Diagnostic microbiology
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Transcript of Diagnostic microbiology
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11/04/2023DR.T.V.RAO MD 1
Dr.T.V.Rao MD
BASIC ORGANIZATION DIAGNOSTIC MICROBIOLOGY
LABORATORY
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11/04/2023DR.T.V.RAO MD 2
• The work environment has changed with the development of new technology. Laboratories have always seen the need for change and development, there has been increased pressure to improve performance, tighten margins, improve quality and reduce costs
Challenges in Diagnostic Laboratories
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11/04/2023DR.T.V.RAO MD 3
Medical Laboratory Managers
• Each laboratory must have a strategic plan that describes its long-term goals, such as a move toward more automation or molecular diagnostic techniques.
• Each employee’s role should be clearly defined, and written job descriptions should be provided so personnel know what they are expected to do. Therefore, it is a not an easy task for a manger to strike a balance among the clinical laboratory regulations, fiscal responsibility, and employee competence and morale to maintain the overall quality of patient care.
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11/04/2023DR.T.V.RAO MD 4
• It is appropriate to remember that the two most important components of management are
• Common sense
• Open communication with laboratory staff
THE LABORATORIES SHOULD FACTION FROM SENSE TO COMMON SENSE
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11/04/2023DR.T.V.RAO MD 5
MICROBIOLOGY LABORATORY
Clinical Microbiology comprises essentially seven sections.• Aerobic and anaerobic bacteriology• Mycology• Mycobacteriology (also called Acid-fast Bacteriology, AFB)
• Parasitology • Virology• Serology• Molecular diagnostics (PCR & DNA probe technology)• :
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11/04/2023DR.T.V.RAO MD 6
Sample Receiving & Processing SectionSamples brought to the clinical microbiology are at first received by this section. Here sample are received and the samples are processed according to the nature of the sample.
• Urinalysis SectionIn this section detailed report of urine samples including physical, chemical, microscopic examination is be prepared.
PLAN FOR THE DIVISION OF MICROBIOLOGY
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11/04/2023DR.T.V.RAO MD 7
• Parasitology SectionParasitology section deals with intestinal parasites. Samples of faeces are examined here for the presence of any intestinal parasite. Slides are prepared here inside a safety cabinet.
DIVISION OF PARASITOLOGY
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11/04/2023DR.T.V.RAO MD 8
• Nose, Throat, Sputum and Urogenital Cultures and Sensitivity SectionHere cultures of respiratory tract and genital tract infections are prepared.
• Blood cultures, Wounds Culture and Sensitivity SectionCulture of wound swab, pus, aspirates, body fluids including CSF are the responsibility of this section.
BACTERIOLOGY CULTURES MAIN PART OF LABORATORY SERVICES
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11/04/2023DR.T.V.RAO MD 9
• Urine Culture & Sensitivity SectionDifferent types of urine culture performed here including mid stream urine and catheter samples of urine. Each sample is processed and evaluated accordingly.
• Blood Culture and Sensitivity SectionIn this section culturing of blood samples is carried out. Nowadays, this section is equipped by machines such as Bactec 9240, flourometric instruments. Each instrument is capable of running 240 samples at a time.
BACTERIOLOGICAL CULTURES IS THE MAJOR GRATIFYING WORK
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11/04/2023DR.T.V.RAO MD 10
• Serology SectionIn serology section immunological and serological tests are performed by different techniques like Latex agglutination, haemagglutination and antibody absorption.
SEROLOGY SECTION
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11/04/2023DR.T.V.RAO MD 11
• Mycobacteriology Culture & Sensitivity SectionIn this section all TB smear, culture and sensitivity performed in two Biosafety II and III cabinets to avoid risk of infection
MYCOBACTERIOLOGY
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11/04/2023DR.T.V.RAO MD 12
• Mycology Culture SectionRequests for fungus smear and culture processed here in a bio safety II cabinet to avoid infection from fungal spore.
• Regardless of the organisation of a Microbiology Laboratory, the main aim is providing the client (the physician) with accurate and reliable results to assist the process of clinical treatment.
MYCOLOGY LABORATORY SERVICES AN EMERGING NEED
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11/04/2023DR.T.V.RAO MD 13
• Quality Control SectionIn this section quality control of water, food products and environment is performed with the help of different media and colony counters.
QUALITY CONTROL IN MICROBIOLOGY LABORATORY
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MANAGING THE SAMPLE: TECHNICAL• Macroscopic evaluation• Split sample for different laboratory disciplines• Two possible approaches:• perform only those tests requested by sender• perform diagnostics for syndromes/clinical description
(laboratory initiative) • Storage of samples
• refrigerator or freezer
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11/04/2023DR.T.V.RAO MD 15
DIRECT METHODS1. Macroscopic evaluation
2. Direct microscopy
3. Electron microscopy
4. Staining
5. Rapid tests
6. Molecular methods
7. Propagate the agent
No propagation required
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• Consistency• rice water stools
for Cholera• Blood• Visible parasites• Helminths• segments
MACROSCOPIC EVALUATIONAN EXAMPLE
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DIRECT MICROSCOPY
Wet mount technique
• hanging drop
Dark background microscope
• fragile organisms (e.g. spirochetes)
Viability maintained
• mobility may be observed
Observations
• white blood cells (denotes invasion)
• red blood cells
• parasites
• protozoa
• Helminths
• eggs
• moving bacteria
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11/04/2023DR.T.V.RAO MD 18
STAINING• A specific staining
• Gram staining
• Specific staining with chemicals
• Ziehl Neelsen staining (Mycobacteria)
• Modified Ziehl Neelsen staining (Cryptosporidium)
• Specific staining with labelled antibodies
• Immunofluorescence - used when gram stain cannot help in diagnosis (e.g. Legionella too small to be visible in a Gram stain)
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11/04/2023DR.T.V.RAO MD 19
RAPID TESTS• Goals
• bacterial, viral or parasite antigen (surface antigen, soluble antigen)
• toxin in biological fluids (e.g. cerebrospinal fluid, blood, urine)
• Main techniques
• direct agglutination: slides, cards
• latex agglutination: slides, cards
• immuno-chromatography: dipsticks
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11/04/2023DR.T.V.RAO MD 20
. DIRECT METHODS - PROPAGATION REQUIRED
• Bacteriology and mycology
• most agents can be propagated on culture media
• Virology• most agents can be propagated on cells
• Parasitology• monocellular organisms can be propagated in
culture media
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11/04/2023DR.T.V.RAO MD 21
PERSONNEL
• The following directorial functions are :1- interpretation , correlation , and communication of lab data
2- interaction with physicians and/or medical staff , patient , administration .
3- monitoring of standard of performance , QC , QI.
4- provision of education programs , planning , research.
5- ensuring sufficient personnel with adequate documented
training and experience to meet the needs of the lab .
6- he/she must be decision-maker in the selection of all lab equipment's and supplies .
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PROPAGATION: ADVANTAGES/DISADVANTAGES
• Advantages• allows anti-microbial susceptibility testing
• allows typing of the micro-organism
• allows storage of the strain
• Disadvantages• depends upon the viability/condition of the agent
• takes time
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11/04/2023DR.T.V.RAO MD 23
DETECTING ANTIBODIES• Precipitation
• Agglutination
• Haemagglutination and haemagglutination inhibition
• Viral neutralization test
• Radio-immunoassays
• ELISA
• Immunofluorescence
• Immmunoblotting
• Immunochromatographic
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ANTIBODY DETECTION: ADVANTAGES/DISADVANTAGES
• Advantages • inexpensive• easy to perform• allows identification of• IgM (acute infection)• IgG (past infection)
• Disadvantages• delayed response
(false negative results during sero-conversion window) • time of infection not always clear
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11/04/2023DR.T.V.RAO MD 25
STANDARDS AND CHECKLISTS
• Purposes:• Standards are the broad principles the
laboratory must meet in order to achieve accreditation• Checklists provide detailed requirements
that inspectors use to determine whether laboratories meet the Standards
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LABORATORY INSPECTIONS:MAINTAINING BALANCE
Quality Improvement
Education
Regulatory Compliance
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THE STANDARDS FOR LABORATORY ACCREDITATION
• Standard I Director / Head of the Divison
• Standard II Physical Facilities & Safety
• Standard III Quality Control and Performance Improvement
• Standard IV Inspection Requirements
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CHECKLISTS• Guide the inspection by assisting with the
interpretation of desired Standards• Provide guidelines for development of
laboratory policies, procedures and processes
• Help ensure accurate, reliable test results• Ensure a focus on patient safety
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• The QC / QI program should be clearly defined and well-organized.
• The QI program must provide the system design and evaluation of proper patient identification and preparation ; specimen collection ; preservation ; transportation ; storage before testing ; processing ; and accurate results reporting.
• This system must ensure optimum patient specimen and integrity of the result throughout the pre-analytical , analytical , and post-analytical process.
Quality control and Quality improvement
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11/04/2023DR.T.V.RAO MD 30
Judgment of the acceptability of QC data must be made at least monthly by the lab director .
Because of many variables , the CAP makes no specific recommendations on the frequency of any additional assessment / review of QC data.
There must be evidence of active review of records of instrument function , temp , and maintenance , for all routine procedures on all shifts.
QI / QA
“SUPERVISION
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• The lab must have documented system in operation to detect and correct significant clerical and analytical errors.
• One common method is review of results by a qualified person before release from the lab , but there is no requirement for supervisory review of all reported data.
• The selective use of delta checks also may be useful in detecting clerical errors in consecutive samples from the same patient.
• In computerized lab , there should be automatic alarm for improbable results.
QI / QA“SUPERVISION”
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11/04/2023DR.T.V.RAO MD 32
EFFECTS OF PRE-ANALYTICAL VARIABLES ON THE QUALITY OF LABORATORY TESTING
• Modern laboratories around the world are now enjoying the benefit of decades of development in technology
• “State of the Art" instrumentation are common in most laboratories
• Walkway, high throughput analyzers are employed for routine and specialized testing
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11/04/2023DR.T.V.RAO MD 33
• Programme Created by Dr.T.V.Rao MD for Medical Microbiologists