Laboratory Techniques in Microbiology
A number of techniques are routine in microbiology laboratories that enable microorganisms to be cultured, examined and identified.
[ندعوك للتسجيل في المنتدى أو التعريف بنفسك لمعاينة هذه الصورة]
An indispensable tool in any microbiology
laboratory is the inoculating loop. The loop is a piece of wire that is
looped at one end. By heating up the loop in an open flame, the loop
can be sterilized before and after working with bacteria. Thus, contamination of the bacterial sample is minimized. The inoculating loop is part of what is known as aseptic (or sterile) technique.
Another staple piece of equipment is called a petri plate. A petri
plate is a sterile plastic dish with a lid that is used as a receptacle
for solid growth media.
In order to diagnose an infection or to conduct research using a
microorganism, it is necessary to obtain the organism in a pure culture. The streak plate technique is useful in this regard. A sample of the bacterial population is added
to one small region of the growth medium in a petri plate and spread in
a back and forth motion across a sector of the plate using a sterile
inoculating loop. The loop is sterilized again and used to drag a small
portion of the culture across another sector of the plate. This acts to
dilute the culture. Several more repeats yield individual colonies. A colony can be sampled and streaked onto another plate to ensure that a pure culture is obtained.
Dilutions of bacteria can be added to a petri plate and warm growth medium added to the aliquot of culture. When the medium hardens, the bacteria grow inside of the agar.
This is known as the pour plate technique, and is often used to
determine the number of bacteria in a sample. Dilution of the original
culture of bacteria is often necessary to reach a countable level.
Bacterial numbers can also be determined by the number of tubes of media that support growth in a series of dilutions of the culture. The pattern of growth is used to determine what is termed the most probable number of bacteria in the original sample.
As a bacterial population increases, the medium becomes cloudier and
less light is able to pass through the culture. The optical density of
the culture increases. A relationship between the optical density and
the number of living bacteria determined by the viable count can be
established.
The growth sources for microorganisms such as bacteria can be in a liquid form
or the solid agar form. The composition of a particular medium depends
on the task at hand. Bacteria are often capable of growth on a wide
variety of media, except for those bacteria whose nutrient or
environmental requirements are extremely restricted. So-called
non-selective media are useful to obtain a culture. For example, in water quality
monitoring, a non-selective medium is used to obtain a total
enumeration of the sample (called a heterotrophic plate count). When it
is desirable to obtain a specific bacterial species, a selective medium
can be used. Selective media support the growth of one or a few
bacterial types while excluding the growth of other bacteria. For
example, the growth of the bacterial genera Salmonella and Shigella are selectively encouraged by the use of Salmonella-Shigella agar. Many selective media exist.
Liquid cultures of bacteria can be nonspecific or can use defined
media. A batch culture is essentially a stopped flask that is about one
third full of the culture. The culture is shaken to encourage the
diffusion of oxygen from the overlying air into the liquid. Growth occurs until the nutrients are exhausted. Liquid cultures can be kept growing indefinitely by adding
fresh medium and removed spent culture at controlled rates (a
chemostat) or at rates that keep the optical density of the culture
constant (a turbidostat). In a chemostat, the rate at which the bacteria
grow depends on the rate at which the critical nutrient is added.
Living bacteria can also be detected by direct observation using a light microscope,
especially if the bacteria are capable of the directed movement that is
termed motility. Also, living microorganisms are capable of being
stained in certain distinctive ways by what are termed vital stains.
Stains can also be used to highlight certain structures of bacteria, and
even to distinguish certain bacteria from others. One example is the
Gram's stain, which classifies bacteria into two camps, Gram positive
and Gram negative. Another example is the Ziehl-Neelsen stain, which
preferentially stains the cell wall of a type of bacteria called
Mycobacteria.
Techniques also help detect the presence of bacteria that have become
altered in their structure or genetic composition. The technique of
replica plating relies on the adhesion
of microbes to the support and the transfer of the microbes to a series
of growth media. The technique is analogous to the making of
photocopies of an original document. The various media can be tailored
to detect a bacteria that can grow in the presence of a factor, such as
an antibiotic, that the bacteria from the original growth culture cannot tolerate.
Various biochemical tests are utilized in a microbiology laboratory.
The ability of a microbe to utilize a particular compound and the nature
of the compound that is produced are important in the classification of
microorganisms, and the diagnosis of infections. For example, coliform
bacteria were traditionally identified by a series of biochemical
reactions that formed a presumptive-confirmed-completed triad of tests.
Now, media have been devised that specifically support the growth of
coliform bacteria, and Escherichia coli in particular.
Various laboratory tests are conducted in animals to obtain an idea of the behavior of microorganisms in vivo.
One such test is the lethal dose 50 (LD50), which measures the amount
of an organism or its toxic components that will kill 50 percent of the
test population. The lower the material necessary to achieve the LD50,
the more potent is the disease component of organism.
Medical Training and Careers in Microbiology
The world of microbiology overlaps the world of medicine. As a
result, trained microbiologists find a diversity of career paths and
opportunity in medicine.
[ندعوك للتسجيل في المنتدى أو التعريف بنفسك لمعاينة هذه الصورة]
Research in medical microbiology can involve clinical or basic science. Clinical microbiology
focuses on the microbiological basis of various diseases and how to
alleviate the suffering caused by the infectious microorganism. Basic
medical research is concerned more with the molecular events associated
with infectious diseases or illnesses.
Both medical training and microbiology contain many different areas
of study. Medical microbiology is likewise an area of many specialties. A
medical bacteriologist can study how bacteria can infect humans
and cause disease, and how these disease processes can be dealt with. A
medical mycologist can study pathogenic (disease-causing) fungi, molds and yeast to find out how they cause disease. A parasitologist is concerned with how parasitic microorganisms
(those that require a host in order to live) cause disease. A medical
virologist can study the diseases attributed to infection by a virus,
such as the hemorrhagic fever caused by the Ebola virus.
The paths to these varied disciplines of study are also varied. One
route that a student can take to incorporate both research training and
medical education is the combined M.D.-PhD. program. In several years of
rigorous study, students become physician-scientists. Often, graduates
develop a clinical practice combined with basic research. The experience
gained at the bedside can provide research ideas. Conversely,
laboratory techniques can be brought to bear on unraveling the basis of
human disease. The M.D.-PhD. training exemplifies what is known as the
transdisciplinary approach. Incorporating different approaches to an
issue can suggest treatment or research strategies that might otherwise
not be evident if an issue were addressed from only one perspective.
The training for a career in
the area of medicine and medical microbiology begins in high school.
Courses in the sciences lay the foundation for the more in-depth
training that will follow in university or technical institution. With
undergraduate level training, career paths can include research assistant,
providing key technical support to a research team, quality assurance
in the food, industrial or environmental microbiology areas, and medical
technology.
Medical microbiology training at the undergraduate and graduate
levels, in the absence of simultaneous medical training, can also lead
to a career as a clinical microbiologist. Such scientists are employed
in universities, hospitals and in the public sector. For example, the
United Kingdom has an extensive Public Health Laboratory Service. The
PHLS employs clinical microbiologists in reference laboratories, to
develop or augment test methods, and as epidemiologists. The latter are involved
in determining the underlying causes of disease outbreaks and in
uncovering potential microbiological health threats. Training in medical
microbiology can be at the Baccalaureate level, and in research that
leads to a Masters or a Doctoral degree. The latter is usually undertaken if the aim is to do original and independent research, teach undergraduate and graduate students, or to assume an executive position.
Medical technologists are involved in carrying out the myriad of
microbiological tests that are performed on samples such as urine, blood
and other body fluids to distinguish pathogenic microorganisms from the
normal flora of the body. This can be very much akin to detective work, involving the testing of samples by various means to resolve he identity of an organism based on the various biochemical behaviors.
Increasingly, such work is done in conjunction with automated
equipment. Medical technologists must be skilled at scheduling tests
efficiently, independently and as part of a team. Training as a medical technologist is typically at a community college or technical institution and usually requires two years.
As in the other disciplines of medical microbiology, medical technology is a specialized field. Histopathology is the examination of body cells or tissues to detect or rule out disease. This speciality involves knowledge of light and electron microscopic examination
of samples. Cytology is the study of cells for abnormalities that might
be indicative of infection or other malady, such as cancer. Medical immunology
studies the response of the host to infection. A medical immunologist
is skilled at identifying those immune cells that active in combating an
infection. Medical technology also encompasses the area of clinical biochemistry,
where cells and body fluids are analyzed for the presence of components
related to disease. Of course the study of microorganism involvement in
disease requires medical technologists who are specialized
microbiologists and virologists, as two examples.
Medical microbiologists also can find a rewarding career path in
industry. Specifically, the knowledge of the susceptibility or
resistance of microorganisms to antimicrobial drugs is crucial to the
development of new drugs. Work can be at the research and development
level, in the manufacture of drugs, in the regulation and licensing of
new antimicrobial agents, and even in the sale of drugs. For example,
the sale of a product can be facilitated by the interaction of the sales
associate and physician client on an equal footing in terms of
knowledge of antimicrobial therapy or disease processes.
Following the acquisition of a graduate or medical degree,
specialization in a chosen area can involve years of post-graduate or
medical residence. The road to a university lab or the operating room
requires dedication and over a decade of intensive study.
Careers in medical science and medical microbiology need not be
focused at the patient bedside or at the lab bench. Increasingly, the
medical and infectious disease fields are benefiting from the advice of consultants
and those who are able to direct programs. Medical or microbiological
training combined with experience or training in areas such as law or
business administration present an attractive career combination.
A number of techniques are routine in microbiology laboratories that enable microorganisms to be cultured, examined and identified.
[ندعوك للتسجيل في المنتدى أو التعريف بنفسك لمعاينة هذه الصورة]
An indispensable tool in any microbiology
laboratory is the inoculating loop. The loop is a piece of wire that is
looped at one end. By heating up the loop in an open flame, the loop
can be sterilized before and after working with bacteria. Thus, contamination of the bacterial sample is minimized. The inoculating loop is part of what is known as aseptic (or sterile) technique.
Another staple piece of equipment is called a petri plate. A petri
plate is a sterile plastic dish with a lid that is used as a receptacle
for solid growth media.
In order to diagnose an infection or to conduct research using a
microorganism, it is necessary to obtain the organism in a pure culture. The streak plate technique is useful in this regard. A sample of the bacterial population is added
to one small region of the growth medium in a petri plate and spread in
a back and forth motion across a sector of the plate using a sterile
inoculating loop. The loop is sterilized again and used to drag a small
portion of the culture across another sector of the plate. This acts to
dilute the culture. Several more repeats yield individual colonies. A colony can be sampled and streaked onto another plate to ensure that a pure culture is obtained.
Dilutions of bacteria can be added to a petri plate and warm growth medium added to the aliquot of culture. When the medium hardens, the bacteria grow inside of the agar.
This is known as the pour plate technique, and is often used to
determine the number of bacteria in a sample. Dilution of the original
culture of bacteria is often necessary to reach a countable level.
Bacterial numbers can also be determined by the number of tubes of media that support growth in a series of dilutions of the culture. The pattern of growth is used to determine what is termed the most probable number of bacteria in the original sample.
As a bacterial population increases, the medium becomes cloudier and
less light is able to pass through the culture. The optical density of
the culture increases. A relationship between the optical density and
the number of living bacteria determined by the viable count can be
established.
The growth sources for microorganisms such as bacteria can be in a liquid form
or the solid agar form. The composition of a particular medium depends
on the task at hand. Bacteria are often capable of growth on a wide
variety of media, except for those bacteria whose nutrient or
environmental requirements are extremely restricted. So-called
non-selective media are useful to obtain a culture. For example, in water quality
monitoring, a non-selective medium is used to obtain a total
enumeration of the sample (called a heterotrophic plate count). When it
is desirable to obtain a specific bacterial species, a selective medium
can be used. Selective media support the growth of one or a few
bacterial types while excluding the growth of other bacteria. For
example, the growth of the bacterial genera Salmonella and Shigella are selectively encouraged by the use of Salmonella-Shigella agar. Many selective media exist.
Liquid cultures of bacteria can be nonspecific or can use defined
media. A batch culture is essentially a stopped flask that is about one
third full of the culture. The culture is shaken to encourage the
diffusion of oxygen from the overlying air into the liquid. Growth occurs until the nutrients are exhausted. Liquid cultures can be kept growing indefinitely by adding
fresh medium and removed spent culture at controlled rates (a
chemostat) or at rates that keep the optical density of the culture
constant (a turbidostat). In a chemostat, the rate at which the bacteria
grow depends on the rate at which the critical nutrient is added.
Living bacteria can also be detected by direct observation using a light microscope,
especially if the bacteria are capable of the directed movement that is
termed motility. Also, living microorganisms are capable of being
stained in certain distinctive ways by what are termed vital stains.
Stains can also be used to highlight certain structures of bacteria, and
even to distinguish certain bacteria from others. One example is the
Gram's stain, which classifies bacteria into two camps, Gram positive
and Gram negative. Another example is the Ziehl-Neelsen stain, which
preferentially stains the cell wall of a type of bacteria called
Mycobacteria.
Techniques also help detect the presence of bacteria that have become
altered in their structure or genetic composition. The technique of
replica plating relies on the adhesion
of microbes to the support and the transfer of the microbes to a series
of growth media. The technique is analogous to the making of
photocopies of an original document. The various media can be tailored
to detect a bacteria that can grow in the presence of a factor, such as
an antibiotic, that the bacteria from the original growth culture cannot tolerate.
Various biochemical tests are utilized in a microbiology laboratory.
The ability of a microbe to utilize a particular compound and the nature
of the compound that is produced are important in the classification of
microorganisms, and the diagnosis of infections. For example, coliform
bacteria were traditionally identified by a series of biochemical
reactions that formed a presumptive-confirmed-completed triad of tests.
Now, media have been devised that specifically support the growth of
coliform bacteria, and Escherichia coli in particular.
Various laboratory tests are conducted in animals to obtain an idea of the behavior of microorganisms in vivo.
One such test is the lethal dose 50 (LD50), which measures the amount
of an organism or its toxic components that will kill 50 percent of the
test population. The lower the material necessary to achieve the LD50,
the more potent is the disease component of organism.
Medical Training and Careers in Microbiology
The world of microbiology overlaps the world of medicine. As a
result, trained microbiologists find a diversity of career paths and
opportunity in medicine.
[ندعوك للتسجيل في المنتدى أو التعريف بنفسك لمعاينة هذه الصورة]
Research in medical microbiology can involve clinical or basic science. Clinical microbiology
focuses on the microbiological basis of various diseases and how to
alleviate the suffering caused by the infectious microorganism. Basic
medical research is concerned more with the molecular events associated
with infectious diseases or illnesses.
Both medical training and microbiology contain many different areas
of study. Medical microbiology is likewise an area of many specialties. A
medical bacteriologist can study how bacteria can infect humans
and cause disease, and how these disease processes can be dealt with. A
medical mycologist can study pathogenic (disease-causing) fungi, molds and yeast to find out how they cause disease. A parasitologist is concerned with how parasitic microorganisms
(those that require a host in order to live) cause disease. A medical
virologist can study the diseases attributed to infection by a virus,
such as the hemorrhagic fever caused by the Ebola virus.
The paths to these varied disciplines of study are also varied. One
route that a student can take to incorporate both research training and
medical education is the combined M.D.-PhD. program. In several years of
rigorous study, students become physician-scientists. Often, graduates
develop a clinical practice combined with basic research. The experience
gained at the bedside can provide research ideas. Conversely,
laboratory techniques can be brought to bear on unraveling the basis of
human disease. The M.D.-PhD. training exemplifies what is known as the
transdisciplinary approach. Incorporating different approaches to an
issue can suggest treatment or research strategies that might otherwise
not be evident if an issue were addressed from only one perspective.
The training for a career in
the area of medicine and medical microbiology begins in high school.
Courses in the sciences lay the foundation for the more in-depth
training that will follow in university or technical institution. With
undergraduate level training, career paths can include research assistant,
providing key technical support to a research team, quality assurance
in the food, industrial or environmental microbiology areas, and medical
technology.
Medical microbiology training at the undergraduate and graduate
levels, in the absence of simultaneous medical training, can also lead
to a career as a clinical microbiologist. Such scientists are employed
in universities, hospitals and in the public sector. For example, the
United Kingdom has an extensive Public Health Laboratory Service. The
PHLS employs clinical microbiologists in reference laboratories, to
develop or augment test methods, and as epidemiologists. The latter are involved
in determining the underlying causes of disease outbreaks and in
uncovering potential microbiological health threats. Training in medical
microbiology can be at the Baccalaureate level, and in research that
leads to a Masters or a Doctoral degree. The latter is usually undertaken if the aim is to do original and independent research, teach undergraduate and graduate students, or to assume an executive position.
Medical technologists are involved in carrying out the myriad of
microbiological tests that are performed on samples such as urine, blood
and other body fluids to distinguish pathogenic microorganisms from the
normal flora of the body. This can be very much akin to detective work, involving the testing of samples by various means to resolve he identity of an organism based on the various biochemical behaviors.
Increasingly, such work is done in conjunction with automated
equipment. Medical technologists must be skilled at scheduling tests
efficiently, independently and as part of a team. Training as a medical technologist is typically at a community college or technical institution and usually requires two years.
As in the other disciplines of medical microbiology, medical technology is a specialized field. Histopathology is the examination of body cells or tissues to detect or rule out disease. This speciality involves knowledge of light and electron microscopic examination
of samples. Cytology is the study of cells for abnormalities that might
be indicative of infection or other malady, such as cancer. Medical immunology
studies the response of the host to infection. A medical immunologist
is skilled at identifying those immune cells that active in combating an
infection. Medical technology also encompasses the area of clinical biochemistry,
where cells and body fluids are analyzed for the presence of components
related to disease. Of course the study of microorganism involvement in
disease requires medical technologists who are specialized
microbiologists and virologists, as two examples.
Medical microbiologists also can find a rewarding career path in
industry. Specifically, the knowledge of the susceptibility or
resistance of microorganisms to antimicrobial drugs is crucial to the
development of new drugs. Work can be at the research and development
level, in the manufacture of drugs, in the regulation and licensing of
new antimicrobial agents, and even in the sale of drugs. For example,
the sale of a product can be facilitated by the interaction of the sales
associate and physician client on an equal footing in terms of
knowledge of antimicrobial therapy or disease processes.
Following the acquisition of a graduate or medical degree,
specialization in a chosen area can involve years of post-graduate or
medical residence. The road to a university lab or the operating room
requires dedication and over a decade of intensive study.
Careers in medical science and medical microbiology need not be
focused at the patient bedside or at the lab bench. Increasingly, the
medical and infectious disease fields are benefiting from the advice of consultants
and those who are able to direct programs. Medical or microbiological
training combined with experience or training in areas such as law or
business administration present an attractive career combination.
