TERMS Def.

CHAPTER 1
Microbiology Derived from the Greek words mikros (small), bios
(life), and logia (study of)
Microorganisms or Microbes Study of organisms that cannot be seen with the
naked eye
Categorized into two: 1) cellular: prokaryotes (bacteria,
cyanobacteria, and archeans); eukaryotic
(fungi, protozoa, algae)
2) acellular: viruses
Bacteriology Study of bacteria
Virology Study of viruses
Mycology Study of fungi
Parasitology Study of protozoa and parasitic worms
Phycology Study of algae
Immunology Study of the immune system and the immune response
Normal flora or indigenous flora Inhabit the human body
Robert Hooke Discovery of microscope; Discovered the cell-basic unit of
living organisms; cell theory: living organisms are made
up of cells
Anton Von Leeuwenhoek Single-lens microscope; animalcules; “Father of
Microbiology”; first provided accurate descriptions of
bacteria, protozoa, and fungi
Louis Pasteur Germ theory of disease; microorganisms were in the
environment and cause infectious diseases;
pasteurization: kills microorganisms in different types of
liquid
Robert Koch Proved Louis Pasteur’s theory; Koch’s Postulates
1800-1900 Golden Age of Microbiology
Edward Jenner Discovered the vaccine for smallpox
Joseph Lister Applied the theory to medical procedures paving the way
for the development of aseptic surgery
Paul Ehrlich Salvarsan; treatment of syphilis; “magic bullet”
Alexander Fleming Antibiotic penicillin from the mold Penicillium notatum
Microscope – optical instrument that can magnify organisms a hundredfold or even a
thousandfold
PARTS OF THE COMPOUND MICROSCOPE
Ocular Lens of eyepiece Topmost part of the microscope which is the lens the
viewer looks through to see the specimen
Revolving nose piece Located above the stage, it holds the objective lens
Diopter adjustment Used to change focus on one eyepiece in order to correct
any difference in vision between the two eyes
Body tube or head Connects the eyepiece to the objective lens
Arm Connects the body tube to the base of the microscope
Course adjustment General focus
Fine adjustment Fine-tunes the focus and increases the details of the
specimen
Objective lenses Geld in place above the stage by the revolving nosepiece
and are the lenses that are closest to the specimen;
contains 3 to 5 objectives ranging from 4X to 100X
Stage Located beneath the revolving nose piece; flat platform
 on which the specimen is placed
Stage Clips Situated above the stage; metal clips that hold the slide
in place
Stage Control Found beneath the stage; these knobs move the stage
either left or right or forward or backward
Aperture Hole in the middle of the stage that allows light from the
illuminator to reach the slide containing the specimen
On/Off switch Switch located at the base of the microscope that turns
the illuminator on or off
Illuminator Light source of the microscope
Iris diaphragm Found in the condenser; adjust the amount of light
coming through the condenser
Condenser Found beneath the stage, contains a lens system that
focuses light onto the specimen; gathers and focuses
light onto the specimen
Base Supports the microscope and it is where the illuminator
is found
TYPES OF MICROSCOPES
Compound Microscope Contains more than one magnifying lens; magnify objects
approximately a thousand times; visible light main
source; compound light microscope; compound
microscope
2 lens system:
Ocular lens – 10x
Second lens – located on the objective right above the
organism
Brightfield Microscope 1000 – 1500 times; visualize bacteria and fungi; specimen
appears dark against the surrounding bright viewer field
of this microscope
Darkfield Microscope Reflected light instead of transmitted light; appears
bright against a dark background; studying specimens
that are unstained or transparent and absorb little light;
outlining; used to view spirochetes
Phase-contrast Microscope Differences in refractive and light waves passing through
transparent objects assume different phases; Frits
Zernike, dutch scientist in 1934; contrast enhancing
optical technique in order to produce high contrast
images of specimens that are transparent which include
thin tissue slices, living cells in culture, and subcellular
particles (such as nuclei and organelles).
Differential Inference Contrast utilizes two beams of light instead of one and therefore
Microscope has higher resolution; Georges Nomarski in 1952; is
useful in examining living specimens when normal
biological processes might be inhibited by standard
staining procedures
Fluorescence Microscope makes use of ultraviolet light and fluorescent dyes called
fluorochromes; to shine against a dark background; s a
higher intensity of light source and this in turn excites a
fluorescent species; used to visualize structural
components of small specimens such as cells and to
detect the viability of cell populations; used to visualize
 the genetic material of the cell (DNA and RNA)
Confocal Microscope Confocal laser scanning microscope (CLSM) or laser
confocal scanning microscope (LCSM); an optical imaging
technique that increases optical resolution and contrast
of the micrograph by using a spatial pin hole to block out
of focus light in image formation; stained with a
fluorescent dye to make it emit or return light; It is also
useful in the study of cell physiology.
Electron Microscope utilizes a beam of electrons to create an image of the
specimen; German Engineer Ernst Ruska in 1933, which
had a resolution power of up to 50 nm; used to visualize
viruses and subcellular structures of the cell
Two Types
1. transmission electron microscope (TEM) – og
form, two dimensional, black and white images,
and magnifies objects up to 200,000 times
2. scanning electron microscope (SEM) -
interactions at the surface rather than
transmission; magnify bulk samples with greater
depth of view so that the image produced
represents the 3D structure of the sample, but
the image is still only black and white. Generally,
it can magnify the object 10,000 times.
Scanning Probe Microscope developed in the 1980s by the Swiss scientists Dr. Gerd
Binnig and Dr. Heinrich Rohre; study the molecular and
atomic shapes of organisms on a nanoscale.
Staining - facilitate visualization, staining procedures have been developed by various scientists;
staining procedures are meant to give color to the organisms, making them easier to see under
the microscope
Simple Stains make use of a single dye which can either be aqueous
(water based) or alcohol based; basic dyes such as
safranin, methylene blue, or crystal violet; stains give up
or accept hydrogen ion, leaving the stain positively
charged; Most bacterial cells and cytoplasm are
negatively charged and since the dye is positively
charged, it adheres readily to the cell surface enabling
the visualization of bacterial cell morphology
Differential Stains Differentiate one group of bacteria from another
1. Gram Stain - distinguishes gram positive bacteria
from gram negative bacteria, gram positive
bacteria stain blue or purple, while gram
negative bacteria stain red or pink; all cocci are
gram positive except Neisseria, Veilonella, and
Branhamella. On the other hand, all bacilli are
gram negative except Corynebacterium,
Clostridium, Bacillus, and Mycobacterium.
2. Acid-fast stain - stain used for bacteria with high
lipid content in their cell wall
a. Ziehl-Neelsen Stain - “hot method” because
it requires steam bathing the prepared
smear after addition of the primary dye; Acid
fast organisms will appear red on a blue
background.
b. Kinyoun Stain - “cold method” as it does not
utilize heat after addition of the primary
stain, which is oil-based. The acid fast
organisms will appear red on a green
background.
Special Stains Demonstrate specific structures in a bacterial cell;
metachromatic granules can be visualized using the
LAMB (Loeffler Alkaline Methylene Blue) stain. Other
special stains include Hiss stain (capsule or slime layer);
Dyer stain (cell wall), Fischer Conn stain (flagella), Dorner
and Schaeffer Fulton stain (spores) and India ink or
nigrosine (capsule of the fungus Cryptococcus
neoformans).
Culture Media s basically an aqueous solution to which all the necessary
nutrients essential for the growth of organisms are
added; physical state, chemical composition, and
functional type.
PHYSICAL STATE
Liquid Media broths, milk, or infusions, these are water-based
solutions that do not solidify at temperatures above the
freezing point; contain specific amounts of nutrients but
do not contain gelling agents such as gelatin or agar;
propagation of a large number of organisms,
fermentation studies, and other tests.
Semi-Solid Media exhibit a clot like consistency at ordinary room
temperature and contain agar at concentrations of 0.5%
or less that allows thickening of the media without
producing a firm substance; soft consistency similar to
custard and are best suited for culture of microaerophilic
bacteria or for the study of bacterial motility
Solid Media contain a solidifying agent such as 1.5%–2% agar, giving
them a firm surface on which cells can form discrete
colonies; used for isolation of bacteria and fungi or for
determining the colony characteristics of the organism
understudy. Solid media come in two forms: (a)
liquefiable (or reversible) solid media and (b) non-
liquefiable(or non-reversible) solid media.
CHEMICAL COMPOSITION
Synthetic Media contain chemically defined substances which are pure
organic and/or inorganic compounds; simple or complex,
depending on what supplement is added to it
Non-synthetic Media complex media that contain at least one ingredient that
is not chemically defined, which means that it is neither a
simple or pure compound; extracts of animals, plants, or
yeasts; Non synthetic media can support the growth of
more fastidious organisms
FUNCTIONAL TYPE
General Purpose Media primary isolation of a broad spectrum of microbes and
contain a mixture of nutrients that support the growth of
both pathogenic and non-pathogenic organisms;
peptone water, nutrient broth, and nutrient agar.
Enrichment Media contain complex organic substances such as blood,
serum, or special growth factors, and are designed to
increase the number of desired microorganisms without
stimulating the rest of the bacterial population;
fastidious or nutritionally exacting bacteria.
 A. Blood agar - contains general nutrients with 5%–
10% (by volume) blood added to a blood agar
base;
i. Beta Hemolysis – complete lysis of red
blood cells
ii. Alpha hemolysis – incomplete lysis of red
blood cells. Producing a greenish
discoloration of the blood agar around
the colonies
iii. Gamma hemolysis – no hemolysis
Selective Media contain one or more substances that encourage the
growth of only a specific target microorganism and
inhibit the growth of others; designed to prevent the
growth of unwanted contaminating bacteria or
commensals so only the target bacteria will grow
a. Thayer-Martin agar - contains the antibiotics
trimethroprim, nystatin, vancomycin, and
colistin. It is used for the isolation of Neisseria.
b. Mannitol Salt Agar – contains 10% NaCl and used
for the isolation of Staphylococcus aureus
c. MacConkey’s agar - promotes the growth of
gram negative bacteria, primarily those
belonging to the family Enterobacteriaceae, and
inhibits the growth of gram positive bacteria
through the addition of bile salts; both selective
and differential.
d. Löwenstein-Jensen medium – selective medium
used to recover Mycobacterium tuberculosis;
selective by the incorporation of malachite green
e. Saborauds‘s dextrose agar – used for the
isolation of fungi
Differential Media allow the growth of several types of microorganisms;
designed to show visible differences among certain
groups of microorganisms; variations in colony size or
color, changes in color of culture media, or formation of
precipitates or gas bubbles
Transport Media used for clinical specimens that need to be transported
to the laboratory immediately after collection; a prevent
the drying of specimen and inhibit the overgrowth of
commensals and contaminating organisms
Anaerobic Media media used specifically for organisms that cannot survive
in the presence of oxygen and require reduced oxidation
reduction potential and other nutrients.

CHAPTER 2: PROKARYOTIC AND EUKARYOTIC CELLS

Living cells can be classified into two general categories—prokaryotes and eukaryotes
Prokaryotes do not possess a true nucleus and membrane bound
organelles (e.g., bacteria)
Eukaryotes are those that possess a true nucleus and membrane
bound organelles. They are usually multicellular
 organisms and include plants, animals, fungi,
parasites, and algae

MEDICALLY IMPORTANT MICROORGANISMS

Viruses acellular organisms; outer surface is called capsid,
which is composed of repeating sub units called
capsomeres; never both RNA and DNA; rely on the
cellular machinery of the host cell for protein and
energy production
classified by:
1. Type of nucleic acid they possess
2. Shape of the capsid (isosahedral, helical,
polyhedral, or complex)
 3. Number of capsomeres
4. Size of the capsid
5. Presence or absence of an envelop
6. Type of host they infect
7. Type of disease they produce
8. Target cell or tropism
9. Immunologic or antigenic properties
Bacteriophages Special type of viruses that primarily infect bacteria
Similar to bacteria
1. Obligate intracellualar parasites
2. Similarly shaped like other viruses
3. They play a role in the acquisition of virulence
factors of certain bacteria
Bacteria Prokaryotic cells with majority having an outer
covering called the cell wall that is composed mainly of
peptidoglycan; both RNA and DNA; a possess a
nucleoid instead of a true nucleus, smaller ribosomes,
and lack mitochondria
May be categorized into
1. Gram-negative bacteria w/ cell wall
2. Gram-positive bacteria w/ cell wall
3. Acid-fast bacteria with lipid-rich cell wall
4. Bacteria without cell wall

Fungi eukaryotic cells with an outer surface composed

mainly of chitin. Their cell membrane is made up
mostly of ergosterol. Like bacteria, fungi possess both
DNA and RNA. Unlike bacteria, they possess a true
nucleus that is enclosed by a nuclear membrane and
mitochondria that function for ATP production. Fungal
ribosomes are also larger than bacterial ribosomes (80
Svedberg units)
Protozoa Representatives for parasites; outer surface called a
pellicle; unicellular organisms that usually divide
through binary fission, similar to bacteria; two
morphologic forms-cysts and trophozoites; both RNA
and DNA
Algae are eukaryotic organisms whose outer surface consists
primarily of cellulose; described as plant-like because
most of them have chlorophyll are thus capable of
photosynthesis; no true roots, stems, and leave

Diatoms are unicellular algae that inhabit both fresh and

saltwater; silicone dioxide; utilized in filtration
systems, insulation, and as abrasives
Dinoflagellates unicellular algae that are important members of the
phytoplankton group; contribute greatly to the oxygen
in the atmosphere and serve as important links in the
food chain; “red tide”; paralytic shellfish poisoning

CHAPTER 3: Bacteria Morphology

Coccus (pl. cocci) Spherical or round-shaped organisms (Staphylococcus)
Pairs – diplococci
 Chains – Streptococci
Clusters – staphylococci
Grps of 4 – tetrad
Grps of 8 - octad
Bacilli Rod-shaped organisms
Curved or spiral-shaped organisms Comma-shaped, spiral in shape, club-shaped
ENVELOPE STRUCTURES - contribute greatly to the oxygen in the atmosphere and serve as
important links in the food chain; serves to protect the bacteria from harsh environmental
conditions
Glycocalyx outermost covering of some bacteria; gelatinous
substance that is located external to the cell wall,
composed of polysaccharide or polypeptide, or both.
Capsule – strongly attached to the cell wall and slime
layer if it is loosely attached; defends from
phagocytosis; stimulate an antibody response from
the immune system; serves to protect the organism
from dehydration
Cell wall Murein sacculus; s principal components
peptidoglycan, which is also called murein or
mucopeptide; multi-layered in gram positive bacteria
and single layered in gram negative bacteria; provides
rigid support and gives shape to the bacteria; protects
the bacteria from osmotic damage and plays an
important role in cell division
Special Components of gram-positive 1. Teichoic acids - comprise major surface
cell walls antigens of gram positive organisms and can
elicit antibody response; teichoic acids
function for the attachment of the organism to
the host cell; also provide tensile strength to
gram positive bacterial cell walls
2. Polysaccharides - neutral sugars such as
mannose, arabinose, rhamnose, and
glucosamine; also includes some acidic sugars
such as glucuronic acid and mannuronic acid.
Special components of gram-negative 1. Outer membrane - a bi layered structure
cell walls where the inner leaflet is composed of a
lipopolysaccharide (LPS); special protein
channels that allow the passage of small or
low molecular weight hydrophilic substances
such as sugars and amino acids. Lipid A:
endotoxin activity; located in the outer leaflet
of the outer membrane
2. Lipoprotein - functions to anchor the outer
membrane to the peptidoglycan layer and
stabilizes the outer membrane.
3. Periplasmic space - fluid filled space between
the outer membrane and the inner plasma
membrane; contains enzymes for the
breakdown of large non transportable
molecules into transportable ones and
enzymes that serve to detoxify and inactivate
 antibiotics
Acid-fast cell wall composed of large amounts of waxes that are known
as mycolic acids; inner layer of the cell wall is also
made up of peptidoglycan but because the outermost
layer is lipid rich, cell walls of acid-fast organisms are
hydrophobic
Projecting Structures
Flagella thread like structures made up entirely of molecules of
the protein sub unit flagellin.; project from the capsule
and are organs for motility
a. Monotrichous (single polar flagellum)
b. Lophotrichous (tuft of flagella at one end of
the bacterium)
c. Amphitrichous (flagella at both ends of the
bacterium)
d. Pertitichous (flagella all around the bacterium)
e. Atrichous (no flagella)
Pili or Fimbriae rigid surface appendages found on many gram
negative bacteria; fine and short in comparison with
flagella;
pilins – structural protein sub-units
common pili – adherence to cell surface
sex pili – attachment to another bacterium during a
form of bacterial gene exchange called conjugation
Axial Filaments Endoflagella; found in spirochetes; e composed of
bundles of fibrils; arise from the ends of the bacterial
cell and spiral around the cell; rotate producing
movement of the outer sheath of the spirochetes
propelling them forward
Cytoplasmic membrane Cell membrane or plasma membrane; located beneath
the cell wall; sometimes called the cell sac because it
encloses the cytoplasm of the cell; selectively
permeable membrane that allows for transport of
selected solutes
In Aerobic organisms - the site of the electron
transport chain and serves as the site of ATP
production; serves the function of the mitochondria,
which are not found in prokaryotic cells
contains the enzymes needed for the biosynthesis of
DNA, cell wall components, and membrane lipids.
INTERNAL STRUCTURES
Nucleiod genetic material is packaged in a structure called the
nucleoid. Bacteria possess a single, circular, double
stranded DNA.
Mesosomes Cell division; involved in the secretion of substances
produced by bacteria
Ribosomes Protein synthesis; bacterial ribosome is smaller (70S)
Granules or Inclusion Bodies Serve for storage of food and energy
Endospores structures produced by many bacteria when they are
placed in a hostile environment; composed of
dipicolinic acid which confers resistance to heat,
 drying, chemical agents, and radiation; making it very
difficult to destroy;
Sporulation – process of spore production

CHAPTER 4: BACTERIAL GROWTH REQUIREMENTS

Growth an orderly and organized increase in the sum of all
components of the organism; entails the replication of
all cellular structures, organelles, and components
Microbial Growth concerned with the increase in the number of cells
and not an increase in the size of the organism
Nutritional Requirements
Carbon the structural backbone or skeleton of all organic
molecules
a. Autotrophs - utilize inorganic compounds and
inorganic salts as their sole carbon source
b. Organotrophs – sugars or glucose as their
carbon source
Chemoorganotrophs Most medically important bacteria
Nitrogen, Sulfur, Phosphorus are necessary for the synthesis of cellular materials
like proteins and nucleic acids;
Nitrogen + sulfur = required for the synthesis of
proteins
Nitrogen + phosphorus = essential for the synthesis of
nucleic acids and ATP
Inorganic Ions magnesium, potassium, calcium, iron, and trace
elements (e.g., manganese, zinc, copper, cobalt)
i. Magnesium - stabilizes ribosomes, cell
membranes, and nucleic acids; co-factor in
the activity of many enzymes
ii. Potassium - required for the normal
functioning and integrity of ribosomes and
participates in certain enzymatic activities
of the cell.
iii. Calcium – important component of gram-
positive bacterial cell wall and contributes
to the resistance of bacterial endospores
against adverse environmental conditions
iv. Iron - a component of cytochrome, a
component of the electron transport
chain, and functions as a co factor for
enzymatic activities
v. Trace Elements - a component of
cytochrome, a component of the electron
transport chain, and functions as a co
factor for enzymatic activities
Growth Factors Growth factors are essential to promote the growth
and development of the bacterial cell. These include
vitamin B complex and amino acids
Physical Requirements
Moisture/Water The bacterial cell is composed mainly of water. It
 serves as the medium from which bacteria acquire
their nutrients.
Oxygen used by aerobic bacteria for cellular respiration and
serve as the final electron acceptor;
i. Aerobes – utilize molecular oxygen
ii. Obligate anaerobes – cannot survive in the
presence of oxygen
iii. Facultative – both aerobic and anaerobic
iv. Microaerophiles - grow at low oxygen
tension but their rate of growth is
diminished
v. Capnophiles - some organisms though that
may require the addition of carbon dioxide
to enhance their growth
Temperature Enhanced enzyme activity requires certain
temperatures;
1. Thermophiles – x > 40 degr. C
2. Mesophiles – 20 – 40 degr. C
3. Psychrophiles – 10 – 20 degr. C
pH the extent of acidityor alkalinity of their environment;
alkalophiles – pH 8.4-9.0
neutrophiles – pH 6.5-7.5
Acidophiles – pH x < 6.0
Osmotic conditions determined by the salt concentration; normal
microbial cytoplasmic salt concentration is
approximately 1%;
halophiles – high salt concentrations for growth
osmophiles – require high osmotic pressure
BACTERIAL GROWTH CURVE - illustrates the phases in the growth of the population of bacteria
when they are grown in a culture of fixed volume; lag phase, log phase, stationary phase, and
death or decline phase
Lag Phase period of adjustment for the bacteria in the new
environment; no appreciable increase in the number
of microorganisms; show increased metabolic activity
in order to synthesize DNA as well as secrete enzymes
which might not be present in their new environment
but which are needed by the organism; attain their
maximum size toward the end of the lag phase; 1 – 4
hours
Log/Logarithmic/Exponential Phase rapid cell division, resulting in an increase in the
number of bacteria; exhibits high metabolic activity;
generation time or doubling time of the organism (i.e.,
the time required for the bacterial cells to double in
number) is determined; 8 hours
Stationary Phase period of equilibrium; rate of growth slows down,
nutrients start to deplete, and toxic wastes begin to
accumulate; some might die; number of living cells
equals the number of dead cells; gram-positive to
gram-negative; sporulation
Death or Decline Phase period of rapid cell death where the number of dead
cells is greater than the number of living cells; is due to
 the continuous depletion of nutrients and
accumulation of waste materials; few hours to a few
days

CHAPTER 5: NORMAL FLORA OF THE HUMAN BODY

Microbial Ecology Study of relationships between microorganisms and their
environment
Normal flora or Group of organisms that inhabit the body of a normal healthy
indigenous flora individual in the community
Resident flora organisms that are relatively of fixed types and are regularly found in
a given area of the body at a given age
Transient flora those that inhabit the skin and mucous membrane temporarily for
hours, days, or weeks and are derived from the environment
Skin constant contact with the environment, making it the most exposed
to microorganisms;
1) Lysozyme in the skin
2) Acidic pH of the skin due to sweat
3) Free fatty acids in sebaceous secretions
4) Constant sloughing off of the skin

Mainly bacteria and fungi

Divided into three regions:
1) axilla, perineum, and toe webs
2) hand, face, and trunk
3) upper arms and legs
Skin of the axilla, perineum and toe webs - having higher moisture
levels, higher body temperature, and higher levels of surface lipids;
regions have more microorganisms compared to the others and are
predominantly inhabited by gram negative bacilli
Hands, forearms, feet, legs – Staphylococcus epidermis and
Staphylococcus hominis
Mouth and Respiratory tongue and buccal mucosa are inhabited mostly by Streptococcus
Tract viridans group, which includes S. mutans, S. milleri, S. salivarius, and
S. sanguis
Potentially pathogenic organisms - Haemophilus. influenzae,
Streptococcus pneumoniae, Neisseria meningitidis and Mycoplasma
Upper respiratory tract – Neisseria meningitidis, Corynebacterium
diphtheriae, and Bordetella pertussis.
Conjunctiva very scanty because they are held in check by the flow of tears that
contain lysozyme; The lysozyme may interfere with the cell wall
synthesis of organisms; Neisseria, Moraxella, and Corynebacterium.
Staphylococci and streptococci may also be present.
Digestive Tract esophagus contains transient mouth flora;
helicobacter pylori – able to survive the acidic environment of the
stomach; most common cause of duodenal ulcer
Small Intestine Scanty flora due to the constant peristaltic movement of the
intestines; streptococci, lactobacilli, and Bacteroides which are all
transient.
Large Intestine Bacteroides fragilis (most common),Bifidobacterium/Lactobacillus
bifidum (predominant in breast fed infants), Eubacterium,
Peptostreptococcus, and Clostridium
Bottle-fed infants – lactobacillus acidophilus

Important roles of 1) synthesis of vitamin B complex and vitamin K

intestinal flora 2) conversion of bile into bile acids
3) competition with transient flora for nutrients
4) prevention of colonization of the intestines by transient flora
5) production of potentially pathogenic end-products of
metabolism that are toxic to transient flora
Genitourinary Tract anterior urethra, the predominant flora isolated are S. epidermidis,
enterococci, and diphtheroids.
Both males and females – Mycobacterium smegmatis
Penile urethra – Gardnerella vaginalis, bacteroides, and alpha
streptococci
Female urethra – sterile or contains Staphylococcus epidermis
Female infants – predominant vaginal flora Lactobacillus spp.
1 month of age – Staphylococcus epidermidis, Streptococci,
diphtheroids, and Escherichia coli
Predominant flora - Lactobacillus acidophilus, corynebacteria,
peptostreptococci, streptococci, Bacteroides, and staphylococci.
Lactobacillus crucial role in preventing gonococcal infection by producing lactic
acid that adds to the acidity of the vagina.
 CHAPTER 6: MEDICAL AND SURGICAL ASEPSIS
Infection control one of the major concerns that healthcare workers in healthcare
facilities and hospitals constantly address
Chain of infection how an individual acquires the infectious agents and includes the
infectious agent, the source of infection or its reservoir, how the
organisms transmitted, and the organism’s portal of entry into the
susceptible host.
Mode of transmission the manner in which the infectious organisms is acquired by the
host.
Standard precautions specific measures used to prevent the spread of infection among all
patients and healthcare workers, including measures to protect
them from contaminated blood and other body fluids
Contamination denotes contact of a sterile or aseptic item with microorganisms.
Medically aseptic items become contaminated if they get in contact
with disease producing organisms
Decontamination Process where physical or chemical means are used to remove,
inactivate, or destroy pathogens on a surface or item making them
safe for handling or use and incapable of transmitting infectious
agents
Disinfection process of using physical or chemical means to destroy pathogens,
excluding the spores.
Sterilization
Antiseptic
Healthcare-associated
infection
Iatrogenic Infection
Occupation exposure
Personal protective
equipment (PPE)
Asepsis
Sepsis
Factors that a role in the
occurrence of infection
among patients
Most commonly occurring
pathogenic
microorganisms
Primary locations of
infections
Medical or clean asepsis
Surgical or sterile asepsis
General aseptic
procedures
Kamo nay basa ani ay
process by which all pathogens are destroyed, including the spores.
a chemical solution that inhibits the growth of some
microorganisms; used directly on skin
any infection that is acquired during the time a patient is admitted
in a healthcare facility.
infection that is acquired in the course of undergoing diagnostic
tests or therapeutic procedures.
the acquisition or exposure to an infectious agent of a healthcare
worker during the course of his/her work.
specialized equipment and attire used by healthcareworkers to
protect them from infections. These include gloves, masks, gowns,
and goggles.
a condition in which the individual and his/her surrounding
environment are free of any microorganisms
refers to the clinical condition where an individual develops a
systemic reaction to a bacterial infection that starts from a localized
infection in one part of the body
1. suppression of the immune system
2. prolonged duration of illness
3. procedures that patients undergo in the healthcare facility
such as insertion of in-dwelling catheters, use of antibiotics
and insertion of intravenous lines or endotracheal tubes
1. Escherichia coli,
2. Staphylococcus aureus
3. Pseudomonas aeruginosa,
4. Candida albicans
5. Enterococcus
surgical wounds, the urinary tract, the respiratory tract, and the
bloodstream.
Absence of disease-producing microorganisms; infection control
process that aims to reduce the spread of infection; involves certain
procedures aimed to decrease the number of organisms and prevent
their spread in the general clinical setting;
the absence of all microorganisms; aim to eliminate microorganisms
from an area in the body where surgical procedures will be
performed as well as the location where the surgical procedure will
be carried out
1. Frequent handwashing of hospital personnel
2. Prompt and safe disposal of contaminated materials like
bandages and needles
3. Regular checking and emptying of containers for surgical
drains
4. Prompt cleaning of soiled or moist areas
5. Proper labelling of containers regarding the date and time of
disposal
1. Handwashing
2. Personal Protective Equipment
a. Gloves
b. Masks
c. Sterile gowns
 Isolation Precautions
Isolation process of separating an individual with an infectious disease from
the rest of the healthy population to prevent the spread of the
infection to other individuals
Universal precautions 1. Proper handwashing
2. Use of PPE
3. Proper handling and disposal of secretions excluding sweat
4. Proper handling and disposal of soiled linen and equipment
5. Environmental control
6. Prevention of injury from sharp devices such as needles
7. Patient placement
Transmission-based Precautions – further prevent the spread of infectious agents
Contact precautions used to prevent the spread of infections or infectious agents that are
transmitted through touching of patients or items in the room where
the infectious agents may be deposited
Droplet Precautions used for diseases or infectious agents that are spread in tiny droplets
caused by coughing and sneezing; used to prevent contact with
secretions from the respiratory tract
Airborne precautions preventing the spread of diseases or infectious agents that are
spread through the air from one person to another
Kamoy basa oi Aseptic Measures in the Operating room and Preventing Infection in
the community

CHAPTER 7: PHYSICAL AND CHEMICAL METHODS OF STERILIZATION

Sterilization Process of killing or removing all microbial forms, including spores
Disinfection process by which most microbial forms on inanimate objects are
killed without necessarily destroying saprophytes and bacterial
endospores which leads to a reduction in the number of organisms
to a level that they cannot produce infection.
Antisepsis use of chemical agents on living tissue (e.g.,skin) to prevent the
spread of microorganisms either by inhibiting their growth or
destroying them.
Bactericidal or germicidal agent, physical or chemical, that kills bacteria.
agent
Bacteriostatic agent agent, physical or chemical, capable of inhibiting the growth of
bacteria without necessarily killing them
Sporicidal, fungicidal, agents capable of destroying spores, fungi, and viruses, respectively
viricidal
PHYSICAL MEHTODS OF STERILZATION
Heating Most common physical method of sterilization
Thermal Death Time Minimum time required to kill a suspension of an organism at a
predetermined temperature and environment
Mechanisms of action of 1. Formation of single-strand breaks in the bacterial DNA
heating 2. Coagulation and denaturation of proteins
3. Accumulation of toxic levels of electrolytes
4. Alteration of cell membrane structure
Several factors can affect 1. Nature of the heat
the process of sterilization 2. Temperature and time
through heating 3. Number of microorganisms
4. Nature of microorganisms
 5. Type of Material
6. Presence of organic material
Types of Heat 1. Moist Heat – preferred over dry heat because of its more
rapid killing action; main mechanism of action is to cause
coagulation and denaturation of proteins

a. Temperature below 100°C

i. Pasteurization - destroying disease producing
organisms in milk and milk products as well as
other beverages; Conventional method: milk is
heated at 60 °C–65 °C followed by rapid cooling.
Flash method: heating at 72 °C for 15 seconds
followed by quick cooling to 13 °C. UHT method:
heating is done at 140 °C for a period of 15
seconds and 149 °C for 0.5 seconds.

ii. Vaccine Bath - destroy contaminating bacteria in

vaccine preparations; heated in a water bath at
60 °C for one hour

iii. Serum Bath - inactivate bacteria contaminating

serum preparations and is done by heating at 56
°C for several successive days

iv. Inspissation - used to solidify and disinfect egg

containing and serum containing media; culture
medium is placed in the slopes of a device called
an inspissator and is heated at 80 °C–85 °C for
30 minutes for three successive days

b. Temperature of 100 °C
i. Boiling - utilizing water at boiling temperature of
100 °C; not sporicidal and will destroy only the
vegetative forms; killing action can be enhanced
by the addition of 2% sodium bicarbonate;
Certain metal articles and glasswares can be
disinfected using this method for 10–20 minutes
without opening the lid of the boiler.

ii. Fractional Sterilization (Tyndallization) - known

as intermittent sterilization and involves
exposing the material to be sterilized to live
steam at 100 °C for 30–90 minutes for three
consecutive days, depending on the material to
be sterilized; used to sterilize culture media such
as TCBS and selenite broth; vegetative forms are
killed on the first day and the spores that will
germinate will be destroyed on the next
successive days; vegetative forms of the bacteria
are destroyed with this method.
 c. Temperature above 100 °C
i. Autoclave (Steam under pressure) - most
efficient method of sterilization because it can
destroy all microbial forms; 15–20 minutes to
sterilize the material; used to sterilize
instruments, surgical bandages, culture media,
and other contaminated materials that can
withstand high temperature and high pressure.

2. Dry Heat - depends on the penetration of heat through the

material to be sterilized; used to sterilize materials in
enclosed tubes, oils, jellies, powders, and glasswares such as
test tubes and Petri dishes.
i. Red flame - sterilize articles like bacteriological
wire loops, straight wires, tips of forceps, and
searing spatulas; materials are held over the
flame of a Bunsen burner until they become red
hot; limited only to articles that can be heated to
redness in flame

ii. Open flame (flaming) - also makes use of the

Bunsen burner or alcohol lamp; sterilized is
passed over the flame several times but is not
heated to redness; aimed at burning the
organism into ashes and is used to sterilize such
articles as mouths of test tubes, scalpels, glass
slides, and cover slips; only vegetative forms are
destroyed; cracking of glassware

iii. Incineration - aimed at burning the organism

into ashes; Articles that must be incinerated
include soiled dressings and beddings, animal
carcasses, and pathological material; used only
for articles that have to be disposed

iv. Hot air Oven – Louis Pasteur; placed in the oven

with a temperature of 160 °C for a period of one
hour; sterilize metallic instruments such as
forceps, scalpels, and scissors; used to sterilize
certain glasswares (e.g., petri dishes, pipettes,
flasks) and it is the only method used to sterilize
powders and ointments

v. Infrared rays - articles to be sterilized are placed

in a conveyor belt and passed through a tunnel
that is heated by infrared radiators; 180 °C for a
period of 7.5 minutes; be used to sterilize
metallic equipment and glassware.
Dessication Depriving the microorganism of moisture; mainly for food
preservation, such as in the preparation of dried fish and fruits;
 destroy vegetative forms; endospores are resistant to drying
Freezing not a reliable method of sterilization because most pathogenic
organisms are resistant to low temperatures; main use in the
laboratory is for the preservation of microorganisms in a process
called lyophilization or freeze drying where the organism is rapidly
frozen then dehydrated in high vacuum and stored in a vacuum
sealed container
Filtration Form of mechanical sieving that does not kill microorganisms but
merely separates them from the fluid; cellulose ester filter with a
pore size of 0.22 μm–0.45 μm
Three smallest bacteria – Mycoplasma, Rickettsia and Chlamydia
used for liquid solutions that will be destroyed by heat or freezing
such as serum, antibiotic solutions, sugar solutions, or urea solution;
used to remove bacteria from culture media or to prepare
suspensions of viruses and phages
Radiation 1. Ultraviolet Light (UVL)/Non-ionizing radiation – effective UVL
wavelength is in the range of 200 nm–280 nm, with 260 nm
as the most effective; maximum absorption of bacterial
DNA; acts by inducing formation of thymine-thymine dimers
resulting in lethal frameshift mutations; used to disinfect
hospital wards, operating rooms, laboratories, and other
rooms in the hospital that need to be sterilized

2. Ionizing radiation - ionizing rays have greater penetrance

than UV rays; exerts its effect by causing formation of free
radicals that chemically interact with proteins and nucleic
acids, resulting in cell death
i. Electron beams – particulate in nature; used to
sterilize syringes, gloves, dressing packs, food,
and some pharmaceuticals; lower penetrance
and requires sophisticated instruments.

ii. Electromagnetic rays (Gamma rays) – produced

from nuclear disintegration of selected
radioactive isotopes; greater penetrance than
electron beams but require longer exposure
time; bactericidal, fungicidal, viricidal, and
sporicidal; used commercially to sterilize
disposable Petri dishes, plastic syringes,
vitamins, antibiotics, hormones, fabrics and
glassware.
Sonic and Ultrasonic Exposure to sound waves at a frequency of approximately20,000
Vibrations cycles/second for one hour can kill some bacteria and viruses; used
to disinfect and clean instruments and to reduce microbial load.
Osmotic Pressure when the concentration of the fluid surrounding the organism is
altered, this will cause the bacterial cell to collapse. This is used for
preservation of fruits in syrup and meats in brine.
CHEMICAL METHODS OF STERILIZATION - inhibit the growth of pathogenic organisms, either
temporarily or permanently
Several factors can affect 1) Concentration and potency of the chemical agent
the efficacy of a chemical 2) Duration of exposure
agent
Characteristics
CLASSIFICATION OF CHEMICAL DISINFECTANTS - (1) consistency (liquid or gaseous);(2) spectrum
of activity (high level, intermediate level, low level); or (3) mechanism of action.
(mechanism of action)
Damage to the cell
membrane
Surface active agents
Phenolic compounds
3) Temperature
4) Nature of surrounding medium
5) Nature of organism
6) Number of organisms/sizes of inoculum
1. It should be broad spectrum, able to destroy a wide variety
of microorganisms.
2. . It should be fast acting, able to destroy microbes within a
short period of time.
3. It should be active in the presence of organic matter.
4. It should be active in any pH. 5.
5. It should be stable Physical and Chemical Methods of
Sterilization
6. It should be nontoxic, non-allergenic, non-irritative, and
non-corrosive.
7. It should be soluble in water and easy to apply.
8. It should leave a residual antimicrobial film on the treated
surface.
9. It should have high penetrating power. 10.
10. It should not be expensive and must be easily available.
11. It should be safe under storage and shipping for reasonable
periods of time.
12. 12. It should not have a bad odor.
cause smaller molecules to leak out of the bacterial cell and
interfere with the active transport and energy metabolism within
the cell.
compounds have long chain hydrocarbons that are fat soluble and
charged ions that are water-soluble; concentration the surface of
membranes and disrupt membrane resulting in leakage of cell
components; e active against vegetative microbial forms including
Mycobacterias well as enveloped viruses; disinfectant in homes and
hospitals; activity is reduced in the presence of hard water and
organic matter
1. Cationic agents - detergents where the fat-soluble portion is
positively charged due to combination with a quaternary
nitrogen atom; quaternary ammonium compounds and are
effective at alkaline pH; cetrimide and benzalkonium
chloride
2. Anionic Agents - negatively charged agents that contain long
chain hydrocarbons; soaps and bile salts; remove dirt
through the process of emulsification and are most effective
at acidic pH.
act by disrupting cell membranes as well as causing precipitation of
proteins and inactivation of enzymes; e coal tar derivatives that act
as disinfectants at high concentration and as antiseptic at low
concentrations; bactericidal and fungicidal with good activity against
 Mycobacteria but have poor activity against spores and most
viruses.

A. Phenol is no longer used as a disinfectant because it is toxic

to human cells. It is used as a gold standard in the chemical
evaluation of new chemical agents using the phenol
coefficient test.

B. Cresols are phenol derivatives more potent and safer than

phenol. An example is Lysol®

C. Chlorhexidine is used as a skin disinfectant if in isopropanol

solution. The aqueous preparation is used for wound
irrigation. Its main use is as antiseptic hand wash.

D. Chloroxylenols are used for topical purposes. They are

effective against gram positive bacteria.

E. Hexachlorophenes a chlorinated diphenyl which has greater

activity against gram positive bacteria similar to
chloroxylenols.

F. Triclosan, an organic phenyl ether, has good activity against

gram positive bacteria and a number of gram-negative
bacteria including Pseudomonas. It has some activity on
fungi and viruses.
Alcohols disorganize the lipid structure of the cell membrane, dehydrate cells,
and cause denaturation and coagulation of cellular proteins; skin
irritants and are also flammable.

a. Ethyl alcohol - skin antiseptic, it is bactericidal and removes

lipids from skin surfaces

b. Isopropyl alcohol - greater bactericidal activity than ethyl

alcohol and is less volatile; used to disinfect surfaces;
inhalation of its fumes can cause narcosis

c. Benzyl alcohol – mainly used as a preservative

d. Methyl alcohol - fungicidal and sporicidal used in disinfecting

inoculation hoods.
Denaturation of cellular pave the way for the eventual destruction of the bacterial cell;
proteins Denaturing agents include:
(1) acids and alkalis,
(2) alcohol and acetone, and
(3) phenol and cresol
Heavy Metals cause damage to the enzyme activity of bacteria; o cause
precipitation of proteins and oxidation of sulfhydryl group/s

a. Mercurials (e.g., mercurochrome and merthiolate) - biocidal

and are used as antiseptics; active against viruses at dilution
 of 1:500 to 1:1000.

b. Silver compounds (e.g silver nitrate) – bactericidal; 1% silver

nitrate solution is used clinically as treatment for ophthalmia
neonatorum; Silver sulfadiazine is used topically in the
treatment of burn wounds
Halogens bactericidal oxidizing agents that cause oxidation of essential
sulfhydryl groups of enzymes causing inactivation of the enzymes

a. Iodine - (tincture of iodine, iodophors) is considered the best

antiseptic because it is sporicidal, bactericidal, fungicidal,
viricidal, and amoebicidal; combined with neutral carrier
polymers to produce iodophores

b. Chlorine - used in the treatment of water; Hypochlorites are

used for sanitizing dairy and food processing equipment

c. Hydrogen peroxide - weak antiseptic and used only for

cleaning wounds and in the disinfection of surgical devices
and soft plastic contact lenses.
Alkylating agents a. Aldehydes - damage nucleic acids by alkylation of amino,
carboxyl, or hydroxyl groups. It kills all microorganisms
including spores.
i. Formaldehyde - used for surface disinfection; used
to sterilize bedding and furniture; used to kill
Mycobacterium tuberculosis in sputum and fungi in
athlete’s foot.

ii. Glutaradehyde - sporicidal and used as a cold

sterilant in sterilizing medical equipment such as
respiratory therapy machines and other equipment
that can be damaged by heat; more potent than
aldehyde; requires alkaline pH for its action and
exposure time of at least 3 hours to be effective

iii. Ethylene oxide - sporicidal and is used in the

gaseous sterilization of heat sensitive materials or
equipment like heart lung machine, respiratory and
dental equipment, and polyethylene tubes in
anesthesia machines; more potent than
glutaraldehyde but slower acting; highly flammable
and is usually combined with 10% CO2. It causes eye
irritation and is mutagenic and carcinogenic.

CHAPTER 8: ANTIMICROBIAL AGENTS