Clean Room
1st Lecture

Dr. Mohamed Kamal

Lecturer of Microbiology
Botany and Microbiology Department
Faculty of Science, Cairo University
 Agenda
• ***IAQ ( indoor air quality)
• ***Nosocomial infection ( MDR, quorum sensing, …..)
• ***What is ( CLEAN ROOM )
• ***Clean Room Standards
• ***Definition of (clean room) According to Ideal Standards
• ***Classification Of Clean Rooms
• Clean Room Guide Lines
• Basic principles behind How a cleanroom works.
• Importunacy Of Clean Room In Different Fields.
• What’s the applications of Clean rooms (Types)
• How To Achieve (Clean Room ) Standards ?
• Bio Contamination Control
• Advantage of integration to reducing Microbial Resistance
INDOOR AIR QUALITY (IAQ)
 WHAT IS IAQ?

Indoor Air Quality refers to the nature of

conditioned (Heat/Cool) air that
circulates throughout the space/area
where we live, work, make our food and
treat our patients

i.e. the air we breathe during most of our lives.

 IAQ – an area of concern
• US EPA pointed out that indoor air pollution
poses a greater risk than outdoor air pollution -
people spend 80-90% of their time indoors (Yu
and Browers, 2013)

• Presence of air pollutants in indoor environment

a global issue due to adverse effects on human
health (Tsakas, Siskos and Siskos, 2011)

• Indoor air pollution - ranked among the top five

environmental health risks to the public by EPA.
 NIOSH categorized major sources, causes,
or factors of indoor air quality episodes as follows*:

• 50% related to deficiencies in building ventilation (e.g., lack of outside air,

poor air distribution, uncomfortable temperatures and humidity, and
contaminants in the system)

• 30% related to some indoor air contaminant

(e.g., formaldehyde, solvent vapors, dusts, microbiological agents)

• 10% were attributed to an outdoor source

(motor vehicle exhaust, pollen, fungi, smoke, construction dusts)

• 10% had no observable cause. However, microbial contamination is

suspected for about half of these.

*estimates are rounded to nearest 10% and assessed from 2,200 episodes since the
 Microbial Contamination Issues

• Some experts suggest the microbial

contamination is higher than estimated
because NIOSH evaluations did not always
consider microorganisms

• In addition, a number of recent studies

suggest that microbiological air contaminants
may be more important and widespread than
previously believed, especially in humid and
coastal areas.

Source: IAQ and HVAC Workbook, 4th edition, by D. Jeff Burton (IVE, Inc. 2011)
IAQ, refers not only to comfort, which is affected by:
• Temperature
• Humidity
• VOCs
But also to harmful Biological and Non Biological air borne particles
present in the conditioned space.
 IAQ

• Sick leave • Salmonella • Hospital Acquired

• Sick building • Costs for care • Listeria infection
syndrome (SBS) • • Clean room app.
health E. coli
• Diminished • Molds
• Building related performance at • Fungi
illness (BRI) work caused by
adverse health
• Costs of
investigation
 Health Effects
• Building related illness (BRI)
• Clinically recognized disease
• Exposure to indoor air pollutants
• Recognizable causes
• Building related illness:
• Pontiac fever - legionella spp.
• Legionnaire’s disease
• Hypersensitivity pneumonitis
• Asthma
• Allergy
• Respiratory disease
• Chronic obstructive pulmonary
disease (COPD)
 Health Effects

• SICK BUILDING SYNDROME • (SBS) SYMPTOMS

(SBS) • Eye, nose, or throat irritation
• A persistent set of symptoms in >
• Headaches
20%
• Cause(s) not recognizable • Fatigue
• Complaints/symptoms relieved • Irritability
after exiting building
• Dry skin

• Nasal congestion

• Difficulty breathing

• Nose bleeds

• Nausea
 Indoor Air Quality
(IAQ)
Fisk & Rosenfeld noticed improved productivity by
changing indoor environment.
Potential annual benefits were:
10-30% reduction in acute respiratory infections &
reduced allergy & asthma Symptoms
20-50% reduction in acute non-specific health
symptoms
0.5-5% increase in the performance of office work
Annual cost savings & productivity gains US$30-170
billion.
 Main Causes Of Poor Indoor Air Quality

Unplanned building design and usage.

Pollutant sources (indoor or outdoor)

Air-conditioning and ventilation

systems (contamination and
malfunction)
Factors affecting indoor air quality
management

Types of Airborne Particles

Sizes Of Airborne Particles

Sources Of Airborne Contamina
nts
 FACTORS AFFECTING IAQ

The indoor environment in any building is a result of the interaction between

the
site
climate
construction techniques
Ventilation system
contaminant sources
Pressurization and Airflow
building occupants

(USEPA and NIOSH, 1993).

Eight Sources Of Airborne Microorganisms

In The Built Environment
Source Source strength Reference
Humans 3.7 × 107 bacterial genome copies per person- [33]
houra
7.3 × 106 fungal genome copies per person-houra

31 mg total per person-houra

2.2 × 102 (median) and 2.6 × 105 (max) influenza [41]
genome copies (<5 μm) per person-houra
Pets TBD N/A
Plants TBD N/A
Plumbing systems TBD N/A
Heating, ventilation, and air- TBD N/A
conditioning (HVAC) systems
Mold TBD N/A
Resuspension of settled dust TBD from walking N/A
4 × 104 (median) and 7.4 × 105 (max) bacterial [99]
genome copies per min from vacuum cleaners

Outdoor air Depends on concentration in outdoor air, N/A

ventilation rate, and penetration efficiency
TBD to be determined (not yet reported in the literature), N/A not
 Airborne food
contamination a constant
worry for food
manufacturers

• Proper safety measures can help prevent the growth of

microorganisms and the accumulation of particulates
such as dust.

• Microorganisms that can harm food and, consequently,

people are airborne and live within droplets, according to
Food Safety Magazine.

As a best practice for food manufacturers,

facilities should safely remove airborne
contaminants and improve the air quality of
the building.
 Healthcare associated infections

(HAIs) (Nosocomial infections or hospital-
acquired infections)

• Healthcare associated infections (HAIs) kill

more people in this country than AIDS,
breast cancer and auto accidents
combined.
HOSPITAL MICROBIAL POLLUTION

Nosocomial infections (also known as hospital associated/acquired

infections) are those infections that develop in patients during their
stay in hospitals or other type of clinical facilities, which were not
present at the time of admission.

Bacterial pathogens that can able to survive in the hospital

environment for long period of time and resist disinfection are
particularly more important for nosocomial infections.

Bacterial pathogens isolated from hospital environments are also

known to develop resistance to multiple antimicrobial agents. The
emergence of multidrug resistance organisms in hospital resulted in
difficulty to treat nosocomial infections. Despite the advance in
modern medicine nosocomial infection still poses a risk of increased
morbidity and mortality to patients. For this, the hospital environment
may play a significant role. It is thereby important to identify
environmental surfaces that are rich in bacteria and have the
Nosocomial infections It may become clinically apparent either during
the hospitalization or after discharge. Hence, pathogens that cause such
infections are termed nosocomial pathogens. However, an
asymptomatic patient may be considered infected if pathogenic
microorganisms are found in a body fluid or at a body site that is
normally sterile, such as the cerebrospinal fluid or blood. Infections
acquired by staff or visitors to the hospital or other health care setting
and neonatal infection that result from passage through the birth canal
may also be considered nosocomial infections

Commensal bacteria found in normal flora of healthy humans. These

have a significant protective role by preventing colonization by
pathogenic microorganisms. Some commensal bacteria may cause
infection if the natural host is com- promised. For example, cutaneous
coagulase- negative staphylococci cause intravascular line infection and
intestinal Escherichia coli are the most common cause of urinary
infection
Pathogenic bacteria have greater virulence, and cause infections
(sporadic or epidemic) regardless of host status.

For example: Anaerobic Gram-positive rods (e.g. Clostridium) cause

gangrene. Gram positive bacteria: Staphylococcus aureus (cutaneous
bacteria that colonize the skin and nose of both hospital staff and
patients) cause a wide variety of lung, bone, heart and blood- stream
infections and are frequently resistant to antibiotics; beta-hemolytic
streptococci are also important. Gram-negative bacteria:
Enterobacteriacae (e.g. Escherichia coli, Proteus, Klebsiella,
Enterobacter, Serratia marcescens), may colonize sites when the host
defenses are compromised (catheter insertion, bladder catheter,
cannula insertion) and cause serious infections (surgical site, lung,
bacteremia, peritoneum infection)
They may also be highly resistant. Gram negative organisms such as

Pseudomonas spp. are often isolated in water and damp areas. They

may colonize the digestive tract of hospitalized patients. Selected

other bacteria are a unique risk in hospitals. For instance, Legionella

species may cause pneumonia (sporadic or endemic) through

inhalation of aerosols containing contaminated water (air

conditioning, showers, therapeutic aerosols).

Reservoirs and transmission
Bacteria that cause nosocomial infections can be acquired in several ways:

1- The permanent or transient flora of the patient

(Endogenous infection) Bacteria present in the normal flora cause infection
because of transmission to sites outside the natural habitat, damage to
Tissue or in appropriate antibiotic therapy that allows over growth (C. difficile,
yeast spp).For example, Gram-negative bacteria in the digestive tract
frequently cause surgical site infections after abdominal surgery or urinary
tract infection in catheterized patients.

2- Flora from another patient or member of staff

(Exogenous cross-infection) Bacteria are transmitted between patients
through: direct contact between patients (hands, saliva droplets or other body
fluids), in the air (droplets or dust contaminated by a patient’s bacteria),via
staff contaminated through patient care (hands, clothes, nose and throat) who
become transient or permanent carriers, subsequently transmitting bacteria
to other patients by direct contact during care, via objects contaminated by
the patient, the staff’s hands, and visitors or other environmental sources


Quorum sensing
is defined as the way that bacteria use auto-inducer (AI)
molecules for bacterial cell-to-cell communication. AIs include
oligopeptides and N-acyl-homoserine lactones (AHLs) in Gram-
positive and -negative bacteria, respectively.
 Superbug
Non
Biological:
Biological:

MDR
Particulate…..
Dust, dead
skin, hair,
makeup…
Chemical…….
..Oil, grease,
perfume
Bacteria Viruses Fungi
 By 2050 Superbugs could
kill 10 million people a year

More than the number

who currently die from cancer
Based on numerous scientific studies conducted from the mid-20th century, a number of mechanisms
explaining bacterial resistance to antibiotics have been proposed. Bacteria are currently believed to
acquire antibiotic resistance via:
active removal of the antibiotic from the cell,
enzymatic modifications of the antibiotic,
modifications of cell components which are the target of the antibiotic,
overexpression of an enzyme inactivated by the antibiotic,
a change in the permeability of bacteria cell membranes,
production of an alternative metabolic pathway,
an increase in the concentration of a metabolite which is an antagonist of the antibiotic,
a reduction in the amount or activity of an enzyme activating the precursor of the antibiotic,
modifications in regulatory systems not associated with the direct mechanism of action of the antibiotic,
or a reduction in the demand for the product of the inhibited metabolic pathway.
Factors influencing the development of nosocomial infections:

The microbial agent

The patient is exposed to a variety of microorganisms during

hospitalization. Contact between the patient and a microorganism
does not by itself necessarily result in the development of clinical
disease other factors influence the nature and frequency of
nosocomial infections.

Many different bacteria, viruses, fungi and parasites may cause

nosocomial infections. Infections may becaused by a microorganism
acquired from another person in the hospital (cross-infection) or may
becaused by the patient’s own flora (endogenous infection).
Most hospital infections were due to pathogens of external
origin(food borne and airborne diseases, gas gangrene, tetanus,etc.)
or were caused by microorganisms not present in the normal flora of
the patients (e.g. diphtheria, tuberculosis) Progress in the antibiotic
treatment of bacterial infections has considerably reduced mortality
from many infectious diseases. Most infections acquired in hospital
today are caused by microorganisms which are common in the
general population, in whom they cause no or milder disease than
among hospital patients such as Staphylococcus aureus, coagulase
negative staphylococci, enterococci and Enterobacteriaceae
Patient susceptibility

Important patient factors influencing acquisition of infection include age,

immune status, underlying disease, and diagnostic and therapeutic
interventions The extremes of life infancy and old age are associated
with a decreased resistance to infection. Patients with chronic disease
such as malignant tumors, leukemia, diabetes mellitus, renal failure, or
the acquired immunodeficiency syndrome (AIDS) have an increased
susceptibility to infections with opportunistic pathogens

part of the normal bacterial flora in the human, but may become
pathogenic when the body’s immunological defenses are compromised.
Many modern diagnostic and therapeutic procedures, such as biopsies,
endoscopic examinations, catheterization, ventilation and suction and
surgical procedures increase the risk of infection. Contaminated objects
or substances maybe introduced directly into tissues or normally sterile
sitessuch as the urinary tract and the lower respiratory tract
Crowded conditions within the hospital, frequent transfers of patients
from one unit to another, and concentration of patients highly
susceptible to infection in one area (e.g. newborn infants, burn
patients, and intensive care) all contribute to the development of
nosocomial infections. Microbial flora may contaminate objects,
devices, and materials which subsequently contact susceptible body
sites of patients. In addition, new infections associated with bacteria
such as water borne bacteria (atypical mycobacteria) and or viruses
and parasites continue to be identified
Environmental factors

Crowded conditions within the hospital, frequent transfers of patients from one unit
to another, and concentration of patients highly susceptible to infection in one area
(e.g. newborn infants, burn patients, and intensive care) all contribute to the
development of nosocomial infections. Microbial flora may contaminate objects,
devices, and materials which subsequently contact susceptible body sites of
patients. In addition, new infections associated with bacteria such as water borne
bacteria (atypical mycobacteria) and/orviruses and parasites continue to be
identified
Bacterial resistance

microorganisms in the normal human flora sensitive to the given drug are
suppressed, while resistant strains persist and may become endemic in the
hospital. The wide spread use of antimicrobials for therapy or prophylaxis is the
major determinant of resistance. Antimicrobial agents are, in some cases,
becoming less effective because of resistance. As an antimicrobial agent becomes
widely used, a bacterium resistant to this drug eventually emerges and may spread
in the health care setting.
Many strains of pneumococci, staphylococci, enterococci, and
tuberculosis are currently resistant to most or all antimicrobials
which were once effective. Multi resistant Klebsiella and
Pseudomonas aeruginosa are prevalent in many hospitals. This
problem is particularly critical in developing countries where more
expensive second-line antibiotics may not be available or affordable.
Nosocomial infections are wide spread. They are important
contributors to morbidity and mortality. They will become even more
important as a public health problem with increasing economic and
human impact because of: Increasing numbers and crowding of
people, more frequent impaired immunity (age, illness, and
Mechanisms of Acquisition of Drug Resistance among Bacteria

The simplest type of resistance is a natural lack of susceptibility, called innate

resistance. This is a constant trait of a species, strain, or whole group of bacteria. A
given microorganism is insensitive to an antibiotic due to its ‘innate’ resistance to
certain groups of antibiotics. It may be linked to the absence of a receptor for the
antibiotic, low affinity, cell wall impermeability, or enzyme production.
Changes in the susceptibility of bacteria can be primary or secondary.
Primary resistance arises as a result of a spontaneous mutation and
can appear without contact with a drug. This type of resistance is
encoded chromosomally and is not transmitted to other bacterial
species. The frequency of occurrence of mutated bacteria is low but
in the presence of an antibiotic, mutants have an advantage over the
rest of the population, and thus they survive and outnumber
susceptible populations. They can spread to other ecological niches
in the same individual or can be transferred to other microorganisms.
While defending themselves against antibacterial agents, including
antibiotics, in the course of their evolution, bacteria have developed a
variety of mechanisms counteracting the effects of antibacterial
agents. As a result of the acquisition of resistance genes, bacteria
become partly or entirely resistant to a given antibiotic by developing
various effector mechanisms.
Surgical wound infections (surgical site Infections)

Surgical site infection (SSI) is a major public health problem. It is the third most common health care
associated infection and contributes to 13–17% of all such infections. The definition is mainly clinical:
purulent discharge around the wound or the insertion site of the drain, or spreading cellulitis from the
wound. The infection is usually acquired during the operation itself; either exogenously (e.g. from the
air, medical equipment, surgeons and other staff), endogenously from the flora on the skin or in the
operative site or, rarely, from blood used in surgery.

SSIs are associated with considerable morbidity and it has been reported that over one-third of
postoperative deaths are related SSI can double the length of time a patient stays in hospital and
thereby increase the costs of health care
WHAT TO KNOW WHEN CONSIDERING

A CLEANROOM

What’s the application? Better quality or better yield is the primary

reason for investing in a cleanroom space. It goes straight to your
bottom line. Numerous manufacturing procedures now require a
controlled environment in which you limit the amount of dust and dirt
in the area of the manufacturing. Medical instrument manufacturing
and packaging, electronics and computer manufacturing, food
preparation and some military applications are but a few of the
instances that have strict requirements for maintaining a clean
environment. You need to know the requirement for your specific
product or process. If the product you are manufacturing is regulated
by a government agency, or you are contracting with a private firm
that requires a certain level of clean manufacturing, they should have
the cleanroom standards already documented. Check with them first.





CLEAN ROOM STANDARDS

EIC
Systems
 USA. FEDERAL STANDARD 209E*

Defines a clean room

as a room in which the

concentration of
AIRBORNE PARTICLES
is controlled to specified limits.

*Is from the U.S. General Service Administration and approved for use by all U.S. agencies. In the absence of
an international standard, FS 209E was broadly used internationally.
 Clean Room applications

• “Federal Standard 209E”

defines a clean room as a
room in which the
concentration of AIRBORNE
PARTICLES is controlled to
specified limits.
 BRITISH STANDARD 5295

Defines a clean room

as a room with control of PARTICULATE CONTAMINATION,
constructed and used in such a way as to minimize the
introduction, generation and retention of particles inside the
room and in which the temperature, humidity,
airflow patterns, air motion and pressure are controlled.
 ISO 14644 (1993)

Room in which the concentration of AIR BORNE

PARTICLES is controlled,
Constructed and used in a manner to minimize the
INTRODUCTION, GENERATION AND RETENTION of
particles inside the room,
And in which other relevant parameters e.g.
temperature, humidity and pressure are controlled as
necessary
ISO 14644

Numbers &
Sizes
There are different levels of cleanrooms. ISO—the
International Standards Organization, ranks cleanrooms ISO
Class 1 (the cleanest) through ISO Class 9. The lower the ISO
rating, the cleaner the environment. Measurement of
contamination is done in “parts-per-cubic-meter.” An ISO
Class 6 cleanroom, for example, is rated at 35,200 parts per
cubic meter. That means the room can have no more than
35,200 particles greater than 0.5 micron in size per cubic
meter. These are particles that are not visible to the human
eye. (As a comparison, a particle of cigarette smoke is
between 0.5 and 2 micron in size. The end of a piece of
human hair is about 60 to 100 microns in size).
ISO Standard for
Classification Of Clean Room

49
Sizes Of Airborne Particles
Human Hair: 50-150 µm
Dust : 0.01-100 µm
Pet dander: 0.1-10 µm
Pollen: 10-110 µm
Tobacco Smoke: 0.01-1 µm
fungal spores: 2.0-5.0 µm
Bacterial cells: 0.3–10 µm

Viruses: 0.02-0.30 µm 1.HEPA Company glossary of terms

2.Originally 'High Efficiency Particulate Arrestment - see thefreedictionary.com
3.American Society of Mechanical Engineers, ASME AG-1a–2004, "Addenda to ASME AG-1–2003 Code on Nuclear Air and Gas Treatment", 2004
4..: a b Gantz, Carroll (2012-09-21). The Vacuum Cleaner: A History. McFarland. p. 128. ISBN 9780786493210.
 Types of Airborne Particles

Non Biological: Biological:

Particulate…..Dus
Chemical……...Oil,
t, dead skin, hair, Bacteria Viruses Fungi
grease, perfume
makeup…
 SIZES AND TYPE OF AIRBORNE
PARTICLES
Human Hair: 50-150 µm

Dust : 0.01-100 µm
Skin cells 4-44 μm
Tobacco Smoke: 0.01-1 µm
Fungal spores: 2.0-5.0 µm

Bacterial cells: 0.3–10 µm

Viruses: 0.02 - 0.30 µm 1.HEPA Company glossary of terms

2.Originally 'High Efficiency Particulate Arrestment - see thefreedictionary.com
3.American Society of Mechanical Engineers, ASME AG-1a–2004, "Addenda to ASME AG-1–2003 Code on Nuclear Air and Gas Treatment", 2004
4..: a b Gantz, Carroll (2012-09-21). The Vacuum Cleaner: A History. McFarland. p. 128. ISBN 9780786493210.
WHO GUIDELINES FOR CLASSIFICATION OF CLEAN ROOM
THANK YOU