Bacteriology at UW-Madison
In a healthy animal, the internal tissues, e.g. blood, brain,
etc., are normally free of microorganisms. However, the
tissues, i.e., skin and mucous membranes, are constantly in contact
environmental organisms and become readily colonized by various
species. The mixture of organisms regularly found at any anatomical
is referred to as the normal flora, except by researchers in
the field who prefer the term "indigenous
microbiota". The normal flora of humans consists of a
few eucaryotic fungi and protists, but bacteria
are the most numerous and obvious microbial components of the normal
|BACTERIUM||Skin||Conjunctiva||Nose||Pharynx||Mouth||Lower Intestine||Anterior urethra||Vagina|
|Staphylococcus epidermidis (1)||++||+||++||++||++||+||++||++|
|Staphylococcus aureus* (2)||+||+/-||+||+||+||++||+/-||+|
|Streptococcus mutans* (3)||+||++|
|Enterococcus faecalis* (4)||+/-||+||++||+||+|
|Streptococcus pneumoniae* (5)||+/-||+/-||+||+||+/-|
|Streptococcus pyogenes* (6)||+/-||+/-||+||+||+/-||+/-|
|Neisseria sp. (7)||+||+||++||+||+||+|
|Neisseria meningitidis* (8)||+||++||+||+|
|Enterobacteriaceae* (Escherichia coli) (9)||+/-||+/-||+/-||+||++||+||+|
|Pseudomonas aeruginosa* (10)||+/-||+/-||+||+/-|
|Haemophilus influenzae* (11)||+/-||+||+||+|
|Bifidobacterium bifidum (12)||++|
|Lactobacillus sp. (13)||+||++||++||++|
|Clostridium sp.* (14)||+/-||++|
|Clostridium tetani (15)||+/-|
(2) Many of the normal flora are either
pathogens, The asterisks indicate members of the normal flora a that
be considered major pathogens of humans.
S. aureus. Gram stain.
(3) Streptococcus mutans is the primary
plaque formation and initiation of dental caries. Viewed as an
infection, dental disease is one of the most prevalent and costly
diseases in the United States.
Streptococcus mutans. Gram stain. CDC
(4) Enterococcus faecalis was formerly
classified as Streptococcus
faecalis. The bacterium is such a regular a component of the
flora, that many European countries use it as the standard indicator of
fecal pollution, in the same way we use E. coli in the
In recent years, Enterococcus faecalis has emerged as a
antibiotic-resistant, nosocomial pathogen.
Vancomycin Resistant Enterococcus faecalis. Scanning E.M. CDC
(5) Streptococcus pneumoniae is present in
tract of about half the population. If it invades the lower
tract it can cause pneumonia. Streptococcus pneumoniae
95 percent of all bacterial pneumonia.
Streptococcus pneumoniae. Direct fluorescent antibody stain. CDC.
(6) Streptococcus pyogenes refers to the
streptococci. Streptococci cause tonsillitis (strep throat), pneumonia,
endocarditis. Some streptococcal diseases can lead to rheumatic fever
or nephritis which can damage the heart and kidney.
Streptococcus pyogenes. Gram stain.
(7) Neisseria and other Gram-negative
frequent inhabitants of the upper respiratory tract, mainly the
meningitidis, an important cause of bacterial meningitis, can
as well, until the host can develop active immunity against the
Neisseria meningitidis. Gram stain.
(8) While E. coli is a consistent resident
of the small
many other enteric bacteria may reside here as well, including Klebsiella,
Enterobacter and Citrobacter. Some strains of E.
pathogens that cause intestinal infections, urinary tract infections
E. coli. Scanning E.M. Shirley Owens. Center for Electron Optics. Michigan State University.
(9) Pseudomonas aeruginosa is the
pathogen of humans that can invade virtually any tissue. It is a
leading cause of hospital-acquired (nosocomial) Gram-negative
but its source is often exogenous (from outside the host).
Colonies of Pseudomonas aeruginosa growing on an agar plate. The most virulent Pseudomonas species produce mucoid colonies and green pigments such as this isolate.
(10) Haemophilus influenzae is a frequent
viral influenza, and was named accordingly. The bacterium was the
leading cause of meningitis in infants and children until the recent
of the Hflu type B vaccine.
Haemophilus influenzae. Gram stain.
(11) The greatest number of bacteria are found in
tract, specifically the colon and the most prevalent bacteria are the Bacteroides,
a group of Gram-negative, anaerobic, non-sporeforming bacteria.
have been implicated in the initiation colitis and colon cancer.
Bacteroides fragilis. Gram stain.
(12) Bifidobacteria are Gram-positive,
acid bacteria. They have been described as "friendly" bacteria in the
intestine of humans. Bifidobacterium bifidum is the predominant
bacterial species in the intestine of breast-fed infants, where it
presumably prevents colonization by potential pathogens. These bacteria
are sometimes used in the manufacture of yogurts and are frequently
incorporated into probiotics.
Bifidobacterium bifidum. Gram stain
(13) Lactobacilli in the oral cavity
probably contribute to
formation that leads to dental caries. Lactobacillus
colonizes the vaginal epithelium during child-bearing years and
the low pH that inhibits the growth of pathogens.
Lactobacillus species and a vaginal squaemous epithelial cell. CDC
(14) There are numerous species of Clostridium
the bowel. Clostridium perfringens is commonly isolated
feces. Clostridium difficile may colonize the bowel and
"antibiotic-induced diarrhea" or pseudomembranous colitis.
Clostridium perfringens. Gram stain.
(15) Clostridium tetani is included in the
table as an
of a bacterium that is "transiently associated" with humans as a
of the normal flora. The bacterium can be isolated from feces in
0 - 25 percent of the population. The endospores are probably
ingested with food and water, and the bacterium does not colonize the
Clostridium tetani. Gram stain.
(16) The corynebacteria, and certain related propionic acid bacteria, are consistent skin flora. Some have been implicated as a cause of acne. Corynebacterium diphtheriae, the agent of diphtheria, was considered a member of the normal flora before the widespread use of the diphtheria toxoid, which is used to immunize against the disease.
Most members of the normal bacterial flora prefer to colonize
certain tissues and not others. This "tissue specificity" is
usually due to properties of both the host and the bacterium. Usually,
specific bacteria colonize specific tissues by one or another of these
1. Tissue tropism is the bacterial preference or predilection for certain tissues for growth. One explanation for tissue tropism is that the host provides essential nutrients and growth factors for the bacterium, in addition to suitable oxygen, pH, and temperature for growth.
|Bacterium||Bacterial adhesin||Attachment site|
|Streptococcus pyogenes||Cell-bound protein (M-protein)||Pharyngeal epithelium|
bound protein (Glycosyl transferase)
||Pellicle of tooth|
|Streptococcus salivarius||Lipoteichoic acid||Buccal epithelium of tongue|
protein (choline-binding protein)
|Staphylococcus aureus||Cell-bound protein||Mucosal epithelium|
|Neisseria gonorrhoeae||N-methylphenylalanine pili||Urethral/cervical epithelium|
|Enterotoxigenic E. coli||Type-1 fimbriae||Intestinal epithelium|
|Uropathogenic E. coli||P-pili (pap)||Upper urinary tract|
|Bordetella pertussis||Fimbriae ("filamentous hemagglutinin")||Respiratory epithelium|
|Vibrio cholerae||N-methylphenylalanine pili||Intestinal epithelium|
|Treponema pallidum||Peptide in outer membrane||Mucosal epithelium|
|Mycoplasma||Membrane protein||Respiratory epithelium|
|Chlamydia||Unknown||Conjunctival or urethral epithelium|
A human first becomes colonized by a normal flora at the moment of birth and passage through the birth canal. In utero, the fetus is sterile, but when the mother's water breaks and the birth process begins, so does colonization of the body surfaces. Handling and feeding of the infant after birth leads to establishment of a stable normal flora on the skin, oral cavity and intestinal tract in about 48 hours.It has been calculated that a human adult houses about 1012 bacteria on the skin, 1010 in the mouth, and 1014 in the gastrointestinal tract. The latter number is far in excess of the number of eucaryotic cells in all the tissues and organs which comprise a human. The predominant bacteria on the surfaces of the human body are listed in Table 3. Informal names identify the bacteria in this table. Formal taxonomic names of organisms are given in Table 1.
||staphylococci and corynebacteria
||sparse, Gram-positive cocci and
||streptococci and lactic
|Upper respiratory tract
|nares (nasal membranes)||staphylococci and corynebacteria|
Gram-negative rods and cocci
|Lower respiratory tract
pylori (up to 50%)
||lactics, enterics, enterococci,
||bacteroides, lactics, enterics,
enterococci, clostridia, methanogens
||lactic acid bacteria during
child-bearing years; otherwise mixed
Normal Flora of the Skin The
human is covered with approximately 2 square meters of skin. The
and composition of the normal flora of the skin varies with anatomical
The high moisture content of the axilla, groin, and areas between the
supports the activity and growth of relatively high densities of
cells, but the density of bacterial populations at most other sites is
fairly low, generally in 100s or 1000s per square cm. Most bacteria on
the skin are sequestered in sweat glands.
The skin microbes found in the most
layers of the epidermis and the upper parts of the hair follicles are
Gram-positive cocci (Staphylococcus epidermidis and Micrococcus
sp.) and corynebacteria such as Propionibacterium
sp. These are generally nonpathogenic and
to be commensal, although mutualistic and parasitic roles have been
to them. For example, staphylococci and propionibacteria produce fatty
acids that inhibit the growth of fungi and yeast on the
skin. But, if
Propionibacterium acnes, a
normal inhabitant of the skin, becomes
trapped in hair follicle, it may grow rapidly and cause inflammation
Figure 4. Colonies of Propionibacterium acnes, found on skin and the conjunctiva.
Normal Flora of the Respiratory Tract A large number of bacterial species colonize the upper respiratory tract (nasopharynx). The nares (nostrils) are always heavily colonized, predominantly with Staphylococcus epidermidis and corynebacteria, and often (in about 20% of the general population) with Staphylococcus aureus, this being the main carrier site of this important pathogen. The healthy sinuses, in contrast are sterile. The pharynx (throat) is normally colonized by streptococci and various Gram-negative cocci. Sometimes pathogens such as Streptococcus pneumoniae, Streptococcus pyogenes, Haemophilus influenzae and Neisseria meningitidis colonize the pharynx.
The lower respiratory tract
and pulmonary tissues) is virtually free of microorganisms,
mainly because of the efficient cleansing action of the ciliated
which lines the tract. Any bacteria reaching the lower respiratory
are swept upward by the action of the mucociliary blanket that lines
bronchi, to be removed subsequently by coughing, sneezing, swallowing,
etc. If the respiratory tract epithelium becomes damaged, as in
or viral pneumonia, the individual may become susceptible to infection
by pathogens such as H. influenzae or
pneumoniae descending from the nasopharynx.
Normal Flora of the Urogenital Tract Urine is normally sterile, and since the urinary tract is flushed with urine every few hours, microorganisms have problems gaining access and becoming established. The flora of the anterior urethra, as indicated principally by urine cultures, suggests that the area my be inhabited by a relatively consistent normal flora consisting of Staphylococcus epidermidis, Enterococcus faecalis and some alpha-hemolytic streptococci. Their numbers are not plentiful, however. In addition, some enteric bacteria (e.g. E. coli, Proteus) and corynebacteria, which are probably contaminants from the skin, vulva or rectum, may occasionally be found at the anterior urethra.
The vagina becomes colonized soon after birth with corynebacteria, staphylococci, streptococci, E. coli, and a lactic acid bacterium historically named "Doderlein's bacillus" (Lactobacillus acidophilus). During reproductive life, from puberty to menopause, the vaginal epithelium contains glycogen due to the actions of circulating estrogens. Doderlein's bacillus predominates, being able to metabolize the glycogen to lactic acid. The lactic acid and other products of metabolism inhibit colonization by all except this lactobacillus and a select number of lactic acid bacteria. The resulting low pH of the vaginal epithelium prevents establishment by most other bacteria as well as the potentially-pathogenic yeast, Candida albicans. This is a striking example of the protective effect of the normal bacterial flora for their human host.
figure 5. A Lactobacillus species, possibly Doderlein's bacillus, in association with a vaginal epithelial cell.
Normal Flora of the Oral Cavity The presence of nutrients, epithelial debris, and secretions makes the mouth a favorable habitat for a great variety of bacteria. Oral bacteria include streptococci, lactobacilli, staphylococci and corynebacteria, with a great number of anaerobes, especially bacteroides.
The mouth presents a succession of different ecological situations with age, and this corresponds with changes in the composition of the normal flora. At birth, the oral cavity is composed solely of the soft tissues of the lips, cheeks, tongue and palate, which are kept moist by the secretions of the salivary glands. At birth the oral cavity is sterile but rapidly becomes colonized from the environment, particularly from the mother in the first feeding. Streptococcus salivarius is dominant and may make up 98% of the total oral flora until the appearance of the teeth (6 - 9 months in humans). The eruption of the teeth during the first year leads to colonization by S. mutans and S. sanguis. These bacteria require a nondesquamating (nonepithelial) surface in order to colonize. They will persist as long as teeth remain. Other strains of streptococci adhere strongly to the gums and cheeks but not to the teeth. The creation of the gingival crevice area (supporting structures of the teeth) increases the habitat for the variety of anaerobic species found. The complexity of the oral flora continues to increase with time, and bacteroides and spirochetes colonize around puberty.
Figure 6. Various streptococci in a biofilm in the oral cavity.
The normal bacterial flora of the oral cavity clearly benefit from
their host who provides nutrients and habitat. There may be
benefits, as well, to the
host. The normal flora occupy available colonization sites which
it more difficult for other microorganisms (nonindigenous species) to
established. Also, the oral flora contribute to host nutrition through
the synthesis of vitamins, and they contribute to immunity by inducing
low levels of circulating and secretory antibodies that may cross react
with pathogens. Finally, the oral bacteria exert microbial antagonism
nonindigenous species by production of inhibitory substances such as
peroxides and bacteriocins.
On the other hand, the oral flora are the usual cause of various
in humans, including abscesses, dental caries, gingivitis, and
periodontal disease. If oral bacteria can gain entrance into
deeper tissues, they may cause abscesses of alveolar bone, lung,
brain, or the extremities. Such infections usually contain mixtures of
with Bacteroides melaninogenicus often playing a dominant role.
If oral streptococci are introduced into wounds created by dental
manipulation or treatment,
they may adhere to heart valves and
subacute bacterial endocarditis.
Flora of the
Figure 7. Colonies of E. coli growing on EMB agar.
The bacterial flora of the gastrointestinal (GI) tract of animals
has been studied more
extensively than that of any other site. The composition differs
various animal species, and within an animal species. In humans, there
are differences in the composition of the flora which are influenced by
age, diet, cultural conditions, and the use of antibiotics. The
greatly perturbs the composition of the intestinal flora.
In the upper GI tract of adult humans, the esophagus contains only
bacteria swallowed with saliva and food. Because of the high acidity of
the gastric juice, very few bacteria (mainly acid-tolerant
can be cultured from the normal stomach. However, at least half
population in the United States is colonized by a pathogenic bacterium,
pylori. Since the 1980s, this bacterium has been known to be
the cause of gastric ulcers, and it is probably a cause of gastric and
duodenal cancer as well. The Australian microbiologist, Barry Marshall,
received the Nobel Prize in Physiology and Medicine in 2005, for
demonstrating the relationship between Helicobacter and gastric
Figure 8. Helicobacter pylori. ASM
The proximal small intestine has a relatively sparse Gram-positive
consisting mainly of lactobacilli and Enterococcus faecalis.
This region has about 105 - 107 bacteria per ml
fluid. The distal part of the small intestine contains greater numbers
of bacteria (108/ml) and additional species, including
coliforms (E. coli and
and Bacteroides, in addition to lactobacilli and enterococci.
flora of the large intestine (colon) is qualitatively similar to that
in feces. Populations of bacteria in the colon reach levels of 1011/ml
feces. Coliforms become more prominent, and enterococci, clostridia and
lactobacilli can be regularly found, but the predominant species are
and anaerobic lactic acid bacteria in the genus Bifidobacterium
(Bifidobacterium bifidum). These organisms may outnumber E.
by 1,000:1 to 10,000:1. Sometimes, significant numbers of
methanogens (up to 1010/gm) may reside in the
of humans. This is our only direct association with archaea as normal
flora. The range of incidence of certain bacteria in the large
of humans is shown in Table 4 below.
|BACTERIUM||Range of Incidence|
It is in the intestinal tract that we see the greatest effect of the
bacterial flora on their host. This is due to their large mass and
numbers. Bacteria in the human GI tract have been shown to produce
vitamins and may otherwise contribute
to nutrition and digestion. But their most important effects are in
their ability to protect their host from establishment and
alien microbes and their ability to stimulate the development and the
activity of the immunological tissues.
On the other hand, some of the bacteria in the colon (e.g. Bacteroides) have been shown to
produce metabolites that are carcinogenic, and there may be an
increased incidence of colon cancer associated with these bacteria.
Alterations in the GI flora brought on by poor nutrition or perturbance
with antibiotics can cause shifts in populations and colonization by
nonresidents that leads to gastrointestinal disease.
Dental plaque, which is material adhering to the teeth, consists of bacterial cells (60-70% the volume of the plaque), salivary polymers, and bacterial extracellular products. Plaque is a naturally-constructed biofilm, in which the consortia of bacteria may reach a thickness of 300-500 cells on the surfaces of the teeth. These accumulations subject the teeth and gingival tissues to high concentrations of bacterial metabolites, which result in dental disease.
The dominant bacterial species in dental plaque are Streptococcus sanguis and Streptococcus mutans, both of which are considered responsible for plaque.
Streptococcus mutans. Gram stain. CDC.
Plaque formation is
initiated by a weak attachment of the
cells to salivary glycoproteins forming a pellicle on the surface of
teeth. This is followed by a stronger attachment by means of
sticky polymers of glucose (glucans) which are synthesized by the
from dietary sugars (principally sucrose). An enzyme on the cell
of Streptococcus mutans,
glycosyl transferase, is involved
initial attachment of the bacterial cells to the tooth surface and in
conversion of sucrose to dextran polymers (glucans) which
Dental Caries is the destruction
of the enamel, dentin or cementum of teeth due to bacterial activities.
Caries are initiated by direct demineralization of the enamel of teeth
due to lactic acid and other organic acids which accumulate in dental
Lactic acid bacteria in the plaque produce lactic acid from the
of sugars and other carbohydrates in the diet of the host.
mutans and Streptococcus
sanguis are most consistently been associated with the
caries, but other lactic acid bacteria are probably involved as well.
organisms normally colonize the occlusal fissures and contact points
the teeth, and this correlates with the incidence of decay on these
Cross section of a
tooth illustrating the various structural regions susceptible to
colonization or attack by microbes.
Streptococcus mutans in particular has a number of physiological and biochemical properties which implicate it in the initiation of dental caries.
2. It contains a cell-bound protein, glycosyl transferase, that serves an adhesin for attachment to the tooth, and as an enzyme that polymerizes dietary sugars into glucans that leads to the formation of plaque.
3. It produces lactic acid from the utilization of dietary carbohydrate which demineralizes tooth enamel. S. mutans produces more lactic acid and is more acid-tolerant than most other streptococci.
4. It stores polysaccharides made from dietary sugars which can be utilized as reserve carbon and energy sources for production of lactic acid. The extracellular glucans formed by S. mutans are, in fact, bacterial capsular polysaccharides that function as carbohydrate reserves. The organisms can also form intracellular polysaccharides from sugars which are stored in cells and then metabolized to lactic acid.
Periodontal Diseases are bacterial infections that affect the supporting structures of the teeth (gingiva, cementum, periodontal membrane and alveolar bone). The most common form, gingivitis, is an inflammatory condition of the gums. It is associated with accumulations of bacterial plaque in the area. Increased populations of Actinomyces have been found, and they have been suggested as the cause.Diseases that are confined to the gum usually do not lead to loss of teeth, but there are other more serious forms of periodontal disease that affect periodontal membrane and alveolar bone resulting in tooth loss. Bacteria in these lesions are very complex populations consisting of Gram-positive organisms (including Actinomyces and streptococci) and Gram-negative organisms (including spirochetes and Bacteroides). The mechanisms of tissue destruction in periodontal disease are not clearly defined but hydrolytic enzymes, endotoxins, and other toxic bacterial metabolites seem to be involved.