‫&‬

‫وحدة السالمة ومكافحة العدوى‬

 Medical Microbiology Laboratory

▪ Plays An Integral Role In The Practice Of Infection

Control Because It Defines One Of The Major
Components Of The Disease Process (The Agent )
▪ Determine the classification of bacteria and how
merging microbiology in stewardship

▪ Define the antibiogram to guide empiric antibiotic

selection and to detect bacterial resistance patterns

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 WHAT IS ANTIBIOGRAM?

• An antibiogram Is An Overall Profile Of Antimicrobial

susceptibility Testing Results Of A Specific Microorganism
To A Battery Of Antimicrobial Drugs ……Only Results For
Antimicrobial Drugs That Are routinely Tested And
Clinically Useful Should Be Presented To Clinician

• Used For Clinical Decision Making

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COLLECTION OF SPECIMENS

Many Mistakes Can Alter The Results Including :

➢Unsuitable Specimen
➢Inadequate Specimen
➢Wrong Timing
➢Wrong container
➢Introduction Of Contaminants

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 Specimen Collection Guidelines

• The proper collection of a specimen for culture is the most

important step in the recovery of pathogenic organisms responsible
for infectious disease.
• A poorly collected specimen may lead to failure in isolating the
causative organism(s) and/or result in the recovery of
contaminating organisms.
 Basic Concepts for Specimen Collection
• Collect the specimen from the actual site of infection, avoiding contamination
from adjacent tissues or secretions
Ensure adherence to safety precautions( gloves, laboratory coat, and face
wear)
• Collect the specimen at optimal times (for example, early morning sputum for
AFB culture).
• Collect a sufficient quantity of material. Use appropriate collection devices:
sterile, leak-proof specimen containers. Use appropriate transport media
Check expiration date before inoculating collection device.
• Whenever possible, collect specimens prior to administration of antibiotic”the
flora changes and leading to potentially misleading culture result
Properly label the specimen and complete the test request form. Hospital
identification number
➢ Collection date and time
➢ Ordering clinician ( Age - Gender )
➢ Exact nature and source of the specimen
➢ Provisional diagnosis
➢ Current antimicrobial history
• Minimize transport time. Maintain an appropriate environment between
collection of specimens and delivery to the laboratory.
• If appropriate, decontaminate the skin surface. Use 70-95% alcohol (ALC) and 2%
chlorhexidine or 1-2% tincture of iodine (TIO) to prepare the site. Allow a contact
time of two minutes to maximize the antiseptic effect.
 TRANSPORT OF SPECIMENS

Delay In Transport Can Lead To :

• Death Of Delicate Organisms

• Overgrowth Of Pathogens By Fast Growing Commensals
• Variation In Bacterial Count
• To Avoided
• Do Culture At Bed
• Use Transport Media
• Adjust Temperature Condition
 Definitions

• MDR: Multidrug Resistant

The isolate is non-susceptible to at least 1 agent in ≥ 3 antimicrobial categories
listed in antimicrobial families

• XDR: Extensively Drug Resistant

The isolate is non-susceptible to at least 1 agent in all but 2 or fewer
antimicrobial categories

• PDR: Pan Drug Resistant

Non-susceptibility to all agents in all antimicrobial categories for each
bacterium

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 • Clinical importance of MDROs

• Increased use of broad-spectrum agents may result increased incidence of

MDROs
• Unavailability of antibiotics to treat MDROs in the near future due to lack of
development of newer antimicrobial agents.
• Options for treating patients with these infections are often extremely limited
• Infections by MDROs have been associated with:
• increased lengths of stay,
• Additional costs,
• and morbidity and mortality

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 Epidemiology of MDROs

Factors Affecting Prevalence :

• Host
• Place
• Type And Level Of Care
• MDROs burden is greatest in adult
hospital patients, in ICUs , in tertiary hospital

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Transmission of MDROs:
Transmission and Persistence Of MDRO is determined by:
• The availability of vulnerable patients with :

• severe diseases
• compromised host defenses
• recent surgery
• indwelling medical devices

• Selective pressure exerted by antimicrobial use

• colonization pressure
• The impact of implementation and adherence to prevention efforts
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Main Sources:
• Contaminated hands of healthcare personnel (HCP)
• Contaminated environmental surfaces in close
proximity to the patient

Role of colonized HCP in MDRO transmission: HCP can become

persistently colonized with an MDRO, but these HCP have a limited role in
transmission, unless other factors are present to facilitate transmission, e.g.
chronic sinusitis, upper respiratory infection, and dermatitis.

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 Types of resistance

1. Intrinsic Resistance (natural )

micro organismes naturally do not posses target sites for drugs and ther
fore the drug does not affect them or they naturally have low
permeability to htose agents because of the differences in the chemical
nature of the drug and the microbial membrane structuras especially for
those that requier entry in to the microbial cell in order to affect their
action
Exemple :
resistance of klebsiella SPP to ampicillin

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 2. Acquired Resistance
Acquired Resistance is when a naturally susceptible microorganism acquires ways of not being
affected by the drug
▪ vertical transmission (Mutation )
Changes in the composition or structure of the target in the bacterium (resulting from
mutations in the bacterial DNA ) can stop the antibiotic from interacting with the target
▪ Horizontal gene transfer
➢ transformation = ability of microorganisms to utilize free DNA from their
surroundings
➢conjugation = contact between bacteria
➢transduction = transfer genetic material from one organism to another by a phage

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How does antimicrobial resistance occur?
• Person-person
• Animals-humans
• Food
• Water
• Within healthcare
facilities
• Travelling
• Trade

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Use of antibiotics for animals

Over 70% of all medically important

antibiotics in the United States are sold
for use in animals.

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 WHAT ARE THE MOST IMPORTANT RESISTANCE PATTERNS?

❑ Methicillin Resistant Staphylococcus Aureus (MRSA)

❑ Vancomycin Resistant Staphylococcus Aureus (VRSA)
❑ Vancomycin Resistant Enterococcus (VRE)
❑ Multidrug-resistant Gram- negative bacteria (Acinetobacter spp klebsilla
spp.Psuedomonas spp.)
Carbapenems Producing Organisms (CPO)
➢ Staphylococcus aureus

➢ is very common, but also very deadly if it gets into the blood stream.

➢ Staphylococcal Enterotoxins, Toxic Shock Syndrome Toxin Tssts

And Exfoliative Toxin Are ‘Superantigens’ , Resulting In Over
Production Of Cytokines, Giving Rise To A Toxic Shock-like
Presentation. It Was Formerly A Major Cause Of Death Following
Surgery.

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Methicillin-resistant S. aureus (MRSA)
• MRSA is defined as S. aureus strains that are resistant to methicillin,
oxacillin, penicillin and amoxicillin. They can also be resistant to all ß-
lactam agents, including cephalosporins and carbapenems.
 Community- associated MRSA
• In recent years, community- associated MRSA (CA- MRSA) has
emerged as a major pathogen.
• Infection with CA- MRSA presents most commonly as relatively minor
skin and soft tissue infections, but severe disease, including
necrotizing pneumonia, necrotizing fasciitis, osteomyelitis, and sepsis
syndrome have also been reported both in children and adults.
• Community strains of Staph. aureus (both MRSA and MSSA) also
produce several toxins which are not commonly found in hospital
strains notably Panton-Valentine leukocidin (PVL), which causes
leukocyte destruction and tissue necrosis.
Screening swabs
• The value of ‘universal’ MRSA screening in the prevention and control of
MRSA infections remains controversial as its cost- effectiveness has not
been fully established. Currently, screening of MRSA is recommended in
the following ‘high- risk group’ of patients.
1. Preoperative patients in certain surgical procedure, e.g. elective
orthopedic, cardiothoracic, vascular, and neurosurgery.
2. Emergency orthopedic and trauma patients.
3. Critical care units, i.e. intensive care, neonatal, burn, renal, transplant,
and oncology units.
4. All previously known MRSA positive patients.
 Screening swabs

• A swab moistened with sterile water should be used to sample carrier

sites and lesions.

• The screening swabs should be taken from the nose, perineum/

groin, operative and wound sites, abnormal or damaged skin, invasive
devices, catheter urine samples, and sputum
• Detection of MDROs : through either phenotypic methods or
• Genotypic methods (detection of resistant genes by molecular techniques e.g
PCR) •
MRSA:
• Disk diffusion
• Oxacillin agar screen
• MIC Broth dilution
• E test
• Automated systems
• Chromogenic agar for MRSA screening
• • Detection of mecA gene by PCR30
Vancomycin- resistant Enterococci
• Enterococci are anaerobic Gram- positive COCCI.

• They are commensal bacteria with low virulence. They are present mainly in the
gastrointestinal tract, but also found in the anterior urethra, vagina, skin, and
oropharynx of healthy individuals.

• The first clinical strains of vancomycin resistant enterococci (VRE) were reported
in 1988 (Enterococcus faecium and Enterococcus faecalis)
Risk factors for VRE colonization and infection include:
• Prior use of multiple courses of antibiotics,
• Colonization pressure, i.e. the proportion of VRE- positive patient days in
a unit
• Exposure to other colonized or infected patients, contaminated items,
equipment, and environmental surfaces with VRE,
• Presence of an invasive device
• Chronic skin ulcers
• Severity of illness,
• Admission to the intensive care unit
• Prolonged stay in healthcare facility
Multidrug-resistant Gram- negative bacteria
• These organisms can develop resistance during treatment against 3rd
generation cephalosporins (cefotaxime, cefriaxone, cefazidime, etc.)
• The most common MDRGN bacteria which are extensively resistant
to antibiotic belong to the Enterobacteriaceae family, especially
Klebsiella spp. and Escherichia coli, and ‘SCEMP’ microorganisms,
i.e. including Serratia spp., Citrobacter freundii, Enterobacter spp.,
Morganella morganii, and Providencia spp.
• Non- fermenters multi- resistant Gram- negative bacteria which
especially pose problems include Pseudomonas aeruginosa,
Acinetobacter baumannii, and Stenotrophomonas maltophia.
 Non- fermentative Gram- negative bacteria

• Acinetobacter spp. Gram- negative coccobacilli.

There are over 30 species and the organism is found in
soil, sewage, and water. Various species colonize the
human skin and they may be readily recovered from the
forearm, forehead, and toe webs of healthy volunteers.
 Klebsiella pneumoniae Carbapenemases
• Klebsiella pneumoniae carbapenemase (KPC)-producing bacteria are a group
of emerging highly drug-resistant Gram-negative bacilli causing infections
associated with significant morbidity and mortality.
• High motrtality 38% and 57% in Isreal and US (Carmeli et al, 2010)
• Infection control emergency!!!
• May test sensitive to carbapenems though!
• Extensive multidrug resistance (XDR)
• Very rapid spread
• Empiric therapy: colistin + tigecycline
 Pseudomonas aeruginosa
• Is a true opportunistic pathogen and is responsible for causing a variety
of infections in clinical settings in both immunocompetent, e.g.
folliculitis resulting from bathing in hot tubs (‘Jacuzzi’) and
immunocompromised hosts.
• It causes malignant external otitis, and it colonizes and infects the lower
respiratory tract in patients with cystic fibrosis. In hospitalized patients,
it particularly infects those who are intubated and undergoing
mechanical ventilation and it also figures prominently in infections of
indwelling devices, such as urinary catheters. In addition, it has also
been responsible for outbreaks in neonatal units
What is the threat of antimicrobial resistance?
❖Every year, over 2 million people in the United States become infected with
bacteria that are resistant to antibiotics, and around 23,000 people die as a
result of these infections (CDC, 2016).
Multidrug-resistant organisms, are bacteria that are resistant to current
antibiotic therapy and, therefore, difficult to treat.

❖Can MDROS cause serious and local systemic that infections be can
severely debilitating even and life-threatening.

❖In the past, these infections were usually controlled by penicillin

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Antimicrobial Resistance: Key Prevention Strategies
Susceptible Pathogen
Pathogen
Antimicrobial-Resistant Pathogen
Prevent
Prevent Infection
Transmission
Infection
Antimicrobial
Resistance
Effective
Optimize Diagnosis
Use and Treatment
Antimicrobial Use
STRATEGIES OF PREPENTION AND CONTROL OF MDROS

Infection prevention

4 parallel strategies Accurate and promote diagnostic

and treatments

Prudent use of antimicrobials

Prevention of transmission
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1. Prevention of Infections:
✓Vaccinate
✓Optimal management of VAP of CVL – BSIs , urinary and
vascular catheters infections by compliance of bundled
evidence based clinical practices have been recommended (CDC)
2. Effective Diagnosis and Treatment

Target the pathogen by :

➢ Do cultures and use the antibiogram

➢Target empiric therapy to likely
pathogens and local antibiogram
➢Target definitive therapy to known
pathogens and antimicrobial susceptibility
test results
➢Allows to select the most narrow
antibiotics available and reduces
resistance.

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 3. Optimize Use of Antimicrobial

When we have to prescribe antibiotics, we use the 4 D’s

• Drug The correct drug for the type of bacteria you want to treat

• Dose Depends on the type of bacteria and the infection it is

causing (e.g. dose to treat staphylococcus causing osteomyelitis, is different from the
dose used to treat a skin infection).

• Duration You want to clear the infection, prolonging the course

exposes patient to unnecessary side-effects and wastes
resource, but a too short course allows for resistance to develop.

• De-escalation Unnecessary IV antibiotic use is associated with increased

risk of line infection, hospital stay and hospital acquired infection
 4. Prevent Transmission

Control Interventions
The various types of interventions used to control or eradicate MDROs
are grouped into seven categories. These include:

1. Administrative support
2. Education
3. Judicious use of antimicrobials
4. Surveillance (routine and enhanced)
5. Standard and Contact Precautions
6. Environmental measures
7. Decolonization
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1) Administrative support
• Providing the supplies for hand hygien
• Enforcing adherence to recommended infections
control practices (hand hygiene ,standard and
contact precautions
 2) Education

1. Encourage a behavior change through improved understanding of

the problem MDRO that the facility was trying to control
2. Enhance adherence to hand hygiene practices
3. Improving prescribing patterns

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 3) Judicious use of antimicrobial agents :
▪ Effective antimicrobial treatment of infections
▪ Use of narrow spectrum agents
▪ Treatment of infections and not contaminants
▪ Treatment of infection and not colonization
▪ Avoiding excessive duration of therapy
▪ Restricting use of broad spectrum or more potent antimicrobials
to treatment of serious infections when the pathogen is not known
or when other effective agents are not available
▪ Antimicrobial cycling
4) Surveillance of MDROs: Main Target:

• detection of newly emerging pathogens

• monitoring epidemiologic trends
• measuring the effectiveness of interventions

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 5) Standard and Contact Precautions

 They are the foundation of all precautions to prevent transmission of

infections associated with healthcare.

 They are applied to all patients receiving care in hospitals , regardless

of their diagnosis or infection status.

 They are designed to reduce the risk of transmission of microorganisms

from both recognized and unrecognized sources of infection in hospitals
by isolate the pathogen and break the chain of infection

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 component of
standard precautions New elements

 Hand hygiene  Safe injection practices

 Personal protective equipment  Infection control practices for special
 Soiled patient care equipment lumber puncture procedures .
 Environmental control  Respiratory hygiene / cough etiquette

 Textiles and laundry

 Needles and other sharps
 Patient resuscitation
 Patient placement
 Contact precautions

 Apply to specified patients known or suspected to be infected or

colonized with epidemiologically important microorganisms that can
be transmitted by direct or indirect contact .

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How Transmission Occurs
 Transfer from contaminated environment/ reusable medical
equipment (RME) to a compromised patient

 By healthcare worker’s hands

 By direct contact with the organism (in an open wound)

52 Dr. Rami H. Alabadla

In addition to standard precautions

 Patient placement
 PPE and hand hygiene
 Patient transport
 Patient – care equipment
 Environmental measures .

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6) Environmental Measures

• Use of dedicated noncritical medical equipment

• Assignment of dedicated cleaning personnel to the affected patient
care unit
• Enhanced environmental cleaning
• Increased cleaning and disinfection of frequently-touched surfaces

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 Infection vs Contamination vs Colonization

Bacteria in Sample Presence of

tissues contains bacteria at a
associated with bacteria from site without
acute an external related signs
inflammation source, e.g. and
and a defined mistakes in symptoms
clinical sampling
syndrome technique
• Decolonization:
❖When decolonization for MRSA is used, perform susceptibility testing for the
decolonizing agent against the target organism or the MDRO strain epidemiologically
implicated in transmission.

❖Decolonization means treatment of persons colonized with a specific MDRO usually

MRSA to eradicate carriage of that organism

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Decolonization therapy for MRSA
• Duration of treatment: Treatment should be prescribed by a medical
practitioner and should be continued for 5 days.
• Clothing and bed linen: For effective decolonization, it essential that
towels, bed linen, and patients’ clothing must be washed and
changed daily until the end of the treatment.
• Nose colonization: For nose colonization, apply 2% nasal mupirocin
ointment three times a day for 5 days.
• Body bathing: During the shower, the antiseptic body wash should be
applied directly to the skin