Bacteria in health and disease

Laboratory – 1
1
INTRODUCTION
 Because most microbiological laboratory procedures
require the use of living microorganisms, an integral
part of all laboratory sessions is the use of aseptic

Bacteria in health and disease

techniques.

 The virulence of microorganisms used in the academic

laboratory environment has been greatly diminished
because of their long-term maintenance on artificial
media. But all microorganisms should be treated as
potential pathogens (organisms capable of producing
infectious diseases).

 Thus, microbiology students must follow aseptic

techniques for personal and environmental safety. 2
BASIC STEPS TO ACHIEVE ENVIRONMENTAL
SAFETY
 Upon entering the laboratory, place coats, books, and other paraphernalia in
specified locations-never on bench tops.

 Keep doors and windows closed during the laboratory session to prevent

Bacteria in health and disease

contamination from air currents (especially during preparation of culture
media).

 At the beginning and termination of each laboratory session, wipe bench tops
with a disinfectant solution provided by the instructor.

 Do not place contaminated instruments, such as inoculating loops, needles, and

pipettes on bench tops. Loops and needles should be sterilized by incineration,
and pipettes should be disposed of in designated receptacles.

 Upon completion of the laboratory session, place all cultures and materials in
the disposal area as designated by the instructor.

 Proper disposal of infectious waste (used gloves, contaminated tissue papers,

any disposable contaminated materials) in special red plastic bags.
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GENERAL RULES TO PREVENT ACCIDENTAL
INJURY AND INFECTION

 Wash your hands with liquid detergent and then with paper
towels upon entering and prior to leaving the laboratory.

Bacteria in health and disease

 Wear a paper cap or tie back long hair to minimize its
exposure to open flames.

 Wear a laboratory coat while working in the laboratory to

protect clothing from contamination or accidental discoloration
by staining solutions.

 Closed shoes should be worn at all times in the laboratory

setting.

 Never apply cosmetics or insert contact lenses in the

laboratory.
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 Do not smoke, eat, or drink in the laboratory.
GENERAL RULES TO PREVENT ACCIDENTAL
INJURY AND INFECTION – CONT’D

 Carry cultures in a test-tube rack on the bench tops when not in use.
This serves a dual purpose: to prevent accidents and to avoid
contamination of yourself and the environment.

Bacteria in health and disease

 Never remove media, equipment, or especially, bacterial cultures
from the laboratory.

 Immediately cover spilled cultures or broken culture tubes with

paper towels and then saturates them with a disinfectant solution
(1% Na hypochlorite). After 15 minutes of reaction time, remove the
towels and dispose them of in a manner indicated by the instructor.

 Report accidental cuts or burns to the instructor immediately.

 Do not lick labels. Use only self-stick labels for the identification of
experimental cultures.

 Speak quietly and avoid unnecessary movement around the 5

laboratory to prevent distractions that may cause accidents.
HAND HYGIENE
Hand washing should be done
1-Before putting on gloves in the
clinical environment.
2-After interacting with each
patient.
3-When hands are visibly dirty or
soiled with blood or other body
fluids.
HAND ANTISEPSIS:
 Rubbing hands with an alcoholic hand
preparations (solutions or gels)
 or washing hands with water and antiseptic
soap e.g. iodophore (betadine) or
chlorhexidine.
SURGICAL HAND ANTISEPSIS

 Washing hands and forearms with antimicrobial

soap for 2-6 minutes (Surgical Scrub)
 Rubbing hands and forearms with alcoholic
preparation after washing with plain soap (Surgical
Rub).
 High efficiency Mask
STANDARD FACE MASK e.g. N95 mask

 Protects the wearer Protects the wearer

from splashes into from infectious air
mouth or nose by borne particles, e.g TB
fluid.
 Protects the wearer
from microorganisms
that are transmitted
by droplets
 Gloves

Sterile gloves Disposable non sterile gloves (latex)

surgical procedures Worn when hands
may contact infectious
materials,
contaminated surfaces
or equipment.

Heavy duty gloves

• Washing contaminated items,

• Collecting and transporting wastes
• Performing environmental cleaning.
PRECAUTIONS OF HANDLING SHARPS
Used disposable needles must NOT be
bent, broken, recapped by two hands,
removed from disposable syringes, or
otherwise manipulated by hand before
disposal.
Used disposable needles must be placed in
puncture-resistant containers.
SAFE WASTE DISPOSAL
A- General waste B- Bio hazardous
waste includes items
or non-hazardous that are contaminated
waste: wastes that with blood and saliva
pose no risk of during treatment and
infection as papers, pose a risk to humans or
the environment
boxes, hand towels.
Types of hand hygiene:
Routine
Aseptic

Bacteria in health and disease

Surgical
Alcohol based hand rub

Putting on and taking off gloves.

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 IMPORTANT DEFINITIONS

 Pathogens:
- They are microorganisms capable of producing

Bacteria in health and disease

disease in man.

 Infection:
- Lodgment and multiplication of a microorganism
in or on the tissue of a host.

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TYPES OF INFECTIONS
 Primary infection:
It is initial infection of the host by a
microorganism.
 Re-infection:

Bacteria in health and disease

- It is the subsequent infection with the same
microorganism in the same host.
 Secondary infection:
- When the primary infection lowers the resistance of
the host and the latter gets infection with another
organism.
 Cross infection:
- An infection transfer from infected or colonized patient
or staff member to another patient or staff member in 16
hospital by direct or indirect contact.
TYPES OF INFECTIONS – CONT’D
 Health care associated infection:
An infection which is acquired during medical

Bacteria in health and disease

intervention and which was not present or
incubating at the time of admission.

 Subclinical infection:
- When the clinical symptoms of an infection are
not apparent.

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COMMENSALS (NORMAL BACTERIAL FLORA)
 They are the microorganisms which live in
complete harmony with the host without causing
any damage to it.

Bacteria in health and disease

 At birth skin and mucous membranes are sterile.
However these surfaces become colonized by
organisms present in the environment. The
organisms differ at different parts of the body.

 These organisms obtain their nutrition from the

secretions and waste products of the body.
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NORMAL FLORA AT VARIOUS SITES OF THE
BODY
Skin Staphylococcus epidermidis, S. aureus, Micrococcus, α-hemolytic
streptococcus, diphtheroids, some Gram negative bacilli

Bacteria in health and disease

Nares and S.aureus, S.epidermidis, Streptococcus spp, Neisseria spp
nasopharynx

Mouth S.epidermidis, S.aureus, Viridans streptococcus, Lactobacilli, Neisseria spp

Oropharynx S.epedermidis, S.aureus, Viridans streptococcus, Neisseria spp, Moraxella,

haemophilus, gram negative bacilli
Gastrointestinal Bacteroids, enterobacteriacae (E.coli), Lactobacilli, Enterococcus spp
tract
Genitourinary Lactobacillus spp, Bacteroids spp, S.aureus, S.epedermidis, Enterococcus spp,
tract Enterobacteriacae
Conjunctiva Diphtheroids, S.epedermidis, Neisseria, non haemolytic streptococcus
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SOURCES OF INFECTION
 Endogenous infections (auto infections):
- Organisms of normal flora are usually non
pathogenic, but occasionally they may lead to

Bacteria in health and disease

infection (e.g. Viridans streptococcus which are
normally present in the mouth can lead to infective
endocarditis when there is rheumatic heart disease
and injury to the oral cavity like tooth extraction).

 Exogenous infections:
- Most of infections are exogenous in origin. The sources
of exogenous infections are:
- Human cases and carriers.
- Animal cases and carriers.
- Insects.
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- The environment.
MODES OF SPREAD OF INFECTION
 Inhalation:
- Respiratory infections such as influenza, common cold, tuberculosis
are acquired by inhalation. These organisms are shed into the
environment by patients in secretions of nose or throat during

Bacteria in health and disease

sneezing, or coughing, or talking. These activities expel a spray of
droplets.

 Ingestion:
- Intestinal infections like typhoid fever, cholera, dysentery, food
poisoning, poliomyelitis, hepatitis A are acquired by ingestion of
contaminated water or food from the feces of infected patients or
carriers.

 Parenteral:
- Microorganisms can be transmitted through minor abrasions in the
skin as HBV, HCV, HIV in some occasions as blood transfusion with
contaminated blood or contaminated syringes in drug abusers or by
needle stick injury among health care workers.
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MODES OF SPREAD OF INFECTION –
CONT’D
 Contact:
- Infection may be acquired by direct contact and
indirect contact with patients. Direct contact as

Bacteria in health and disease

sexually transmitted diseases (venereal diseases as
syphilis and gonorrhoeae), and indirect contact as
operative wound infections by contaminated
instruments.

 Blood-sucking arthropods:
- As malaria which can be transmitted by mosquitoes.

 Congenital:
- Some microorganisms as toxoplasma, rubella virus,
HIV can cross the placenta and infect the foetus in
utero (vertical transmission). 22
Staphylococcus GNB – E.coli

Bacteria in health and disease

Mix Gram +ve and -ve

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Candida sp. Sterile swab

Bacteria in health and disease

Viridans streptococcus
Normal flora of the mouth

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SUMMARY
 Laboratory safety

Bacteria in health and disease

 Types of infections

 Normal flora at various sites of the body

 Modes of spread of infection

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PRACTICAL WORK
1- Microscopic and morphological demonstration:
of some important normal bacterial flora.
- Staphylococcus: gram positive cocci arranged in

Bacteria in health and disease

punches.
- Streptococcus: gram positive cocci arranged in chains.
- Pneumococcus: gram positive diplococci – flame
shaped.
- Neisseria: gram negativediplococci – coffee been
shaped.
- E.coli: gram negative bacilli, no special arrangement.
- Candida spp: gram like, oval shape, no special
arrangement.

2- Demonstration of mouth swab or nasal swab: 26

collection and culture (isolation).
REVIEW QUESTIONS
1- Enumerate three examples of normal bacterial
flora of the following sites: skin, mouth, and
gastrointestinal tract.

Bacteria in health and disease

2- What are the sources of infections?

3- Enumerate modes of spread of infections. Give

examples?

4- Define primary and secondary infection (or other

types of infections). 27
FURTHER READINGS
 www.textbookofbacteriology.net/normalflora.html

Bacteria in health and disease

 bioinfo.bact.wisc.edu/the
microbialworld/Normalflora.html

 www.rch.org.au/washup/prof.cfm

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