually
transmitted disease and can lead to blindness.
All members of the phylum Spirochetes have spiral-shaped cells. Most are free-living anaerobes,
but some are pathogenic. Flagella run lengthwise in the periplasmic space between the inner and
outer membrane. Representative species include Treponema pallidum, the causative agent of
syphilis and Borrelia burgdorferi, the causative agent of Lyme disease Micrograph shows
corkscrew-shaped Trepanema pallidum, about 1 micron across.
Bacteria in the phylum Cyanobacteria, also known as blue-green algae, obtain their energy
through photosynthesis. They are ubiquitous, found in terrestrial, marine, and freshwater
environments. Eukaryotic chloroplasts are thought be derived from bacteria in this group. The
cyanobacterium Prochlorococcus is believed to be the most abundant photosynthetic organism on
 earth, responsible for generating half the world’s oxygen. Micrograph shows a long, thin rod-
shaped species called Phormidium.
Gram-positive Bacteria have a thick cell wall and lack an outer membrane. Soil-dwelling
members of this subgroup decompose organic matter. Some species cause disease. Representative
species include Bacillus anthracis, which causes anthrax; Clostridium botulinum, which causes
botulism; Clostridium difficile, which causes diarrhea during antibiotic therapy; Streptomyces,
from which many antibiotics, including streptomyocin, are derived; and Mycoplasmas, the
smallest known bacteria, which lack a cell wall. Some are free-living, and some are pathogenic.
Micrograph shows Clostridium difficile, which are rod-shaped and about 3 microns long.
_Image credit: "Structure of prokaryotes: Figure 4," by OpenStax College, Biology, CC BY 4.0.
Original work credits: ―Rickettsia rickettsia‖: modification of work by CDC; credit ―Spirillum
minus‖: modification of work by Wolframm Adlassnig; credit ―Vibrio cholera‖: modification of
work by Janice Haney Carr, CDC; credit ―Desulfovibrio vulgaris‖: modification of work by
Graham Bradley; credit ―Campylobacter‖: modification of work by De Wood, Pooley, USDA,
ARS, EMU; scale-bar data from Matt Russell._
The other four major groups of bacteria are similarly diverse. Chlamydias are pathogens that live
inside host cells, while cyanobacteria are photosynthesizers that make much of Earth's oxygen.
Spirochetes include both harmless bacteria and harmful ones, like the Borrelia burgdorferi that
cause Lyme disease. The same is true of Gram-positive bacteria, which range from probiotic
bacteria in yogurt to the Bacillus anthracis that cause anthrax.^44start superscript, 4, end
superscript
 Chlamydia, Spirochetes, Cyanobacteria, and Gram-positive bacteria are described in this table.
Chlamydias: all members of this group are obligate intracellular parasites of animal cells. Cell
walls lack peptidoglycan. Representative organism: Chlamydia trachomatis, common sexually
transmitted disease that can lead to blindness. Representative micrograph: in this pap smear,
Chlamydia trichomatis appear as pink inclusions inside cells.
Spirochetes: Most members of this species, which has spiral-shaped cells, are free-living
anaerobes, but some are pathogenic. Flagella run lengthwise in the periplasmic space between the
inner and outer membrane. Representative organisms: Treponema pallidum, causative agent of
syphilis, and Borrelia burgdorferi, causative agent of Lyme disease. Representative micrograph:
Treponema pallidum, a corkscrew-shaped bacterium.
Cyanobacteria: also known as blue-green algae, these bacteria obtain their energy through
photosynthesis. They are ubiquitous, found in terrestrial, marine, and freshwater environments.
Eukaryotic chloroplasts are thought to be derived from bacteria in this group. Representative
organism: Prochlorococcus, believed to be the most abundant photosynthetic organism on earth;
responsible for generating half the world's oxygen. Representative micrograph: Phormidium, a
long, thin, rod-shaped bacterium.
Gram-positive bacteria: soil-dwelling members of this subgroup decompose organic matter. Some
species cause disease. They have a thick cell wall and lack an outer membrane. Representative
organisms: Bacillus anthracis, causes anthrax; Clostridium botulinum, causes botulism;
Clostridium difficile, causes diarrhea during antibiotic therapy; Streptomyces, many antibiotic,
including streptomycin, are derived from these bacteria; and Mycoplasmas, tiny bacteria, the
smallest known, lacking a cell wall. Some are free-living, and some are pathogenic.
Representative micrograph: Clostridium dificile, a rod-shaped bacterium.
_Image credit: "Structure of prokaryotes: Figure 5," by OpenStax College, Biology, CC BY 4.0.
Original image credits: ―Chlamydia trachomatis‖: modification of work by Dr. Lance Liotta
Laboratory, NCI; credit ―Treponema pallidum‖: modification of work by Dr. David Cox, CDC;
credit ―Phormidium‖: modification of work by USGS; credit ―Clostridium difficile‖:
modification of work by Lois S. Wiggs, CDC; scale-bar data from Matt Russell._

Archaea

Domain Archaea contains 444 major groups. Intriguingly, so far, no archaea that are human
pathogens have yet been discovered.
Archaea do live in our bodies and those of animals—for instance, in the gut—but all of them
seem to be harmless or beneficial. Although there are hypotheses, no one yet knows exactly why
archaea are all "friendly," i.e., why no disease-causing species have evolved.^55start superscript,
5, end superscript
Alongside the archaea that enjoy the comfy environment of the human gut, there are
many extremophile species that live in much more inhospitable places. These include volcanic
hot springs, undersea hot vents, and very salty places like the Dead Sea.
 Characteristics of the four phyla of archaea are described. Euryarchaeotes includes methanogens,
which produce methane as a metabolic waste product, and halobacteria, which live in an extreme
saline environment. Methanogens cause flatulence in humans and other animals. Halobacteria can
grow in large blooms that appear reddish, due to the presence of bacterirhodopsin in the
membrane. Bacteriorhodopsin is related to the retinal pigment rhodopsin. Micrograph shows rod-
shaped Halobacterium. Members of the ubiquitous Crenarchaeotes phylum play an important role
in the fixation of carbon. Many members of this group are sulfur-dependent extremophiles. Some
are thermophilic or hyperthermophilic. Micrograph shows cocci-shaped Sulfolobus, a genus
which grows in volcanic springs at temperatures between 75° and 80°C and at a pH between 2
and 3. The phylum Nanoarchaeotes currently contains only one species, Nanoarchaeum equitans,
which has been isolated from the bottom of the Atlantic Ocean, and from the a hydrothermal vent
at Yellowstone National Park. It is an obligate symbiont with Ignococcus, another species of
archaebacteria. Micrograph shows two small, round N. equitans cells attached to a larger
Ignococcus cell. Korarchaeotes are considered to be one of the most primitive forms of life and so
far have only been found in the Obsidian Pool, a hot spring at Yellowstone National Park.
Micrograph shows a variety of specimens from this group which vary in shape.
_Image credit: "Structure of prokaryotes: Figure 6," by OpenStax College, Biology, CC BY 4.0.
Original image credits: ―Halobacterium‖: modification of work by NASA; credit
―Nanoarchaeotum equitans‖: modification of work by Karl O. Stetter; credit ―korarchaeota‖:
modification of work by Office of Science of the U.S. Dept. of Energy; scale-bar data from Matt
Russell._
[More about extremophiles]

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The many "mystery prokaryotes"

For many years, the main approach to studying prokaryotes was to grow them in the lab. If an
organism could be grown on an agar plate or in a liquid culture, then it could be studied,
analyzed, and added to our growing catalog of prokaryotic species and strains.
Some prokaryotes, however, can't grow in a laboratory setting (at least, not under the conditions
scientists have tried). In fact, an estimated 99\%99%99, percent of bacteria and archaea are
unculturable!

Two bacterial plates with red agar are shown. Both plates are covered with bacterial colonies. On
the right plate, which contains hemolytic bacteria, the red agar has turned clear where bacteria are
growing. On the left plate, which contains non-hemolytic bacteria, the agar is not clear.
In these agar plates, the growth medium is supplemented with red blood cells. Blood agar
becomes transparent in the presence of hemolytic Streptococcus bacteria, as shown on the plate
at right. _Image credit:Prokaryotic diversity: Figure 6, by OpenStax College, Biology, (CC BY
4.0). Original image by Bill Branson, NCI._
This represents a pretty huge gap in our understanding of what prokaryotes are out there. For
context, there are 8.78.78, point, 7 \text{million}millionm, i, l, l, i, o, n known eukaryotic
species^66start superscript, 6, end superscript. If the culturability problem applied to eukaryotes
in the same degree as prokaryotes, we would only know of 87,87,87, comma000000000 of these
species. This would make for a very empty tree of life, and a very incomplete understanding of
what eukaryotes (as a group) are like. For instance, we might know that there were animals, but
be in the dark about plants or fungi!
What is a prokaryotic species?

In order to talk about finding prokaryotic species, we probably need to define what they are. This
may seem like a basic question, but it's a complex and even controversial one if you're a
microbiologist.
For eukaryotes, most scientists define a species as a group of organisms that can interbreed and
have fertile offspring. This definition makes sense for species that reproduce sexually, but it
doesn't work so well for organisms like bacteria. Bacteria reproduce asexually to make clones of
themselves—they don't interbreed.
Scientists instead classify bacteria and archaea into taxonomic groups based on similarities in
appearance, physiology, and genes.^77start superscript, 7, end superscript Many are given names
using traditional Linnean taxonomy, with a genus and species. Still, the question of how and
whether prokaryotes should be grouped into species remains a topic of debate among scientists.
The right ―species concept‖ for these organisms is still a work in progress.^88start superscript, 8,
end superscript

Metagenomics: A new window on microbes

Scientists estimate there may be millions of prokaryotic species (or species-like groups), but we
know very little about most of them.^11start superscript, 1, end superscript This is starting to
change thanks to large-scale DNA sequencing.
DNA sequencing makes it possible for scientists to study entire prokaryotic communities in their
natural habitats – including the many prokaryotes that are unculturable, and would previously
have been "invisible" to researchers.
The collective genome of such a community is called its metagenome, and the analysis of
metagenome sequences is known as metagenomics. Prokaryotic metagenomics is one of the
areas of biology that I find coolest and most mysterious.
For example, a DNA sample can be taken from a hot spring microbial mat, such as the beautiful,
multicolored mats found in Yellowstone National Park. Even a tiny sample from this rich
community includes many, many individuals of different species.^99start superscript, 9, end
superscript
Image credit: "Bacteria mat," by sevenblock CC BY-NC-SA 2.0.
By sequencing and analyzing metagenome DNA samples, scientists can sometimes piece
together entire genomes of previously unknown species. In other cases, they use sequence
information from specific genes to figure out what types of prokaryotes are present (and how
they are related to each other or to known species). The genes found in the DNA samples can
also provide clues about the metabolic strategies of the organisms in the
community.^{10}10start superscript, 10, end superscript
[Attribution and references]