THEME:
II. Human microflora and methods of its studying.
Dysbacteriosis, causes of its appearance.
Microflora of the environment
Microbes are distributed everywhere in the environment surrounding us.
They are found in the soil, water, air, in plants, animals, food products,
various utensils, in the human body, and on the surface of the human body.
The relationship of micro-organisms with the environment has been named ecology (Gr.
oikos home, native land, logos
idea, science). This is an-adaptive relationship. Micro-organisms have a
remarkable ability to adapt themselves to certain environmental conditions.
Individual ecology, population ecology, and association community ecology are
distinguished. The study of micro-organism ecology forms the basis for
understanding parasitism, zoonotic diseases, and particularly diseases with
natural nidality, as well as for elaborating measures for the control of
various infectious diseases.
Soil Microflora
Types of Microorganisms
in Soil
Living organisms both plants
and animals, constitute an important component of soil. The pioneering
investigations of a number of early microbiologists showed for the first time
that the soil was not an inert static material but a medium pulsating with
life. The soil is now believed to be a dynamic or rather a living system,
containing a dynamic population of organisms/microorganisms. Cultivated soil
has relatively more population of microorganisms than the fallow land, and the
soils rich in organic matter contain much more population than sandy and eroded
soils. Microbes in the soil are important to us in maintaining soil fertility /
productivity, cycling of nutrient elements in the biosphere and sources of
industrial products such as enzymes, antibiotics, vitamins, hormones, organic
acids etc. At the same time certain soil microbes are the causal agents of
human and plant diseases.
The soil organisms are broadly classified in to two groups viz soil flora and soil fauna, the detailed
classification of which is as follows.
Soil Organisms
A. Soil Flora
a)
Microflora: 1.
Bacteria 2. Fungi, Molds, Yeast, Mushroom 3. Actinomycetes, Stretomyces 4.
Algae eg. BGA, Yellow Green Algae, Golden Brown Algae.
1. Bacteria is
again classified in I) Heterotrophic eg. symbiotic & non -
symbiotic N2 fixers, Ammonifier, Cellulose Decomposers, Denitrifiers II)
Autrotrophic eg. Nitrosomonas, Nitrobacter, Sulphur oxidizers, etc.
b) Macroflora: Roots of higher plants
B. Soil Fauna
a) Microfauna: Protozoa, Nematodes
b) Macrofauna: Earthworms. moles, ants & others.
- Macroflora: Vascular
plants, Mosses, etc.
- Microflora: Bacteria, Actinomycetes, Fungi, Algae
Influences on Microbial Activity
- Temperature (70°-100°F
most active microbes)
- Moisture (Field capacity
is optimal)
- Aeration (want a nice mix
of pores filled with water and air)
- pH (optimal pH is 6-7)
- Soil organic matter
Soil Microorganisms:
Bacteria
1. Most numerous in soil
2. Most diverse metabolism
3. Can be aerobic or anaerobic
4. Optimal growth at pH 6-8
5.
Examples: Nitrosomonas and Nitrobacter
in nitrification processes, N2 fixers, fire blight is caused by a bacterium
Soil Microorganisms: Actinomycetes
1.
Transitional group between bacteria and fungi
2.
Active in degrading more resistant organic compounds
3. Optimal growth at alkaline pH
4.
2 important products:
– produce antibodies (streptomycin is produced by an actino)
– produce geosmin
5.
Negative impact - potato scab (Streptomyces scabies)
Soil Microorganisms: Fungi
1. Dominate the soil biomass
2. Obligate aerobes
- Can survive desiccation
- Dominate in acid soils
- Negative impacts:
– Apple replant disease (Rhizoctonia, Pythium, Fusarium, and Phytophtora)
– Powdery mildew is caused by a fungus - Beneficials:– Penicillium
Nematode-trapping Fungus
Plant root / Soil / Microbial Interactions
Beneficial
- Symbiotic associations
such as that found with Rhizobia (N2 fixing bacteria, ex. legumes)
- Fungi-mycorrhizal associations:
important for water and P uptake; also improves soil structure
- Earthworm channels:
improve permeability and aeration
Deleterious
- Agrobacterium (bacteria)
cause gall formation in plants
- Fungi causing soil-borne
plant rot diseases
- Rhizoctonia and Pythium
(involved with replant disease)
Soil
Nutrient Cycling
- Materials are broken down
by macro and meso-fauna
- Nutrients are taken up
and converted by lower life forms in the soil
- They convert these nutrients
to organic forms within the cell or to inorganic forms released to soil
- These organisms die and
are decomposed by other organisms
- This also releases
inorganic ions for plant or other microbe uptake and…
- The cycle starts all over
Nitrogen Cycle: Nitrogen is the
nutrient needed in largest amounts by plants and is the most commonly applied
fertilizer. Excess N can have negative affects on plant growth and crop quality
as well as harming the environment, especially water quality.
Nitrogen is present
in one of five forms in soil:
- Organic N: 90% of N is in
organic form. It must be mineralized to become available.
- Ammonium N (NH4+):
Inorganic, soluble form
- Nitrate (NO3-): Inorganic, soluble form
- Atmospheric N (N2):
80% of atmosphere but unavailable to most plants except N-fixers
- Nitrite (NO2-):
only under anaerobic conditions. This form is toxic to plants and normally
will not be present in significant amounts in soil.
N: Nutrient cycling and composting
Soil science was founded by V. Dokuchaev, P. Kostychev. S. Vinogradsky,
V. Williams, and others. Soil fertility depends not only on the presence of
inorganic and organic substances, but also on the presence of various species
of micro-organisms which influence the qualitative composition of the soil. Due
to nutrients and moisture in the soil the number of microbes in
Soil
microflora
Soil microflora consists algae
nitrifying nitrogen-fixing, denitrifying, cellulose-splitting and sulfur bacteria, pigmented microbes
fungi, protozoa, etc.
The blue-green algae play an important part in enrichment of the soil
with nitrogen. The extent to which the soil is contaminated with microbes
depends on its nature and chemical composition (Table 1).
The greatest amount of microbes (1 000000 per cu cm) is found in the top
layer of soil at a depth of 5-
Table 1
Total Amount of Microbes in Different Soils according
to the Direct Counting Method
|
Kind of soil |
Number of
microbes |
Number of spores
in |
|
Clayey podsol |
801 800000 |
4000 |
|
|
1219000000 |
12000 |
|
Chernozem |
4771000000 |
100000-180000 |
|
|
2854000000 |
200000-400000 |
|
Light soil |
2661 100000 |
700000 |
|
Loose sand |
904000000 |
600000-1200000 |
|
Gray soil |
896000000 |
750000-1500000 |
Oil' bacteria live in oil wells. Using paraffins (distillates of oil) as
nutrients, they turn part of the oil into a thick asphalt-like mass with the
formation of which natural oil reservoirs become clogged up. It has been
calculated that in the ploughed layer of cultivated soil over an area of
The number of microorganisms in the soil depends on the extent of
contamination with faeces and urine, and also on the nature of treating and
fertilizing the soil. For example, ploughed soil contains 2.5 times more
microbes than forest soil.
Saprophytic
spores (B. cereus. B, meguterium,
etc.) survive for long periods in the
soil.
Pathogenic bacteria which do not
produce spores due to lack of essential nutrients, and also as a result of the
lethal activity of light, drying, antagonistic microbes, and phages do not live
long in the soil (from a few days to a few months) (Table 2).
Usually the soil is an unfavourable
habitat for most pathogenic species of bacteria, rickettsiae, viruses, fungi,
and protozoa. The survival period of some pathogenic bacteria is shown in Table
2. However, the soil as a factor of transmitting a number of causative agents
of infectious diseases is quite a complex substrate. Thus, for example, anthrax
bacilli after falling on the soil produce spores which can remain viable for
many years. In favourable conditions (in dark brown soil and chernozem) they
pass through the whole cycle of development: during the summer months the
spores germinate into the vegetative forms and then this cycle is repeated.
Table 2
Survival Period
of Pathogenic Bacteria in the Soil
|
Species of
bacteria |
Average
period |
Maximal
period |
|
Salmonella typhi |
2-3 |
12 |
|
Shigella |
1,5-5 |
9 |
|
Vibrio cholerae |
1-2 |
4 |
|
Vibrio cholerae El Tor |
4 |
6 |
|
Mycobacterium tuberculosis |
13 |
7 |
|
Brucella |
0,5-3 |
2 |
|
Yersnia pestis |
0,5 |
1 |
|
Francisella tularensis |
1,5 |
2,5 |
As is known, the spores of clostridia causing tetanus, anaerobic infections,
and botulism, and of many soil microbes survive for long periods in the soil.
The soil is the habitat for various animals (rodents) which are parasitized by
the carriers of the causative agents of plague, tularaemia, the viruses of
mosquito fever, haemorrhagic fever, encephalitis, agricultural leishmaniasis,
etc. The cysts, of intestinal
protozoa (amoeba, balantidium, etc.) spend a certain stage in the soil. The
soil plays an important role in transmitting worm invasions (ascarids,
hook-worms, nematode worms, etc.). Some fungi live in the soil. Entering the
body they cause fusariotoxicosis, ergotism, aspergillosis, penicilliosis
mucormycosis, etc.
Taking into consideration the
definite epidemiological role played by the soil in spreading some infectious
diseases of animals and man, sanitary-epidemiological practice involves
measures directed at protecting the soil from pollution and infection with
pathogenic species of microorganisms.
S. Vinogradsky, V. Omeliansky, N.
Kholodny and others devised a method of
investigating soil microbes and used the results obtained in
agriculture.
A valuable index of the sanitary condition of the soil is the discovery
of the colibacillus and related bacteria, also enterococci, and Clostridium perfringens. The presence of
the latter indicates an earlier faecal contamination.
Microflora
of the Water
Water microbiology is concerned with
the microorganisms
that live in water,
or can be transported from one habitat to
another by water.
Water can
support the growth of many types of microorganisms. This can be advantageous.
For example, the chemical activities of certain strains of yeasts provide us
with beer and bread. As well, the growth of some bacteria in contaminated water can
help digest the poisons from the water.
However,
the presence of other disease causing microbes in water is
unhealthy and even life threatening. For example, bacteria that live in the
intestinal tracts of humans and other warm blooded animals, such as Escherichia coli, Salmonella,
Shigella, and Vibrio, can contaminate water if
feces enters the water. Contamination of drinking water
with a type of Escherichia coli known as O157:H7 can be fatal. The
contamination of the municipal water supply of Walkerton, Ontario, Canada in
the summer of 2000 by strain O157:H7 sickened 2,000 people and killed seven
people.
The
intestinal tract of warm-blooded animals also contains viruses that can
contaminate water and cause disease. Examples include rotavirus, enteroviruses,
and coxsackievirus.
Another
group of microbes of concern in water microbiology are protozoa. The two protozoa of the most
concern are Giardia and Cryptosporidium. They live normally in
the intestinal tract of animals such as beaver and deer. Giardia
and Cryptosporidium form dormant and hardy forms called cysts during
their life cycles. The cyst forms are resistant to chlorine, which is the most popular
form of drinking water disinfection, and can pass through the filters used in
many water treatment plants. If
ingested in drinking water they can cause debilitating and prolonged diarrhea
in humans, and can be life threatening to those people with impaired immune
systems. Cryptosporidium contamination of the drinking water of
Milwaukee, Wisconsin with in 1993 sickened more than 400,000 people and killed
47 people.
Many
microorganisms are found naturally in fresh and saltwater. These include
bacteria, cyanobacteria, protozoa, algae, and tiny animals such as
rotifers. These can be important in the food chain that forms the basis of life
in the water. For example, the microbes called cyanobacteria can convert the energy of the sun into the energy it
needs to live. The plentiful numbers of these organisms in turn are used as
food for other life. The algae that thrive in water is also an important food
source for other forms of life.
A variety
of microorganisms live in fresh water. The region of a water body near the
shoreline (the littoral zone) is well lighted, shallow, and warmer than other
regions of the water. Photosynthetic algae and bacteria that use light as energy thrive in this
zone. Further away from the shore is the limnitic zone. Photosynthetic microbes
also live here. As the water deepens, temperatures become colder and the oxygen concentration and light in
the water decrease. Now, microbes that require oxygen do not thrive. Instead,
purple and green sulfur bacteria, which can grow without
oxygen, dominate. Finally, at the bottom of fresh waters (the benthic zone),
few microbes survive. Bacteria that can survive in the absence of oxygen and
sunlight, such as methane producing bacteria, thrive.
Saltwater
presents a different environment to microorganisms. The higher salt concentration,
higher pH, and lower nutrients, relative to freshwater,
are lethal to many microorganisms. But, salt loving (halophilic) bacteria
abound near the surface, and some bacteria that also live in freshwater are
plentiful (i.e., Pseudomonas and Vibrio). Also, in 2001, researchers
demonstrated that the ancient form of microbial life known as archaebacteria is one of the
dominant forms of life in the ocean. The role of archaebacteria
in the ocean food chain is not yet known, but must be of vital importance
Another
microorganism found in saltwater are a type of algae known as
dinoflagellelates. The rapid growth and multiplication of dinoflagellates can
turn the water red. This "red tide" depletes the water of nutrients
and oxygen, which can cause many fish to die. As well, humans can
become ill by eating contaminated fish.
Water can
also be an ideal means of transporting microorganisms from one place to
another. For example, the water that is carried in the hulls of ships to
stabilize the vessels during their ocean voyages is now known to be a means of
transporting microorganisms around the globe. One of these organisms, a
bacterium called Vibrio cholerae, causes life threatening diarrhea in
humans.
Drinking
water is usually treated to minimize the risk of microbial contamination. The
importance of drinking water treatment has been known for centuries. For
example, in pre-Christian times the storage of drinking water in jugs made of metal was practiced. Now, the
anti-bacterial effect of some metals is known. Similarly, the boiling of
drinking water, as a means of protection of water has long been known.
Chemicals
such as chlorine or chlorine derivatives has been a popular means of killing
bacteria such as Escherichia coli in water since the early decades of
the twentieth century. Other bacteria-killing treatments that are increasingly
becoming popular include the use of a gas called ozone and the disabling of the
microbe's genetic material by the use of ultraviolet light. Microbes can also
be physically excluded form the water by passing the water through a filter.
Modern filters have holes in them that are so tiny that even particles as
miniscule as viruses can be trapped.
An
important aspect of water microbiology, particularly for drinking water, is the
testing of the water to ensure that it is safe to drink. Water quality testing
can de done in several ways. One popular test measures the turbidity of the
water. Turbidity gives an indication of the amount of suspended material in the
water. Typically, if material such as soil is present in the water then
microorganisms will also be present. The presence of particles even as small as
bacteria and viruses can decrease the clarity of the water. Turbidity is a
quick way of indicating if water quality is deteriorating, and so if action
should be taken to correct the water problem.
In many
countries, water microbiology is also the subject of legislation. Regulations
specify how often water sources are sampled, how the sampling is done, how the
analysis will be performed, what microbes are detected, and the acceptable
limits for the target microorganisms in the water sample. Testing for microbes
that cause disease (i.e., Salmonella typhymurium and Vibrio cholerae)
can be expensive and, if the bacteria are present in low numbers, they may
escape detection. Instead, other more numerous bacteria provide an indication
of fecal pollution of the water. Escherichia
coli has been used as an indicator of fecal pollution for decades. The
bacterium is present in the intestinal tract in huge numbers, and is more
numerous than the disease-causing bacteria and viruses. The chances of
detecting Escherichia coli is better than detecting the actual disease
causing microorganisms. Escherichia coli also had the advantage of not
being capable of growing and reproducing in the water (except in the warm and
food-laden waters of tropical countries). Thus, the presence of the bacterium
in water is indicative of recent fecal pollution. Finally, Escherichia coli
can be detected easily and inexpensively.
See also Chlorination; Oil spills; Sewage treatment; Water pollution.
Read more: Water Microbiology -
Bacteria, Microorganisms, Life, and Drinking - JRank Articles http://science.jrank.org/pages/7311/Water-Microbiology.html#ixzz2NVtehNPi
Read more: Water Microbiology -
Bacteria, Microorganisms, Life, and Drinking - JRank Articles http://science.jrank.org/pages/7311/Water-Microbiology.html#ixzz2NVtT1OJa
Pseudomonas fluorescens, Micrococcus
roseus, etc., are among the specific aquatic aerobic microorganisms. Anaerobic
bacteria are very rarely found in water.
The microflora of rivers depends on the
degree of pollution and the quality of purification of sewage waters flowing
into river beds. Micro-organisms are widespread in the waters of the seas and
oceans. They have been found at different depths (3700-
The degree of contamination of the water with organisms is expressed as saprobity which designates the total
of all living matter in water containing accumulations of animal and plant
remains. Water is subdivided into three zones. Polysaprobic zone is strongly polluted water, poor in oxygen and
rich in organic compounds. The number of bacteria in 1 ml reaches 1 000000 and
more. Colibacilli and anaerobic bacteria predominate which bring about the
processes of putrefaction and
fermentation. In the mesosaprobic
zone (zone of moderate pollution) the mineralization of organic substances with
intense oxidation and marked nitrification takes place. The number of bacteria
in 1 ml of water amounts to hundreds of thousands, and there is a marked
decrease in the number of colibacilli. The ohgosaprobic
zone is characteristic of pure water. The number of microbes is low, and in 1
ml there are a few tens or hundreds; this zone is devoid of the colibacillus.
Depending
on the degree of pollution pathogenic bacteria can survive in reservoirs and
for a certain time can remain viable. Thus, for example, in tap water, river,
or well water salmonellae of enteric
fever can live from 2 days to 3 months, shigellae — 5-9 days, leptospirae —
from 7 to 150 days. The cholera, vibrio El Tor lives in water for many months,
the causative agent of tularaemia — from a few days to 3 months.
Tap water is considered clean if it contains a total amount of 100
microbes per ml, doubtful if there are 100-150 microbes, and polluted if 500
and more are present. In well water and in open reservoirs the amount of
microbes in 1 ml should not exceed 1000. Besides, the quality of the water is
determined by the presence of E. coli
and its variants.
The degree of
faecal pollution of water is estimated by the colititre or coli-index. The colititre is the smallest amount of
water in millilitres in which one E. coli
bacillus is found. The coli-index is
the number of individuals off. coli
found in
Due to the fact that Str. faecalis
(enterococci) are constant inhabitants only of the intestine in man and
warm-blooded animals, and are highly resistant to temperature variations and
other environmental factors, they are taken into account with the colititre and
coli-index for the determination of the degree of faecal pollution of water,
sewage waters, soil, and other objects.
At present new standards of enterococcus indices are being worked out.
Water is an
important factor for the transmission of a number of infectious diseases
(enteric fever, paratyphoids, cholera, dysentery, leptospiroses, etc.).
Due to the enormous
sanitary-epidemiological role of water in relation to the intestinal group of
diseases, it became necessary to work out rapid indicator methods for revealing
colibacillus and pathogenic bacteria in water.
These include the methods of
luminescent microscopy for the investigation of water for the presence of
pathogenic microbes and the determination of the increase of the titre of the
phage. Upon the addition of specific phages to liquids containing a homologous
microbe in 6-10 hours a considerable increase in the amount of phage particles
can be observed.
For a more complete and profound study of the microflora of the soil and
water capillary microscopy is used.
The principle is that very thin capillary tubes are placed in the soil or water
reservoirs after which their contents are exposed to microscopic
investigations. This method reveals those species of micro-organisms which do
not grow in ordinary nutrient media, and which for many years were unknown to
microbiologists.
Microflora of the Air
Of all environments, air is the simplest one and it occurs in a single
phase gas. The relative quantities of various gases in air, by volume
percentage are nitrogen 78%, oxygen 21 %, argon 0.9%, carbon dioxide 0.03%,
hydrogen 0.01 % and other gases in trace amounts. In addition to various gases,
dust and condensed vapor may also be found in air
Various layers can be recognized in the atmosphere upto a height of about
1000km. The layer nearest to the earth is called as troposphere. In temperate
regions, troposphere extends upto about 11 km whereas in tropics up to about
16km. This troposphere is characterized by a heavy load of microorganisms. The
temperature of the atmosphere varies near the earth's surface. However, there
is a steady decrease of about 1 DC per 150m until the top of the troposphere.
Above the troposphere, the temperature starts to increase. The atmosphere as a
habitat is characterized by high light intensities, extreme temperature
variations, low amount of organic matter and a scarcity of available water
making it a non hospitable environment for microorganisms and generally
unsuitable habitat for their growth. Nevertheless, substantial numbers of
microbes are found in the lower regions of the atmosphere.
Microbes Found in Air- In addition to gases, dust particles
and water vapour, air also contains microorganisms. There are vegetative cells
and spores of bacteria, fungi and algae, viruses and protozoan cysts. Since air
is often exposed to sunlight, it has a higher temperature and less moisture.
So, if not protected from desiccation, most of these microbial forms will
die.Air is mainly it transport or dispersal medium for microorganisms. They
occur in relatively small numbers in air when compared with soil or water. The
microflora of air can be studied under two headings outdoor and indoor
microflora.
Sources of Microorganisms in Air - Although a number of
microorganisms are present in air, it doesn't have an indigenous flora. Air is
not a natural environment for microorganisms as it doesn't contain enough
moisture and nutrients to support their growth and reproduction.
Quite a number of sources have been studied in this connection and almost all
of them have been found to be responsible for the air microflora. One of the
most common sources of air microflora is the soil.
Soil microorganisms when disturbed by the wind blow, liberated into the air and
remain suspended there for a long period of time. Man made actions like digging
or plaguing the soil may also release soil borne microbes into the air.
Similarly microorganisms found in water may also be released into the air in
the form of water droplets or aerosols. Splashing of water by wind action or
tidal action may also produce droplets or aerosols. Air currents may bring the
microorganisms from plant or animal surfaces into air. These organisms may be
either commensals or plant or animal pathogens. Studies show that plant
pathogenic microorganisms are spread over very long distances through air. For
example, spores of Puccinia graminis travel over a thousand kilometers.
However, the transmission of animal diseases is not usually important in
outside air.
The main source of airborne microorganisms is human beings. Their surface flora
may be shed at times and may be disseminated into the air. Similarly, the
commensal as well as pathogenic flora of the upper respiratory tract and the
mouth are constantly discharged into the air by activities like coughing,
sneezing, talking and laughing.
The microorganisms are discharged out in three different forms which are
grouped on the basis of their relative size and moisture content. They are
droplets, droplet nuclei and infectious dust. It was Wells, who described the
formation of droplet nuclei. This initiated the studies on the significance of
airborne transmission. A brief description of these agents is given below
Droplets
Droplets are usually formed by
sneezing, coughing or talking. Each consists of saliva and mucus. Droplets may
also contain hundreds of microorganisms which may be pathogenic if discharged
from diseased persons. Pathogens will be mostly of respiratory tract origin.
The size of the droplet determines the time period during which they can remain
suspended.
Most droplets are relatively large, and they tend to settle rapidly in still
air. When inhaled these droplets are trapped on the moist surfaces of the
respiratory tract. Thus, the droplets containing pathogenic microorganisms may
be a source of infectious disease.
Droplet Nuclei
Small droplets in a warm, dry atmosphere tend to evaporate rapidly and
become droplet nuclei. Thus, the residue of solid material left after drying up
of a droplet is known as droplet nuclei. These are small, 1-4µm, and light.
They can remain suspended in air for hours or days, traveling long distances.
They may serve as a continuing source of infection if the bacteria remain
viable when dry. Viability is determined by a set of complex factors including,
the atmospheric conditions like humidity, sunlight and temperature, the size of
the particles bearing the organisms, and the degree of susceptibility or
resistance of the particular microbial species to the new physical environment.
If inhaled droplet nuclei tend to escape the mechanical traps of the upper
respiratory tract and enter the lungs. Thus, droplet nuclei may act as more
potential agents of infectious diseases than droplets.
Infectious Dust
Large aerosol droplets settle out rapidly from air on to various surfaces
and get dried. Nasal and throat discharges from a patient can also contaminate
surfaces and become dry. Disturbance of this dried material by bed making,
handling a handkerchief having dried secretions or sweeping floors in the
patient's room can generate dust particles which add microorganisms to the
circulating air. Microorganisms can survive for relatively longer periods in
dust. This creates a significant hazard, especially in hospital areas.
Infective dust can also be produced during laboratory practices like opening
the containers of freeze dried cultures or withdrawal of cotton plugs that have
dried after being wetted by culture fluids. These pose a threat to the people working
in laboratories
Significance of Air Microflora - Although, when compared with
the microorganisms of other environments, air microflora are very low in
number, they playa very significant role. This is due to the fact that the air
is in contact with almost all animate and inanimate objects.
The significance of air flora has been studied since 1799, in which year Lazaro
Spallanzani attempted to disprove spontaneous generation. In t 837, Theodore
Schwann, in his experiment to support the view of Spallanzani, introduced fresh
heated air into a sterilized meat broth and demonstrated that microbial growth
couldn't occur. This formed the basis of modern day forced aeration
fermentations. It was Pasteur in 1861, which first showed that microorganisms
could occur as airborne contaminants. He used special cotton in his air sampler
onto which the microorganisms were deposited.
He microscopically demonstrated the presence of microorganisms in the cotton.
In his famous swan necked flask experiment, he showed that growth could not
occur in sterile media unless airborne contamination had occurred.
Factors Affecting Air Microflora - A number of intrinsic and
environmental factors influences the kinds and distribution of the microflora
in air. Intrinsic factors include the nature and physiological state of
microorganisms and also the state of suspension. Spores are relatively more
abundant than the vegetative bacterial cells.
This is mainly due to the dormant nature of spores which enables them to
tolerate unfavourable conditions like desiccation, lack of enough nutrients and
ultraviolet radiation. Similarly fungal spores are abundant in the air since
they are meant for the dispersal of fungi.
The size of the microorganisms is another factor that determines the period of
time for which they remain suspended in air. Generally smaller microorganisms
are easily liberated into the air and remain there for longer period. Fungal
mycelia have a larger size and hence mainly fragments of mycelia will be
present in air. The state of suspension plays an important role in the settling
of microorganisms in air. Organisms in the free state are slightly heavier than
air and settle out slowly in a quiet atmosphere. However, microorganisms
suspended in air are only rarely found in the free state.
Usually they are attached to dust particles and saliva. Microorganisms embedded
in dust particle settle out rapidly and in a quiet atmosphere they remain
airborne only for a short period of time. Droplets which are discharged into
the air by coughing or sneezing are also remain suspended in air for a short
period of time. When their size decreases by evaporation they remain for a
longer period in air.
Environmental factors that affect air microflora include atmospheric
temperature, humidity, air current, the height at which the microorganisms are
found etc. Temperature and relative humidity are the two important factors that
determine the viability of microorganisms in aerosol. Studies with Serratia
marcesens and E. coli show that the airborne survival is closely related to the
temperature.
There is a progressive increase in the death rate with an increase in
temperature from -18°C to 49°C. Viruses in aerosols show a similar behaviour.
Particles of influenza, poliomyelitis and vaccinia viruses survive better at
low temperature from 7 to 24°C.The optimum rate of relative humidity (RH) for
the survival of most microorganisms is between 40 and 80 percent. Low and high
relative humidity cause the death of most microorganisms. Almost all viruses
survive better at a RH of 17 to 25 percent.
A notable exception is that of poliomyelitis which survives better at 80 to 81
percent. survival has been found to be a function of both RH and temperature.
At all temperatures, survival is best at the extremes of RH. Irrespective of
RH, an increase in temperature leads to decrease in survival time.Air current
influences the time for which either the microorganisms or the particles laden
with microorganisms remain suspended in air. In still air the particles tend to
settle down. But a gentle air current can keep them in suspension for
relatively long periods. Air current is also important in the dispersal of
microorganisms as it carries them over a long distance.
Air currents also produce turbulence which causes a vertical distribution of
air flora. Global weather patterns also influence the vertical distribution.
High altitudes have a limiting effect on the air microflora. High altitudes are
characterized by severe conditions like desiccation, ultraviolet radiation and
low temperature. Only resistant forms like spores can survive these conditions.
Thus high attitudes are characterized by the presence of spores and other
resistant forms.
Distribution of Microbes in Air - No microbes are indigenous
to the atmosphere rather they represent allochthonous populations transported
from aquatic and terrestrial habitats into the atmosphere. Microbes of air
within 300-1,000 or more feet of the earth's surface are the organisms of soil
that have become attached to fragments of dried leaves, straw or dust
particles, being blown away by the wind. Species vary greatly in their
sensitivity to a given value of relative humidity, temperature and radiation
exposures.
More microbes are found in air over land masses than far at sea. Spores of fungi,
especially Alternaria, Cladosporium, Penicillium and Aspergillus are more
numerous than other forms over sea within about 400 miles of land in both polar
and tropical air masses at all altitudes up to about 10,000 feet.
Microbes found in air over populated land areas below altitude of 500 feet in
clear weather include spores of Bacillus and Clostridium, ascospores of
yeasts, fragments of myceilium and spores of molds and streptomycetaceae,
pollen, protozoan cysts, algae, Micrococcus, Corynebacterium etc.
In the dust and air of schools and hospital wards or the rooms of persons
suffering from infectious diseases, microbes such as tubercle bacilli,
streptococci, pneumococci and staphylococci have been demonstrated.
These respiratory bacteria are dispersed in air in the droplets of saliva and
mucus produced by coughing, sneezing, talking and laughing. Viruses of
respiratory tract and some enteric tract are also transmitted by dust and air.
Pathogens in dust are primarily derived from the objects contaminated with
infectious secretions that after drying become infectious dust.
Droplets are usually formed by sneezing, coughing and talking. Each droplet
consists of saliva and mucus and each may contain thousands of microbes. It has
been estimated that the number of bacteria in a single sneeze may be between
10,000 and 100,000. Small droplets in a warm, dry atmosphere are dry before
they reach the floor and thus quickly become droplet nuclei.
Many plant pathogens are also transported from one field to another through air
and the spread of many fungal diseases of plants can be predicted by measuring
the concentration of airborne fungal spores. Human bacterial pathogens which
cause important airborne diseases such as diphtheria, meningitis, pneumonia,
tuberculosis and whooping cough are described in the chapter "Bacterial
Diseases of Man".
Air Microflora Significance in Hospitals - Although hospitals
are the war fields for combating against diseases, there are certain occasions
in which additional new infectious diseases can be acquired during
hospitalization. Air within the hospital may act as a reservoir of pathogenic
microorganisms which are transmitted by the patients.Infection acquired during
the hospitalization are called nosocomial infections and the pathogens involved
are called as nosocomial pathogens. Infections, manifested by the corresponding
symptoms, after three days of hospitalization can be regarded as nosocomial
infection (Gleckman & Hibert, 1982 and Bonten& Stobberingh, 1995).
Nosocomial infection may arise in a hospital unit or may be brought in by the
staff or patients admitted to the hospital.The common microorganisms associated
with hospital infection are Haemophilus influenzae, Streptococcus pneumoniae,
Staphylococcus aureus, Pseudomonas aeruginosa, members of Enterobacteriaceae
and respiratory viruses. Development of high antibiotic resistance is a
potential problem among nosocomial pathogens. For example, Methicillin
Resistant Staphylococcus aureus (MRSA) and gentamicin resistant Gram-negative bacilli
are of common occurrence. Even antiseptic liquids used would contain bacteria,
for example Pseudomonas, due to their natural resistance to certain
disinfectants and antiseptics and to many antibiotics.
Nosocomial pathogens may cause or spread hospital outbreaks. Nosocomial
pneumonia is becoming a serious problem nowadays and a number of pathogens have
been associated with it. (Bonten & Stobberingh, 1995). Frequent agents are
Staphylococcus aureus, Streptococcus pneumoniae, Pseudomonas aeruginosa, Enterobacter,
Klebsiella, Escherichia coli and Haemophilus influenzae. Other less frequent
agents are enterococci, streptococci other than S. pneumoniae, Serratia
marcescens, Citrobacter freundii, Acinetobacter sp. and Xanthomonas sp.
In addition Legionella, Chlamydia pneumoniae and Mycobacterium tuberculosis
have also been reported. Nosocomial transmissions of tuberculosis from patients
to patients and from patients to health care workers have also been well
documented (Wenger et a/., 1995). There are two main routes of transmission for
nosocomial pathogens, contact (either direct or indirect) and airborne spread.
Airborne spread is less common than the spread by direct or indirect contact.
It occurs by the following mechanisms. The source may be either from persons or
from inanimate objects.
In case of spread from persons the droplets from mouth, skin scales from nose,
skin exudates and infected lesion transmit diseases such as measles,
tuberculosis, pneumonia, staphylococcal sepsis and streptococcal sepsis. Talking,
coughing and sneezing produce droplets. Skin scales are shed during wound
dressing or bed making.
In case of inanimate sources particles from respiratory equipment and
air-conditioning plant may transmit diseases. These include Gram-negative
respiratory infection, Legionnaire's disease and fungal infections.
Air Microflora Significance in Human Health - The significance
of air microflora in human health relies on the fact that air acts as a medium
for the transmission of infectious agents. An adult man inhales about '5m3 of
air per day. Although most of the microorganisms present in air are harmless
saprophytes and commensals, less than I % of the airborne bacteria are
pathogens.
Eventhough the contamination level is very low, the probability of a person
becoming infected will be greatest if he is exposed to a high concentration of
airborne pathogens. Carriers, either with the manifestation of corresponding
symptoms or without any apparent symptoms, may continuously release respiratory
pathogens in the exhaled air.
Staphylococcus aureus is the most commonly found pathogen in air since the
carriers are commonly present. The number of S. aureus in air may vary between
0-l/m3 and 50/m3.
Practically speaking, outdoor air doesn't contain disease causing pathogen in a
significant number to cause any infection. The purity of outdoor air, however,
is an essential part of man's environment. Dispersion and dilution by large
volume of air is an inherent mechanism of air sanitation in outside air.
In the case of indoor air chance for the spread of infectious disease is more,
especially in areas where people gather in large numbers. For example, in
theatres, schools etc.
Air-Borne Microorganisms and Human Diseases
Air-borne microorganisms cause dangerous diseases in human beings. A detailed
study of these diseases falls under the preview of a text book of medical
microbiology. A chart representing air-borne diseases is given below for ready
reference :
The composition of the microbes of
the air is quite variable. It depends on many factors: on the extent to which
air is contaminated with mineral and organic suspensions, on the temperature,
rainfall, locality, humidity, and other factors. The more dust, smoke, and soot
in the air, the greater the number of microbes. Each particle of dust or smoke
is able to adsorb on its surface numerous microbes. Microbes are rarely found
on the surfaces of mountains, in the seas of Arctic lands covered with snow, in
oceans, and in snow.
The microflora of the air consists of very different species which enter
it from the soil, plants, and animal organisms. Pigmented saprophytic bacteria
(micrococci, various sarcinae), cryptogams (hay bacillus, B. cereus, B. megaterium), actinomycetes, moulds, yeasts, etc., are
often found in the air.
The number of microbes in the air
vanes from a few specimens to many tens of thousands per 1 cu m. Thus, for
example, the air of the
According to the investigations of E.
Mishustin, 1 cu m of air in
At present Streptococcus viridans
serves as sanitary indices for the air of closed buildings, and haemolytic
streptococci and pathogenic staphylococci are a direct epidemiological hazard.
Depending on the time of the year,
the composition and the amount of microflora change. If the total amount of
microbes in winter is accepted as 1, then in spring it will be 1.7, in summer—
2 and in autumn — 1.2.
The total amount of
microbes in an operating room before operation should not exceed 500 per 1 cu m
of air, and after the operation not more than 1000. There should be no
pathogenic staphylococci and streptococci in
The number of microbes in factories
and homes is associated closely with the sanitary hygienic conditions of the
building. In overcrowding, poor
ventilation and natural lighting and if the premises are not properly cleaned,
the number of microbes increases. Dry cleaning processes, infrequent floor
washing, the use of dirty rags and brushes, and drying them in the same room
make the conditions favourable for the accumulation of microbes in air. The
causative agents of influenza, measles, scarlet fever, diphtheria, whooping
cough, meningococcal infections, tonsillitis, acute catarrhs of the respiratory
tract, tuberculosis, smallpox, pneumatic plague, and other diseases can be
transmitted through the air together with droplets of mucus and sputum during
sneezing, coughing, and talking.
Microbes can be spread by air
currents, by aerial dust and aerial droplets. During sneezing, coughing and
talking, a sick person can expel pathogenic bacteria together with droplets of
mucus and sputum into the surrounding environment within a radius of 1.0-
On the average a person breathes
about 12000-
The greatest amount of bacteria is
discharged during sneezing, less — during coughing, and still less — during
talking. With each sneeze a man expels from 10000 to 1 000000 droplets. In one
cough from 10 to 1000 droplets containing bacteria are discharged into the environment,
and when a person utters 10-20 words — up to 80 droplets are expelled. The
nature of the bacterial aerosol depends on the viscosity of the secretion
excreted from the respiratory tract. A liquid secretion is dispersed into more
minute droplets more easily than a viscous one. Near the person expelling the
bacteria a more concentrated aerosol of bacterial droplets from 1 to 2000 mcm
in size is produced. Most of the droplets are from 2 to 100 mcm in size. Large
droplets from 100 to 2000 mcm in size are thrown out to a distance of 2-
The air is an unfavourable medium for microbes. The absence of nutrient
substances, the presence of moisture, optimal temperature, the lethal activity
of sunlight, and desiccation do not create conditions for keeping microbes
viable and most of them perish. However, the relatively short period during
which the microbes are in air is quite enough to bring about the transmission
of pathogenic bacteria and viruses from sick to healthy persons, and to cause
extensive epidemics of diseases such as influenza.
For the purpose of prophylaxis
various methods are used in protecting humans from infection via air-borne
dust. Thus, the sputum of tuberculosis patients is burned or decontaminated, the room is frequently
ventilated, and cleaned by mopping, the streets are sprayed, drainage and
absorbers are used, and masks are used during sorting of wool and rags, etc.
The air of operating rooms, isolating rooms, wards, and bacteriological
laboratories is decontaminated by ultraviolet radiation (mercury-quartz, uviol
lamps, etc.).
The laboratory investigation of air
is carried out to determine the qualitative and quantitative composition of its
microflora. This is achieved by using simple and complex methods. For a more
accurate investigation of microbial contents of the air special apparatus are
used (Rechmensky's bacterial absorber, Krotov's apparatus, Kiktenko's apparatus
(Fig. 1), and others).
Figure 1. Kiktenko's apparatus for
bacteriological testing of the atmosphere
At present rapid methods for the
indication of microbes in the environment are being devised which will allow
quick determinations of the presence of micro-organisms in the soil, the water
and air.
Microflora
of Food Products
Proteins, carbohydrates,
vitamins and other nutrient substances contained in food products have a
favourable effect not only on the preservation of different micro-organisms but
also on their multiplication.
Products of sour milk and foodstuff's produced by fermentation contain a
great number of microbes which lend them flavour and consistency (specific microflora). Besides, micro-organisms
or their spores may get into foodstuff's from the environment (non-specific microflora).
The reproduction of some micro-organisms may cause spoiling of food
products which become unsuitable for eating. In some cases the foodstuffs may
be seeded with Salmonella and Shigella organisms, staphylococci, Clostridium botulinun, Escherichia coli.
Bacillus cereus, Clostridium perfringens, and other microbes which cause
food toxicoinfections and other diseases among humans.
Milk
may be contaminated with Mycobacterium
bovis, Brucellae, Coxiella burnetti, pathogenic streptococci, and
encephalitis viruses from sick animals. During transportation or when it is
being bottled or treated milk may be infected with Salmonella and Shigella organisms,
pathogenic streptococci and staphylococci, Corynebacterium
diphtheriae. Vibrio cholerae, and other microbes by personnel who are sick
or are microbe carriers.
Meat
may have been contaminated when the animals or poultry were still alive but
sick or it may be infected when they are slaughtered, cut, or when the
carcasses are improperly stored and transported. Cl. perfringens, B. cereus, enteric bacteria. Streptococcus faecalis, Proteus, and other bacteria are usually
found in meat. Meat and meat products, minced meat in particular, are most
frequently contaminated during treatment when pathogenic microbes are found on
the surface of the meat chopper, on the hands, and on the kitchen utensils
(cutting board, etc.).
Bacteria from meat
The flesh
of fish is infected with a wide variety of microbial species found
in water, the scales and guts of fish, on the hands of persons involved in processing
the fish products, and on various objects (knives, tables, boards used in
preparing the fish, the deck of a fishing boat, etc.). The most dangerous
micro-organisms are Cl. botulinum
which produce an exotoxin in canned fish products and Vibrio parahaemolytica. When sanitary regimens are not observed, S. typhi, Sh. flexneri, and in some
cases the El Tor vibrio are found in
the flesh of fish and oysters.
Vegetables and
fruit may be seeded with Shigella and Salmonella organisms,
Vibrio cholerae, and microflora found
in the soil and on the hands of persons who take part in their harvesting,
packing, transportation, and those who sell them. Improperly canned vegetables
(tomatoes, mushrooms, etc.) may sometimes be the cause of botulism.
Various microflora, pathogenic species among others (Salmonella organisms, fungi, actinomycetes), penetrate eggs quite
often; egg powder may be contaminated with staphylococci.
Baker's products
are a relatively rare source of infection of man with pathogenic micro-organisms.
Only those baked from grain left in the field the whole winter cause
fusariotoxicosis due to pathogenic Fusarium
genus moulds.
Among all food poisonings encountered among humans, 70 per cent are due
to pathogenic bacteria. Salmonella
organisms, staphylococci, and streptococci are most dangerous; they multiply
and accumulate in the foodstuff's without causing changes in the organoleptic
properties.
In the different countries the
quality norms of most foodstuffs are set by the All-
State Standard (GOST) or Provisional Technical Specifications (VTU).
Microbiological
methods for testing foodstuffs. Foodstuffs are tested in the
following cases: (1) as a planned measure to control the observance of the
sanitary and hygienic regimen in the preparation, storage, and realization of
food, particularly those foodstuffs which are not subjected to treatment at
high temperature; (2) when there is doubt concerning the quality of the food;
(3) when food toxicosis or diseases due to the intake of food occur.
The main task of microbiological testing of a food product is the
determination of the total content of microbes and the model sanitary microbes.
The model sanitary microbe for most foodstuffs and water is E. coli.
Some foodstuffs are tested for the presence of Proteus vulgaris. Salmonella organisms, aerobic and anaerobic
organisms and for the toxins of these microbes.
The technique of collecting the
samples and the sanitary and bacteriologic examination are fixed strictly by
instructions in the corresponding State Standard. It, for instance, specifies
the methods for collecting samples and all stages in testing milk, cream,
ice-cream, butter, koumiss (fermented mare's milk), yoghurt, sour clotted milk,
sour cream, acidophilin (sour fermented milk), cottage cheese and food prepared
from it, dried dairy products, condensed milk, and cold beverages prepared from
milk. For testing, liquid foodstuffs are diluted with sterile isotonic solution
1:10. Compact products are melted or ground in the mortar and diluted in
sterile water 1:10.
Sanitary and bacteriologic tests of
milk and dairy products consist in determination of the total microbial content
(microbial count) and the coli titre. In sour dairy products (yoghurt, cottage
cheese, cheese, etc.) the microbial count is not determined. The microbial
count in milk is determined by a direct count and by inoculating nutrient media
with 1.0 ml of different dilutions of the product that is tested. The dairy
products should only contain microorganisms specific for the given food, e. g.
lactic streptococci and lactobacilli in sour clotted milk, lactobacilli and
yeasts in koumiss, etc.
The permissible microbial count in
various dairy products ranges between 500 (children's mixtures subjected to pasteurization
and cooking) and 300000 (cow's milk in cans and cisterns). The microbial count
for pausterized milk kept in bottles and packets is 75 000, for ice-cream
250000, for condensed milk 50000, for dried cow's milk 50000 per one
millilitre.
The coli titre of dairy products is
determined by a three-stage fermentation method and for most of them it ranges
from 0.3 to 3; only for children's milk mixtures (pausterized and cooked) it is
above 11.1. Since milk and dairy
products may be vehicles of the causative agents of certain infectious diseases
(typhoid fever, paratyphoid fevers, brucellosis, tuberculosis, Q fever, etc.),
these agents are identified by special methods discussed in the corresponding
sections of the special part of this textbook. If pathogenic micro-organisms
are detected in dairy products it is unquestionable that these products are not
fit to be eaten.
The sanitary and bacteriological testing of meat and meat products
comprises determination of the total
amount of microbes per
No stable norms have been fixed to
date for the sanitary and bacteriological assessment of meat and meat products.
According to the accepted provisional norms, the permissible microbial count
for roast meat should be less than 500, for boiled sausage and meat jelly less
than 1000. The coli titre for roast meat should be above
Canned foodstuffs, such as canned meat, lard, beans, fish, vegetables
and juices are also subjected to sanitary and bacteriological testing. Canned food is tested microbiologically for
aerobic and anaerobic micro-organisms and for the botulinum toxin. When there
are epidemiological indications, canned food is tested for Salmonella organisms, pathogenic staphylococci, and Proteus vulgaris; the presence of these
microbes shows that the canned food is spoiled and cannot be eaten. It is
permissible for canned food to contain non-pathogenic sporulating microbes
provided there is no bulging of the can and the organoleptic properties of the
food are normal.
Fish, vegetables, and eggs are tested
microbiologically usually in cases of food poisoning or diseases among humans.
Tests are performed for detecting pathogenic and conditionally pathogenic
microbes or their toxins by the commonly accepted methods.
Remnants
of food and foodstuffs, vomit, lavage waters, faeces, blood, mucus, washings
and scrapings, and autopsy material may be subjected to bacteriological testing
according to the epidemiological indications or on instruction of a health
officer.
The prevention of food poisoning and other diseases acquired through
foodstuffs consists in the observance of sanitary and hygienic measures in
preparation of food products, their storage, transportation, and realization.
It is also necessary to observe strictly the rules for processing foodstuffs, especially
for canning them. Since foodstuffs may be infected by the service staff among
whom there may be sick persons or carriers of pathogenic microorganisms, all
personnel of food-supplying establishments must be examined regularly. Control
of the vectors of the causative agents of intestinal infections and health
education among the population are extensively carried out.
Microflora
of the body
Significance of the Normal Flora
The normal flora influences the
anatomy, physiology, susceptibility to pathogens, and morbidity of the host.
Skin
Flora
The varied environment of the
skin results in locally dense or sparse populations, with Gram-positive
organisms (e.g., staphylococci, micrococci, diphtheroids) usually
predominating.
Oral
and Upper Respiratory Tract Flora
A varied microbial flora is
found in the oral cavity, and streptococcal anaerobes inhabit the gingival
crevice. The pharynx can be a point of entry and initial colonization for Neisseria,
Bordetella, Corynebacterium, and Streptococcus spp.
Gastrointestinal
Tract Flora
Organisms in the stomach are
usually transient, and their populations are kept low (103 to 106/g
of contents) by acidity. Helicobacter pylori is a potential stomach
pathogen that apparently plays a role in the formation of certain ulcer types.
In normal hosts the duodenal flora is sparse (0 to 103/g of
contents). The ileum contains a moderately mixed flora (106 to 108/g
of contents). The flora of the large bowel is dense (109 to 1011/g
of contents) and is composed predominantly of anaerobes. These organisms
participate in bile acid conversion and in vitamin K and ammonia production in
the large bowel. They can also cause intestinal abscesses and peritonitis.
Urogenital
Flora
The vaginal flora changes with
the age of the individual, the vaginal pH, and hormone levels. Transient
organisms (e.g., Candida spp.) frequently cause vaginitis. The distal
urethra contains a sparse mixed flora; these organisms are present in urine
specimens (104/ml) unless a clean-catch, midstream specimen is
obtained.
Conjunctival
Flora
The conjunctiva harbors few or
no organisms. Haemophilus and Staphylococcus are among the genera
most often detected.
Host
Infection
Many elements of the normal
flora may act as opportunistic pathogens, especially in hosts rendered
susceptible by rheumatic heart disease, immunosuppression, radiation therapy,
chemotherapy, perforated mucous membranes, etc. The flora of the gingival
crevice causes dental caries in about 80 percent of the population.
Introduction
A diverse microbial flora is
associated with the skin and mucous membranes of every human being from shortly
after birth until death. The human body, which contains about 1013
cells, routinely harbors about 1014 bacteria (Fig. 6-1).
This bacterial population constitutes the normal microbial flora . The
normal microbial flora is relatively stable, with specific genera populating
various body regions during particular periods in an individual's life.
Microorganisms of the normal flora may aid the host (by competing for
microenvironments more effectively than such pathogens as Salmonella spp
or by producing nutrients the host can use), may harm the host (by causing
dental caries, abscesses, or other infectious diseases), or may exist as
commensals (inhabiting the host for long periods without causing detectable
harm or benefit). Even though most elements of the normal microbial flora
inhabiting the human skin, nails, eyes, oropharynx, genitalia, and
gastrointestinal tract are harmless in healthy individuals, these organisms
frequently cause disease in compromised hosts. Viruses and parasites are not
considered members of the normal microbial flora by most investigators because
they are not commensals and do not aid the host.
Numbers of bacteria that
colonize different parts of the body. Numbers represent the number of organisms
per gram of homogenized tissue or fluid or per square centimeter of skin
surface.
Significance
of the Normal Flora
The fact that the normal flora
substantially influences the well-being of the host was not well understood
until germ-free animals became available. Germ-free animals were obtained by
cesarean section and maintained in special isolators; this allowed the
investigator to raise them in an environment free from detectable viruses,
bacteria, and other organisms. Two interesting observations were made about
animals raised under germ-free conditions. First, the germ-free animals lived
almost twice as long as their conventionally maintained counterparts, and
second, the major causes of death were different in the two groups. Infection
often caused death in conventional animals, but intestinal atonia frequently
killed germ-free animals. Other investigations showed that germ-free animals
have anatomic, physiologic, and immunologic features not shared with
conventional animals. For example, in germ-free animals, the alimentary lamina
propria is underdeveloped, little or no immunoglobulin is present in sera or
secretions, intestinal motility is reduced, and the intestinal epithelial cell
renewal rate is approximately one-half that of normal animals (4 rather than 2
days).
Although the foregoing
indicates that bacterial flora may be undesirable, studies with antibiotic
treated animals suggest that the flora protects individuals from pathogens.
Investigators have used streptomycin to reduce the normal flora and have then
infected animals with streptomycin-resistant Salmonella. Normally, about
106 organisms are needed to establish a gastrointestinal infection,
but in streptomycin-treated animals whose flora is altered, fewer than 10
organisms were needed to cause infectious disease. Further studies suggested
that fermentation products (acetic and butyric acids) produced by the normal
flora inhibited Salmonella growth in the gastrointestinal tract. Figure 6-2
shows some of the factors that are important in the competition between the
normal flora and bacterial pathogens.
Mechanisms by which the normal
flora competes with invading pathogens. Compare this schematic with Figure 6-3.
The normal flora in humans
usually develops in an orderly sequence, or succession, after birth, leading to
the stable populations of bacteria that make up the normal adult flora. The
main factor determining the composition of the normal flora in a body region is
the nature of the local environment, which is determined by pH, temperature,
redox potential, and oxygen, water, and nutrient levels. Other factors such as
peristalsis, saliva, lysozyme secretion, and secretion of immunoglobulins also
play roles in flora control. The local environment is like a concerto in which
one principal instrument usually dominates. For example, an infant begins to
contact organisms as it moves through the birth canal. A Gram-positive
population (bifidobacteria arid lactobacilli) predominates in the
gastrointestinal tract early in life if the infant is breast-fed. This
bacterial population is reduced and displaced somewhat by a Gram-negative flora
(Enterobacteriaceae) when the baby is bottle-fed. The type of liquid diet
provided to the infant is the principal instrument of this flora control;
immunoglobulins and, perhaps, other elements in breast milk may also be
important.
What, then, is the significance
of the normal flora? Animal and some human studies suggest that the flora
influences human anatomy, physiology, lifespan, and, ultimately, cause of
death. Although the causal relationship of flora to death and disease in humans
is accepted, of her roles of the human microflora need further study.
Normal
Flora of Skin
Skin provides good examples of
various microenvironments. Skin regions have been compared to geographic
regions of Earth: the desert of the forearm, the cool woods of the scalp, and
the tropical forest of the armpit. The composition of the dermal microflora
varies from site to site according to the character of the microenvironment. A
different bacterial flora characterizes each of three regions of skin: (1)
axilla, perineum, and toe webs; (2) hand, face and trunk; and (3) upper arms
and legs. Skin sites with partial occlusion (axilla, perineum, and toe webs)
harbor more microorganisms than do less occluded areas (legs, arms, and trunk).
These quantitative differences may relate to increased amount of moisture,
higher body temperature, and greater concentrations of skin surface lipids. The
axilla, perineum, and toe webs are more frequently colonized by Gram-negative
bacilli than are drier areas of the skin.
The number of bacteria on an
individual's skin remains relatively constant; bacterial survival and the
extent of colonization probably depend partly on the exposure of skin to a
particular environment and partly on the innate and species-specific
bactericidal activity in skin. Also, a high degree of specificity is involved
in the adherence of bacteria to epithelial surfaces. Not all bacteria attach to
skin; staphylococci, which are the major element of the nasal flora, possess a
distinct advantage over viridans streptococci in colonizing the nasal mucosa.
Conversely, viridans streptococci are not seen in large numbers on the skin or
in the nose but dominate the oral flora.
The microbiology literature is
inconsistent about the density of bacteria on the skin; one reason for this is
the variety of methods used to collect skin bacteria. The scrub method yields
the highest and most accurate counts for a given skin area. Most microorganisms
live in the superficial layers of the stratum corneum and in the upper parts of
the hair follicles. Some bacteria, however, reside in the deeper areas of the
hair follicles and are beyond the reach of ordinary disinfection procedures.
These bacteria are a reservoir for recolonization after the surface bacteria
are removed.
Staphylococcus
epidermidis
S. epidermidis
is a major inhabitant of the skin, and in some areas it makes up more than 90
percent of the resident aerobic flora.
Staphylococcus
aureus
The nose and perineum are the
most common sites for S. aureus colonization, which is present in 10
percent to more than 40 percent of normal adults. S. aureus is prevalent
(67 percent) on vulvar skin. Its occurrence in the nasal passages varies with
age, being greater in the newborn, less in adults. S. aureus is
extremely common (80 to 100 percent) on the skin of patients with certain
dermatologic diseases such as atopic dermatitis, but the reason for this
finding is unclear.
Micrococci
Micrococci are not as common
as staphylococci and diphtheroids; however, they are frequently present on
normal skin. Micrococcus luteus, the predominant species, usually
accounts for 20 to 80 percent of the micrococci isolated from the skin.
Diphtheroids
(Coryneforms)
The term diphtheroid denotes a
wide range of bacteria belonging to the genus Corynebacterium. Classification
of diphtheroids remains unsatisfactory; for convenience, cutaneous diphtheroids
have been categorized into the following four groups: lipophilic or
nonlipophilic diphtheroids; anaerobic diphtheroids; diphtheroids producing
porphyrins (coral red fluorescence when viewed under ultraviolet light); and
those that possess some keratinolytic enzymes and are associated with
trichomycosis axillaris (infection of axillary hair). Lipophilic diphtheroids
are extremely common in the axilla, whereas nonlipophilic strains are found more
commonly on glabrous skin.
Anaerobic diphtheroids are
most common in areas rich in sebaceous glands. Although the name Corynebacterium
acnes was originally used to describe skin anaerobic diphtheroids, these are
now classified as Propionibacterium acnes and as P. granulosum. P.
acnes is seen eight times more frequently than P. granulosum in acne
lesions and is probably involved in acne pathogenesis. Children younger than 10
years are rarely colonized with P. acnes. The appearance of this
organism on the skin is probably related to the onset of secretion of sebum (a
semi-fluid substance composed of fatty acids and epithelial debris secreted
from sebaceous glands) at puberty. P. avidum, the third species of
cutaneous anaerobic diphtheroids, is rare in acne lesions and is more often
isolated from the axilla.
Streptococci
Streptococci, especially β-hemolytic
streptococci, are rarely seen on normal skin. The paucity of β-hemolytic
streptococci on the skin is attributed at least in part to the presence of lipids
on the skin, as these lipids are lethal to streptococci. Other groups of
streptococci, such as α-hemolytic
streptococci, exist primarily in the mouth, from where they may, in rare
instances, spread to the skin.
Gram-Negative
Bacilli
Gram-negative bacteria make up
a small proportion of the skin flora. In view of their extraordinary numbers in
the gut and in the natural environment, their scarcity on skin is striking.
They are seen in moist intertriginous areas, such as the toe webs and axilla,
and not on dry skin. Desiccation is the major factor preventing the
multiplication of Gram-negative bacteria on intact skin. Enterobacter, Klebsiella,
Escherichia coli, and Proteus spp. are the predominant
Gram-negative organisms found on the skin. Acinetobacter spp also occurs
on the skin of normal individuals and, like other Gram-negative bacteria, is
more common in the moist intertriginous areas.
Nail
Flora
The microbiology of a normal
nail is generally similar to that of the skin. Dust particles and other extraneous
materials may get trapped under the nail, depending on what the nail contacts.
In addition to resident skin flora, these dust particles may carry fungi and
bacilli. Aspergillus, Penicillium, Cladosporium, and Mucor
are the major types of fungi found under the nails.
Oral
and Upper Respiratory Tract Flora
The oral flora is involved in
dental caries and periodontal disease, which affect about 80 percent. of the
population in the Western world. The oral flora, its interactions with the
host, and its response to environmental factors are thoroughly discussed in
another Chapter. Anaerobes in the oral flora are responsible for many of the
brain, face, and lung infections that are frequently manifested by abscess
formation.
The pharynx and trachea
contain primarily those bacterial genera found in the normal oral cavity (for
example, α-and
β-hemolytic
streptococci); however, anaerobes, staphylococci, neisseriae, diphtheroids, and
others are also present. Potentially pathogenic organisms such as Haemophilus,
mycoplasmas, and pneumococci may also be found in the pharynx. Anaerobic
organisms also are reported frequently. The upper respiratory tract is so often
the site of initial colonization by pathogens (Neisseria meningitides, C.
diphtheriae, Bordetella pertussis, and many others) and could be
considered the first region of attack for such organisms. In contrast, the
lower respiratory tract (small bronchi and alveoli) is usually sterile, because
particles the size of bacteria do not readily reach it. If bacteria do reach
these regions, they encounter host defense mechanisms, such as alveolar
macrophages, that are not present in the pharynx.
Gastrointestinal
Tract Flora
The stomach is a relatively
hostile environment for bacteria. It contains bacteria swallowed with the food
and those dislodged from the mouth. Acidity lowers the bacterial count, which
is highest (approximately 103 to 106 organisms/g of
contents) after meals and lowest (frequently undetectable) after digestion.
Some Helicobacter species can colonize the stomach and are associated
with type B gastritis and peptic ulcer disease. Aspirates of duodenal or
jejunal fluid contain approximately 103 organisms/ml in most
individuals. Most of the bacteria cultured (streptococci, lactobacilli, Bacteroides)
are thought to be transients. Levels of 105 to about 107
bacteria/ml in such aspirates usually indicate an abnormality in the digestive
system (for example, achlorhydria or malabsorption syndrome). Rapid peristalsis
and the presence of bile may explain in part the paucity of organisms in the
upper gastrointestinal tract. Further along the jejunum and into the ileum,
bacterial populations begin to increase, and at the ileocecal junction they
reach levels of 106 to 108 organisms/ml, with
streptococci, lactobacilli, Bacteroides, and bifidobacteria predominating.
Concentrations of 109
to 1011 bacteria/g of contents are frequently found in human colon
and feces. This flora includes a bewildering array of bacteria (more than 400
species have been identified); nonetheless, 95 to 99 percent belong to
anaerobic genera such as Bacteroides, Bifidobacterium, Eubacterium,
Peptostreptococcus, and Clostridium. In this highly anaerobic
region of the intestine, these genera proliferate, occupy most available niches,
and produce metabolic waste products such as acetic, butyric, and lactic acids.
The strict anaerobic conditions, physical exclusion (as is shown in many animal
studies), and bacterial waste products are factors that inhibit the growth of
other bacteria in the large bowel.
Although the normal flora can
inhibit pathogens, many of its members can produce disease in humans. Anaerobes
in the intestinal tract are the primary agents of intra-abdominal abscesses and
peritonitis. Bowel perforations produced by appendicitis, cancer, infarction,
surgery, or gunshot wounds almost always seed the peritoneal cavity and
adjacent organs with the normal flora. Anaerobes can also cause problems within
the gastrointestinal lumen. Treatment with antibiotics may allow certain
anaerobic species to become predominant and cause disease. For example, Clostridium
difficile, which can remain viable in a patient undergoing antimicrobial
therapy, may produce pseudomembranous colitis. Other intestinal pathologic
conditions or surgery can cause bacterial overgrowth in the upper small
intestine. Anaerobic bacteria can then deconjugate bile acids in this region
and bind available vitamin B12 so that the vitamin and fats are
malabsorbed. In these situations, the patient usually has been compromised in
some way; therefore, the infection caused by the normal intestinal flora is
secondary to another problem.
More information is available
on the animal than the human microflora. Research on animals has revealed that
unusual filamentous microorganisms attach to ileal epithelial cells and modify
host membranes with few or no harmful effects. Microorganisms have been
observed in thick layers on gastrointestinal surfaces (Fig. 6-3)
and in the crypts of Lieberkuhn. Other studies indicate that the immune
response can be modulated by the intestinal flora. Studies of the role of the
intestinal flora in biosynthesis of vitamin K and other host-utilizable
products, conversion of bile acids (perhaps to cocarcinogens), and ammonia
production (which can play a role in hepatic coma) show the dual role of the
microbial flora in influencing the health of the host. More basic studies of
the human bowel flora are necessary to define their effect on humans.
(A) Scanning electron
micrograph of a cross-section of rat colonic mucosa. The bar indicates the
thick layer of bacteria between the mucosal surface and the lumen (L) (X 262,)
(B) Higher (more...)
Urogenital
Flora
The type of bacterial flora
found in the vagina depends on the age, pH, and hormonal levels of the host. Lactobacillus
spp. predominate in female infants (vaginal pH, approximately 5) during the
first month of life. Glycogen secretion seems to cease from about I month of
age to puberty. During this time, diphtheroids, S. epidermidis,
streptococci, and E. coli predominate at a higher pH (approximately pH
7). At puberty, glycogen secretion resumes, the pH drops, and women acquire an
adult flora in which L. acidophilus, corynebacteria, peptostreptococci,
staphylococci, streptococci, and Bacteroides predominate. After menopause, pH
again rises, less glycogen is secreted, and the flora returns to that found in
prepubescent females. Yeasts (Torulopsis and Candida) are
occasionally found in the vagina (10 to 30 percent of women); these sometimes
increase and cause vaginitis.
In the anterior urethra of
humans, S. epidermidis, enterococci, and diphtheroids are found
frequently; E. coli, Proteus, and Neisseria (nonpathogenic
species) are reported occasionally (10 to 30 percent). Because of the normal
flora residing in the urethra, care must be taken in clinically interpreting
urine cultures; urine samples may contain these organisms at a level of 104/ml
if a midstream (clean-catch) specimen is not obtained.
Conjunctival
Flora
The conjunctival flora is
sparse. Approximately 17 to 49 percent of culture samples are negative.
Lysozyme, secreted in tears, may play a role in controlling the bacteria by
interfering with their cell wall formation. When positive samples show
bacteria, corynebacteria, Neisseriae, and Moraxellae are cultured.
Staphylococci and streptococci are also present, and recent reports indicate
that Haemophilus parainfluenzae is present in 25 percent of conjunctival
samples.
Host
Infection by Elements of the Normal Flora
This chapter has briefly
described the normal human flora; however, the pathogenic mechanisms of various
genera or the clinical syndromes in which they are involved was not discussed.
Although such material is presented in other chapters, note that a breach in
mucosal surfaces often results in infection of the host by members of the
normal flora. Caries, periodontal disease, abscesses, foul-smelling discharges,
and endocarditis are hallmarks of infections with members of the normal human
flora (Fig. 6-4).
In addition, impairment of the host (for example, those with heart failure or
leukemia) or host defenses (due to immunosuppression, chemotherapy, or
irradiation) may result in failure of the normal flora to suppress transient
pathogens or may cause members of the normal flora to invade the host
themselves. In either
situation, the host may die.
Clinical conditions that may be caused by members of
the normal flora
Microbiocenosis is microbial
community of different bacterial populations, which colonize certain
biotope.
Biotope is an area with
relatively homogenous conditions where microbial population can survive.
Ecological niche is the place
or status of microbes in their biotic environment.
Constant (obligate, resident,
indigenous, autochthonous) microflora is native, no imported one of different biotopes.
Transient (temporary,
facultative, allochthonous) microflora is not aboriginal, it is acquired one.
Before birth, the human body has no
normal flora. During the birth process, the body comes in contact with microbes
in the external environment. Later with the initial feedings and exposure to an
expanding environment some microorganisms find their way to a permanent
residence in many parts of the body
Most organisms in the external environment apparently do not find the
body to be a favourable habitat. Charactenstic features of different body
areas, such as temperature,
oxygen availability, nutrient availability,
natural inhibitors and pH influence the population that is
able to survive and establish itself. Because these conditions vary from site
to site in the body different sites acquire considerably
different organisms as their normal flora. Once the normal flora is established
it benefits the body by preventing the overgrowth of undesirable organisms.
Destruction of the normal flora frequently disrupts the status quo resulting in
the growth of harmful organisms. This can be seen after the prolonged administration
of broad spectrum antibiotics. For example if the normal flora of the
intestinal tract and vagina are largely
destroyed, the yeast Candida albicans which is unaffected by these
antibacterial antibiotics can grow unchecked to become the major organism in
these areas. It then cm infect the mucous membranes and the skin, causing a
severe inflammation. Another complication of antibiotic therapy is a
severe gastroenteritis known as pseudomembranous
colitis. This syndrome has been associated with several antimicrobial agents
but the antibiotics clindamycin and lincomycin have been incriminated most
often. The mechanism of this diarrhoea was elucidated when it was observed that
the use of these antibiotics resulted in an overgrowth in the intestine of an
organism identified is Clostridium difficile. This organism produces in
enterotoxin that causes the gastroenteritis but it can do so only when
antibiotic therapy destroys much of the other normal intestinal flora
permitting it to grow unchecked.
Our normal flora can be categorized is helpful (mutualutic symbionts)
harmless (commensals) or potentially harmful (opportunists) However these
groups are not mutually exclusive. Under certain circumstances even a mutualism
cause harm and, thus, become a pathogen. Therefore these categories are of
value only in describing the usual role of the organism in relation to its
host.
In a mutualistic relationship the microbe and the host benefit one
another. This type of relationship is common in the plant kingdom and is essential
in ruminants such as cattle in which microbes are necessary for digestion of
the cellulose in plant material. Few such relation ships exist in humans,
however. Probably the only good example of mutualism in humans is found in the
normal flora of the large intestine where enteric organisms synthesize vitamin
K and the vitamins of the B complex, enabling them to be absorbed through
the intestinal wall and contribute to human nutrition. However considering that
our normal flora provides us with protection by interfering with the growth of
potentially harmful organisms much of our normal flora could be considered
mutualistic symbionts.
The microbe that lives on and benefits from its host without either
benefiting or harming the host is called a commensal. Most of the organisms
that make up the normal flora of a healthy individual could be categorized as
commensals.
Opportunists (microbes that are potential pathogens) are of greatest
interest to us. These organisms seem to lack the ability to invade and cause
disease in healthy individuals but may be able to colonize as pathogens in ill
or injured persons. Staphylococcus aureus is a good example of an opportunist.
Many people (about 25 %) carry staphylococci in their nasopharynx without
suffering any illness. However if these people acquire respiratory tract
infections such as measles or influenza the staphylococci can invade the lung
and cause severe pneumonia. Accidental contamination of the bladder with E coli
or Enterococcus faecalis during a catheterization
procedure also can lead to opportunistic infection. Both these organisms ire
part of the normal flora of the large intestine and usually do not produce
urinary tract infections. How ever if they gain access to the urethra or are
transplanted mechanically to an environment in which they can grow they can
cause disease. The most startling examples of opportunistic infections are
associated with the viral infection known as acquired immunodeficiency syndrome
(AIDS). This syndrome is caused by a virus that destroys certain subsets of T
cells inhibiting the body's ability to mount in
immune response. As a result infection by the virus causing AIDS is
characterized by severe and eventually fatal infections or malignancies that do
not occur in individuals with functional immune systems.
Because much of our normal flora can cause disease under the proper
conditions these organisms could be considered opportunists. This is
particularly true in elderly and debilitated individuals and in patients
receiving immunosuppressive drug therapy to prevent rejection of organ
transplants. Opportunists are especially important as causes of nosocomial
infections (ic those acquired during hospitalization). Under appropriate circumstances most of the
organisms that constitute our normal flora can
cause disease.
Another group of bacteria that is not
really part of our normal flora consists of pathogenic organisms that can exist
in a large percentage of the population without causing disease. This group
includes such organisms is Neisseria meningitidis (also called the
meningococcus) the causative agent of epidemic meningitis. Many individuals
carry this organism in their respiratory tracts without ever having meningitis,
yet they can spread the bacterium to nonimmune individuals and cause disease.
Streptococcus pneumoniae (the pneumococcus) the major cause of lobar pneumonia
also is carried by 10 % to 20 % of normal healthy individuals. Persons who
harbor bacteria such is these without ever exhibiting overt symptoms of disease
are referred to as carriers.
Thus, although our normal flora can be beneficial by preventing the
growth of potential pathogens, it also can be a reservoir from which endemic
and epidemic diseases are spread.
A diverse microbial flora is associated with the skin and
mucous membranes of every human being from shortly after birth until death. The
human body, which contains about 1013 cells, routinely harbors about
1014 bacteria.This bacterial population
constitutes the normal microbial flora. The normal microbial flora is
relatively stable, with specific genera populating various body regions during
particular periods in an individual's life. Microorganisms of the normal flora
may aid the host (by competing for microenvironments more effectively than such
pathogens as Salmonella spp or by producing nutrients the host can use), may
harm the host (by causing dental caries, abscesses, or other infectious
diseases), or may exist as commensals (inhabiting the host for long periods
without causing detectable harm or benefit). Even though most elements of the
normal microbial flora inhabiting the human skin, nails, eyes, oropharynx,
genitalia, and gastrointestinal tract are harmless in healthy individuals,
these organisms frequently cause disease in compromised hosts. Viruses and
parasites are not considered members of the normal microbial flora by most
investigators because they are not commensals and do not aid the host.
Human microflora is the result of a mutual adaptation of micro- and
macro-organisms in the process of evolution. Most bacteria of the normal and constant
microflora of the human body have adapted themselves to life in certain parts
of the body (tabl. 1, 2, 3). Besides,
there are some microbes which make up a temporary (casual) microflora.
With the development of virology and the improvement of virological
technique, our concepts on the microflora of the human body were increased. It
has been established that not only the open cavities, but the tissues of the
human organism are inhabited by numerous persisting viruses which are excreted
into the environment with milk, saliva, sputum, perspiration, urine, and
faeces.
There are different methods of human microbiocenosis studying. Among
them, biopsy, pad method, impression method, sticky film method, swab-washing
method, scrub-washing technique etc. After receiving of tested material it
should be inoculated on different nutrient media.
Etiology of different human infections
Microflora of the
skin. Staphylococci, streptococci, moulds and yeasts,
diphtheroids, and also certain pathogenic and conditionally pathogenic bacteria
live on the surface of the skin. They receive their nutrition from the
secretions of the sebaceous and sweat glands, dead cells, and waste products.
The total number of microbes on the skin of one person varies from
85000000 to 1212000000.
When the human body comes into contact with the soil, the clothes and
skin are seeded with spores of different species of microbes (organisms
responsible for tetanus, anaerobic infections, etc.).
Most frequently the exposed parts of the human body are infected, e. g.
the hands, on the surface of which colibacilli, staphylococci, streptococci,
enterococci, moulds, yeasts, fungi imperfecti, and spores of aerobic and
anaerobic bacilli are found.
Pustular and fungal infections of the skin and gastrointestinal diseases
often occur owing to violation of sanitary-hygienic conditions and normal
conditions of the work and life of people.
Microbes of the
mouth cavity. In
the mouth cavity there are more than 100 species of microbes. There are the
natural inhabitants (acidophilic bacillus, Treponema microdentium, diplococci,
Streptococcus salivarius, Entamoeba gingivalis, etc.). Besides, in the mouth
cavity there are foreign microbes or those which have been carried in from the
environment together with food, water, and air.
Pathogenic and conditionally pathogenic microbes (staphylococci,
streptococci, diphtheria bacilli, diphtheroids, Borrelia organisms,
spindle-shaped bacteria), protozoa (amoebae and trichomonads) are found on the
mucous membrane of the mouth.
The mouth cavity is a favourable medium for many microbes; it has an
optimal temperature, a sufficient amount of food substances, and has a weakly
alkaline reaction.
The greatest amount of microbes can be found at the necks of the teeth
and in the spaces between teeth. Streptococci and diplococci are found on the
tonsils. There are many microbes in other parts of the mouth cavity, which are
inaccessible to the bathing action of saliva and the action of lysozyme (an
enzyme found in the saliva, lacrimal fluid and sputum). The presence of carious
teeth is a condition for increasing the microflora in the mouth cavity, for the
appearance of decaying processes and unpleasant odours.
The microflora of
the gastrointestinal tract (Table 3, 4). When the stomach functions normally, it is almost
devoid of microflora due to the marked bactericidal properties of gastric
juice.
The gastric Juice is considered to be a reliable defense barrier against
the penetration of pathogenic and
conditionally pathogenic microbes into the intestine. However, the degree of
acidity of the gastric juice is not always constant. It varies according to the
character of the food and the amount of water consumed.
Table
3.
BACTERIA CONSIDERED TO BE
AND THEIR ANATOMICAL
LOCATIONS
|
BACTERIUM |
SKIN |
CONJUNCTIVA |
NOSE |
PHARYNX |
MOUTH |
LOWER
INTES-TINE |
ANTERIOR
URETHRA |
VAGINA |
|
Staphylococcus epidermidis |
++ |
+ |
++ |
+ |
++ |
+ |
++ |
+ |
|
Staphylococcus aureus |
+ |
+/– |
+ |
+ |
+ |
++ |
+/– |
+ |
|
Streptococcus mitis |
– |
– |
– |
+ |
++ |
+/- |
+ |
+ |
|
Streptococcus salivarius |
|
|
|
++ |
++ |
– |
|
|
|
Streptococcus mutans |
|
|
|
+ |
++ |
|
|
|
|
Enterococci |
|
|
|
+/– |
+ |
++ |
+ |
+ |
|
Streptococcus pneumoniae |
|
+/– |
+/– |
+ |
+ |
|
|
+/– |
|
Streptococcus pyogenes |
+/– |
+/– |
|
+ |
+ |
+/– |
|
+ |
|
Neisseriae |
|
+ |
+ |
++ |
+ |
|
+ |
+ |
|
Neisseria meningitidis |
|
|
+ |
+ |
++ |
|
|
+ |
|
Veillonellae |
|
|
|
|
+ |
+/– |
|
|
|
Coliforms (E. coli) |
|
+/– |
+/– |
+/– |
+ |
++ |
+ |
+ |
|
Proteus mirabilis |
|
+/– |
+ |
+ |
+ |
+ |
+ |
+ |
|
Pseudomonas aeruginosa |
|
|
|
+/– |
+/– |
+ |
+/– |
|
|
Haemophilus influenzae |
|
+/– |
+ |
+ |
+ |
|
|
|
|
Bacteroides |
|
|
|
+ |
+ |
++ |
+ |
+/– |
|
Spirochetes |
|
|
|
+ |
+ |
+ |
|
|
|
Lactobacilli |
|
|
|
+ |
++ |
+ |
++ |
|
|
Clostridia |
|
|
|
|
+/– |
++ |
|
|
|
Clostridium tetani |
|
|
|
|
|
+/– |
|
|
|
Corynebacteria |
++ |
+ |
++ |
+ |
+ |
+ |
+ |
+ |
|
Mycobacteria |
+ |
|
+/– |
+/– |
|
+ |
+ |
|
|
Actinomycetes |
|
|
|
+ |
+ |
|
|
|
|
Mycoplasmas |
|
|
|
+ |
+ |
+ |
+/– |
+ |
“++” – prominent; “+” – common;
“+/-” –“irregular, occasional or transient.
Together with food, lactic acid bacteria, Sarcina
ventriculi. hay bacillus, yeasts, etc., enter the stomach from the mouth. In
some cases, dysentery, enteric fever, and paratyphoid bacilli and other
pathogenic microbes are capable of penetrating into the stomach and then the
intestine.
Table 3
EXAMPLES OF TISSUE
SPECIFICITIES OF SOME BACTERIA ASSOCIATED WITH HUMANS
|
BACTERIUM |
TISSUE |
Corynebacterium diphtheriae
|
Throat |
|
Neisseria
gonorrhoeae |
Urogenital
epithelium |
|
Streptococcus
mutans |
Tooth
surfaces |
|
Streptococcus
salivarius |
Tongue
surfaces |
|
Vibrio
cholerae |
Small
intestine epithelium |
|
Escherichia
coli |
Small
intestine epithelium |
|
Staphylococcus
aureus |
Nasal
membranes |
|
Staphylococcus
epidermidis |
Skin |
Enterococci, fungi, and various other microbes are relatively rarely
found in the duodenum. There are few microbes in the small intestine.
Enterococci are found more often than others – In the large intestine there are
large amounts of micro-organisms. Almost one-third of the dry weight of the
faeces of certain animal species is made up of microbes. Daily, an adult human
excretes about 17 million billion micro-organisms with the excrements
(Table 4, 5).
Table 4.
EXAMPLES OF SPECIFIC
ATTACHMENTS OF BACTERIA
TO HOST CELL OR TISSUE
SURFACES
|
Bacterium |
Ligand |
Receptor
|
Attachment site |
Disease |
|
Streptococcus pyogenes |
Protein F |
Amino terminus of fibronectin |
Pharyngeal epithelium |
Sore throat |
|
Streptococcus mutans |
Glycosyl transferase |
Salivary glycoprotein |
Pellicle of tooth |
Dental caries |
|
Streptococcus salivarius |
Lipoteichoic acid |
Unknown |
Buccal epithelium of tongue |
None |
|
Streptococcus pneumoniae |
Cell-bound protein |
N-acetylhexosamine-galactose
disaccharide |
Mucosal epithelium |
pneumonia |
|
Staphylococcus aureus |
Cell-bound protein |
Amino terminus of fibronectin |
Mucosal epithelium |
Various |
|
Neisseria gonorrhoeae |
N-methylphenyl- alanine pili |
Glucosamine-galactose carbohydrate |
Urethral/cervical epithelium |
Gonorrhea |
|
Enterotoxigenic E. coli |
Type-1 fimbriae |
Species-specific carbohydrate(s) |
Intestinal epithelium |
Diarrhea |
|
Uropathogenic E. coli |
Type 1 fimbriae |
Complex carbohydrate |
Urethral epithelium |
Urethritis |
|
Uropathogenic E. coli |
P-pili (pap) |
Globobiose linked to ceramide lipid |
Upper urinary tract |
Pyelonephritis |
|
Bordetella pertussis |
Fimbriae ("filamentous hemagglutinin") |
Galactose on sulfated glycolipids |
Respiratory epithelium |
Whooping cough |
|
Vibrio cholerae |
N-methylphenylalanine pili |
Fucose and mannose carbohydrate |
Intestinal epithelium |
Cholera |
|
Treponema pallidum |
Peptide in outer membrane |
Surface protein(fibronectin) |
Mucosal epithelium |
Syphilis |
|
Mycoplasma |
Membrane protein |
Sialic acid |
Respiratory epithelium |
Pneumonia |
|
Chlamydia |
Unknown |
Sialic acid |
Conjunctival or urethral epithelium |
Conjunctivitis or urethritis |
Table 5.
BACTERIA FOUND IN THE LARGE
INTESTINE OF HUMANS
|
BACTERIUM |
RANGE OF INCIDENCE
(%) |
|
Bacteroides
fragilis |
100 |
|
Bacteroides
melaninogenicus |
100 |
|
Bacteroides
oralis |
100 |
|
Lactobacillus |
20-60 |
|
Clostridium
perfringens |
25-35 |
|
Clostridium
septicum |
5-25 |
|
Clostridium
tetani |
1-35 |
|
Bifidobacterium
bifidum |
30-70 |
|
Staphylococcus
aureus |
30-50 |
|
Streptococcus
faecalis |
100 |
|
Escherichia
coli |
100 |
|
Salmonella
enteritidis |
3-7 |
|
Salmonella
typhi |
0.00001 |
|
Klebsiella
species |
40-80 |
|
Enterobacter
species |
40-80 |
|
Proteus
mirabilis |
5-55 |
|
Pseudomonas
aeruginosa |
3-11 |
|
Peptostreptococcus |
common |
|
Peptococcus |
moderate |
|
Methanogens |
common |
The intestinal microflora undergoes essential changes with the age of man.
The intestinal tract of the newly-born baby in the first hours of life is
sterile. During the first days it becomes inhabited by temporary microflora
from the environment, mainly from breast milk. Later on, in the intestine of
the newly-born baby a specific bacterial flora is established consisting of
lactic acid bacteria (biphidobacteria, acidophilic bacillus), which is retained
during the year. It has antagonistic properties in relation to many microbes
capable of causing intestinal disorders in breastfed children, and remains
during the whole period of breast feeding. However, on the 3rd-5th day of life
in the intestine of breast-fed children E.
coli and enterococci can be found, the amount of which sharply increases with
the change to mixed feeding. After breast feeding is stopped the microflora of
the child's intestine is completely replaced by a
microflora typical of adults (E. coli, Clostridium perfringens, Clostridium
sporogenes, Streptococcus faecalis. Proteus vulgaris, etc.).
At present it has been established that such a constant inhabitant of
the intestine of man as Clostridium perfringens is capable of secreting
digestive enzymes. The colibacillus and other species of microbes in the
intestine produce the vitamins essential for the human body (B1, B2,
B12, K). Microbe antagonists (acidophilic, Lactobacillus bulgaricus
etc.) are beneficial to the organism as they hinder the development of
pathogenic bacteria which, together with infected food and water, may enter the
intestine.
The pathogenic serotypes of E. coli which are capable of causing severe
diseases (colienteritis) mostly in children have been found to be present in
the human intestine together with the on pathogenic species.
Anaerobic bacteria which do not produce spores, the so-called bacteroids,
inhabit mainly the lower part of the large intestine in human. They are found
during acute appendicitis, postpartum infection, pulmonary abscesses,
septicaemia of different aetiology, postoperative infectious complications in
the peritoneal cavity, inflammatory processes of the gastrointestinal tract,
respiratory tract, and on the skin
Metchnikoff considered some species of intestinal bacteria to be
harmful, causing chronic intoxications. He suggested the method (if combating
them by introducing lactic acid bacilli (Lactohacillus bulgaricus) bearing
antagonistic properties into the intestine. Besides, Metchnikoff recommended a
diet of vegetables and fruit, rich in sugar, and considered it advisable to
build one's life according to the principles of orthobiosis (normal work,
healthy relaxation, hygienic conditions, and prophylaxis of diseases).
Enteroviruses live in large quantities in the intestine. They may be
found for a long time in healthy persons without causing diseases. In
unfavourable conditions associated with some species of bacteria they cause the
most varied clinical forms of disease.
Microflora of the
respiratory tract. People
breathe in a large number of dust particles and adsorbed micro-organisms.
Experimentally, it has been established that the amount of microbes in inspired
air is 200-500 times greater than in expired air Most of them are trapped in
the nasal cavity and only a small amount enters the bronchi, The pulmonary
alveoli and the terminal branches of bronchi are usually sterile. The upper
respiratory tract (nasopharynx, pharynx) contains relatively constant species
(Staphylococcus epidermidis, streptococci, diphtheroids, Gaffkya tetragena,
etc.).
When the defense mechanisms of the body are weakened as a result of
cooling, starvation, vitamin deficiency, or traumas, the constant inhabitants
of the respiratory tract become capable of causing different diseases (acute
catarrhs of the respiratory tract, tonsillitis, pneumonia, bronchitis, etc.).
The nasal cavity contains a small amount of microbes. The mucous
membrane of the nose produces mucin and lysozyme which have a bactericidal
action. However, in spite of this, the nasal cavity has a relatively constant
microflora (haemolytic or nasal micrococcus, diphtheroids, non-haemolytic
staphylococci, haemolytic staphylococci, saprophytic Gram-negative diplococci,
capsular Gram-negative bacteria, haemoglobinophilic bacteria of influenza,
Proteus, etc). In the respiratory tract, besides the bacterial microflora. many
viruses, in particular adenoviruses. can remain viable for long periods without
causing pathological processes.
Microflora of the
vagina. In the first 2 days after birth the baby's vagina is
sterile. Sometimes it contains a small amount of Gram-positive bacteria and
cocci. After 2-5 days of life the coccal microflora becomes fixed and remains
constant until puberty, when it is replaced by Dodderlein's lactic acid
bacilli.
During the menstrual cycle the contents of the vagina become alkaline which
is favourable for the development of coccal microflora. During sexual life the
microflora of the vagina changes, and many microbes appear which are introduced
from outside.
The microflora of the vagina undergoes profound changes during
gynaecological diseases (endometritis, metritis, ovaritis, etc.), and after
abortions.
The vaginal contents of the healthy woman have a relatively high
concentration of sugar and glycogen, and a low content of the diastatic enzyme
and proteins. The pH is 4.7 during which all other microbes except for
Doderlein's lactic acid bacilli, cannot develop.
As has been established, the acid medium of the vagina depends on the
presence of glycogen which under the influence of vaginal bacteria is
transformed into mono- and disaccharides and then into lactic acid. The amount
of glycogen depends on the function of the ovaries and the condition of the
whole body.
Vaginal bacteria have antagonistic properties; because of this, normal
microflora should be protected and should not be exposed to the harmful effect
of medicines (antibiotics, sulphonamide preparations, rivanol, osarsol.
potassium permanganate, etc.) to which Doderlein's lactic acid bacilli are more
sensitive than the bacteria against which these substances are employed.
Microflora of the
urinary tract. In men in the anterior part of the urethra there
are Staphylococcus epidermidis. diphtheroids and Gram-negative non-pathogenic
bacteria. Mycobacterium smegmatis and mycoplasmas are found on the external
parts of the genitalia, and also in the urine of men and women.
The female urethra is usually sterile, in some cases it contains a small
amount of non-pathogenic cocci.
The bacteria of
the mucous membranes of the eyes
include Staphylococcus epidermidis, Corynebacterium xerosis, mycoplasmas, etc.
When the organism is weakened or when there are visual disturbances and vitamin
deficiency, the normal inhabitants of the mucous membranes may become
relatively pathogenic and may cause different diseases of the mucous membrane,
such as conjunctivitis, blepharitis and other suppurative processes.
The normal microflora is not constant but depends on the age, nutrition
and general condition of the macro-organism. The microflora of the human body
undergoes profound changes, especially during various diseases.
Disturbances in the species composition of the normal microflora
occurring under the influence of
infectious and somatic diseases, and long-term and irrational use of antibiotics
bring about the state of dysbacteriosis. This is characterized by disturbances
in the assimilation of products of digestion, by changes in the enzymatic
processes, and by the cleavage of ready-made physiological secretions. The
territorial deviations of microflora cause a whole series of complications:
intestinal dyspepsia, toxinfections, suppurative processes, catarrhs of the
respiratory tract, pneumonia, candidiasis, etc. In dysbacteriosis the number of
lactic acid bacteria is diminished, the number of anaerobes increased, Gram-positive
bacteria change to Gram-negative and Gram-negative to Gram-positive, etc.
The question arises whether animal life is possible without microbes, It
was already known in the last century that micro-organisms were very rarely
found in birds and animals of the arctic regions. There were cases with
absolutely no microflora found in the body of some birds. Pasteur made an
attempt to raise amicrobic animals, but with the level of techniques of that
time the problem could not be solved.
A new branch of biology, gnotobiology, is now developing. It studies the
microbe-free organisms. Amicrobic chicks, rats, mice, guinea pigs, sucking
pigs. and other animals have been reared.
Amicrobic animals, or gnotobiotes,
are subdivided into several groups, monobiotes. (absolutely microbe-free
animals), dibiotes (animals infected with one microbial species), polyobiotes
(animals harbouring more than one species of microbes in their body).
Germ-free (gnotobiotic) animals
are good experimental models for investigating the interactions of animals and
microorganisms. To determine the role of the normal microbiota, animals can be
delivered by aseptic Caesarean section (the surgical removal of the fetus from
the uterus via the abdomen) so they will not be contaminated by the normal
microbiota of the vagina and birth canal during vaginal delivery. Germ-free
animals can been be raised in the absence of microorganisms by being kept in a
sterile environment. They are fed sterile food and water and given sterile air
to breathe. Comparing animals possessing normal associated microbiota with
germ-free animals permits the exploration of the complex relationships between
microorganisms and host animals.
Germ-free animals develop abnormalities of the gastrointestinal tract.
They are more susceptible to disease than animals with normal associated
microbiota. Germ-free animals are more susceptible to bacterial infection.
Organisms such as Bacillus subtilis and Micrococcus luteus, which are harmless
to other animals, cause disease in germ-free animals. More exotic pathogenic
microorganisms such as Vibrio cholerae and Shigella dysenteriae are far more
readily able to establish infections where there are no normal microbiota. They
do not have to compete for survival within the intestinal tract. At the same
time, though, germ-free animals are resistant to Entamoeba histolytica, the
causative organism of amebic dysentery.
This is because this protozoan requires normal intestinal bacteria as a food
source. Likewise, tooth decay is no problem to germ-free animals, even those on
high sugar diets, because they do not have lactic acid bacteria — the
bacteria that cause tooth decay — in
their oral cavities.
Scientists
focused their attention at gnotobiotes because it was necessary to study deeply
the role of normal microflora in the mechanisms of infectious pathology and
immunity. As compared to the commonly encountered animals, gnotobiotes have a
larger caecum, underdeveloped lymphoid tissue, internal organs of lesser weight,
smaller blood volume, a reduced content of water in the tissue and of
antibodies in blood serum.
Gnotobiology provides the means for revealing more precisely the role of
normal microflora in the synthesis of vitamins and amino acids, in the
production of congenital and acquired immunity, and the relationship of
bacteria and viruses. Much importance is attached to gnotobiology in the study
of space and the life conditions of man and animals during space flight.
Microbiological investigation of soil. For this purpose it is necessary to
select most typical area not more then
After careful mixing take an average sample of weight 100 –
Collection of soil
Examination of the
general microbe number
Microbiological investigation of water. The sanitary - bacteriological investigation of
water includes determination of total number of microbes in 1 ml of water,
determination of a coli-index or coli-titer, and detection of pathogenic
microbes, their toxins and Е. соli bacteriophages.
There are quantitative parameters of
faecal pollution: a coli-titre and coli-index. Coli-titre is an index, which
characterise an amount of water in
millilitres which contain one E.coli. Coli-index characterises a number of E. coli in
one litre of water.
The important part of investigation
is taking of water samples. This procedure can do only by special persons
(sanitary doctor or his assistant).
The samples of water for investigation are taken in
Water collection
Water cocks, pumps, tubes previously
must be sterilize by a flame using a cotton plug moistened before with alcohol.
Then the water on for 10 min to wash off the bacteria which are on their
surface inside. From the open reservoirs water for investigation is taken with
the bathometer. This instrument is metal frame with a sterile bottle inside. It
gives the possibility to take the water samples from any depth. Usually water
is taken from the depth of 15 cms. The bottles with samples of water should
have labels, which should mark the source, place of sample,
time (day, hour), surname of the person who has taken the sampleassay,
and also purpose of research.
Water is delivered to the laboratory immediately after
its receival. The transportation should be done within 2 hours. When doctor examines chlorinated
water, it is taken in sterile bottles with 2 mL of 1,5 % of solutions of Sodium hyposulphitum (Na2S2Оз
• 5Н20). Such quantity of Sodium
hyposulfite binds chlorine at its concentration up to 2 mg/L in 500 ml. In summer water transports in ice,
and in winter in special hot-water bottle (temperature must be 1–5 °C). After delivery of samples to the laboratory
the investigation begins. It includes
determination of total number of microbes and determination of a coli-titre and
coli-index.
Quantitative Analysis.
Bacteria cannot be accurately counted by microscopic examination unless
there are at least 100 million (108) cells per mililiter Natural
bodies of water, however, rarely contain more than 105 cells per
millilitre. The method employed is therefore the plate count. A measured volume
of water is serially diluted (see below), following which 1 mL from each
dilution tube is plated in nutrient agar and the resulting colonies counted.
Since only cells able to form colonies are counted, the method is also known as
the "viable count".
A typical example of serial dilution would be the following. One
millilitre of the water sample is aseptically transferred by pipette to 9 mL of
sterile water. The mixture is thoroughly shaken, yielding a 1:10 dilution (For
obvious reasons, this is also known as the "10–1"
dilution). The process is repeated serially
until a dilution is reached that contains between 30 and 300 colony-forming
cells per millilitre, at which point several 1-mL samples are plated in a
nutrient medium. Since the original sample may have contained up to 1 million
(106) viable bacteria, it is necessary to dilute all the way to 10–5,
plate 1-mL samples from each dilution tube, and then count the colonies only on
those plates containing 30-300 colonies. The reasons for these numerical limits
are that with over 300 colonies the plate becomes too crowded to permit each cell
to form a visible colony, whereas with below 30 colonies the percent counting
error becomes too great. (The statistical error of sampling can be calculated
as follows' The standard deviation of the count equals the square root of N, where N equals the average
of many samples. Ninety-five percent of all samples will give counts within 2
standard deviations of the average. For example, if the average count is 36,
then 95% of all samples will lie between 24 and 48 [36 ± 12]. In other words,
within 95% confidence limits a sample count of 36 has an error of plus or minus
33%.) Assume that the above procedure has been carried out with the results
shown in Table 3. The 10–3
dilution has a suitable number of colonies, the others being either too high or
too low for accuracy. The original water sample is calculated to have contained
72,000 (72 x 103) viable cells per millilitre.
Qualitative
Analysis.
The methods of plating and enrichment culture are used to obtain a picture of the
aquatic bacterial population. Although such methods are satisfactory for
general biologic studies, they are inadequate for the purpose of sanitary water
analysis; this involves the detection of intestinal bacteria in water, since
their presence indicates sewage pollution and the consequent danger of the
spread of enteric diseases. Since any enteric bacteria would be greatly
outnumbered by other types present in the water samples, a selective technique
is necessary in order to detect them. Two widely used procedures for sanitary
water analysis are as follows:
Table.
Example of a viable count
|
Dilution |
Plate
Count* |
|
Undiluted |
Too crowded |
|
10-' |
to count |
|
10 2 |
510 |
|
10 3 |
72 |
|
10 4 |
6 |
|
10 5 |
1 |
*Each count is the average of 3 replicate plates
1.Tube method.
Dilutions of a water sample are inoculated into tubes of a medium which is elective for coliform bacteria and in which all
coliform bacteria but few noncoliform bacteria will form acid and gas. Such
media include MacConkey's medium, which contains bile salts as inhibitors of
noncoliform bacteria; lactose-containing media; and glutamatecontaining media.
Cultures showing both acid and gas may then be subjected to further tests to
confirm the presence of Escherichia coli
or closely related enteric gram-negative rods. Such tests include streaking
cultures on a lactose-peptone agar containing eosin and methylene blue (EMB
agar), on which E coli forms
characteristic blue-black colonies with a metallic sheen; subculturing at
2. Membrane
filtration method. A large measured volume of water is
filtered through a sterilized membrane of a type that retains bacteria on its
surface while permitting the rapid passage of smaller particles and water
(fig.1). The membrane is then transferred to the surface of an agar plate
containing a selective differential medium for coliform bacteria (fig. 2). Upon
incubation, coliform bacteria give rise to typical colonies on the surface of
the membrane. The advantages of this method are speed (the complete test takes
less than 24 hours) and quantitation, the number of coliform cells being
determined for a given volume of water.
Figure. Water samples (100 ml)
are passed through bacteriological filters (0,2 to 0,45 mm pore size) to trap bacteria The filters with trapped
bacteria are placed on a medium containing lactose as a carbon source, an
inhibitor to suppress growth of noncoliforms and indicator substances to facilitate
differentiation of coliforms. Coliform bacteria form distinct colonies on Endo
medium
Figure. Colonies of Е. coli on
membranous filters.
The drinking water should
not have more than 100 microbes in 1 ml. The microbic number in water of wells
and open reservoirs can be up 1000.
During determination of a
coli-index and coli-titre of water it is necessary to take into consideration
the ability of Е. coli of the man and
animal to grow at 43 °C
Microbiological investigation of the air. The sanitary - hygienic investigation of the microflora
of the air includes determination both the total number of microbes in
Plate method (sedimentation method). The Petry’s dishes with meat-peptone agar or another
special nutrient media for staphylococci and streptococci, for example blood
agar, yolk-salt agar are used. They stay in open form at various height from a
floor. It is recommend to take one sample on every
Place the dishes into 37 °C incubator for 24 hrs and then
incubate for 48 hrs at room temperature (18-20 °C). Study colonies, count them,
and isolate pure culture of different microbes.
According to Omeliansky data on
a surface of medium by 100 cm2 sedimentate in 5 minutes as so many
microbes, as they present in
x =
(32 • 100) : 78,5 = 40
This quantity of microbes
contains in
There is a special table for
determination of total number of microbes in
Table
Account of bacteria number in
|
no |
Dish diameter |
Dish area (cm2) |
Multiplier |
|
1 |
8 |
50 |
100 |
|
2 |
9 |
63 |
80 |
|
3 |
10 |
78,5 |
60 |
For
determination of microbial dissemination
degree quantity of the colonies on the dish surface which have been counted
should be multiplied with one of multiplier.
If on a Petry’s dish (78,5 cm2) at a
10-minute exposure 40 colonies have grown, the
quantity of microbes in
Aspiration method. It is
based on a shock action of an air jet about a surface of a medium. Krotov’s
apparatus is used for this purpose. It give us the possibility to let pass 50
–100 L of air with a speed of
For example, 250 colonies are revealed on the surface of dish after 2-minutes
exposure with a 25 l/min speed. Thus the number of microbes
(x) in
There are temporary standards of a sanitary - hygienic state of the
air: in operating room the total number
of microbes prior to the beginning of the operation must be no more than
In preoperative and dressing
rooms limiting number of microbes prior before the beginning of work – 750
microbes in
SUPPLEMENT
http://interactive.usask.ca/Ski/agriculture/soils/soilliv/soilliv_micfl.html
http://www.google.com/search?hl=en&q=soil+microflora
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=647419&dopt=Abstract
http://jac.oxfordjournals.org/cgi/content/abstract/46/suppl_1/41
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=3103209&dopt=Abstract
www.gsbs.utmb.edu/microbook/ch006.htm
www.pedresearch.org/cgi/content/full/54/5/739
References:
1.
Hadbook on Microbiology.
Laboratory diagnosis of Infectious Disease/ Ed by Yu.S. Krivoshein, 1989, P. 29-34.
2. Medical Microbiology and Immunology: Examination and Board Rewiew /W.
Levinson, E. Jawetz.– 2003.– P. 23-27.
3.
Review of Medical Microbiology /E.
Jawetz, J. Melnick, E. A. Adelberg/ Lange Medical Publication, Los Altos,
California, 2002. – P. 176-179.
5. Essential of medical
microbiology /W. A. Volk.. et al.– 5 ed., 1995.