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LAB 7: RESPIRATORY SYSTEM
RESPIRATORY SYSTEM: UPPER AND LOWER
THE RESPIRATORY SYSTEM Host Defense Mechanisms
Anatomical characteristics/mechanisms to prevent/fight infection:– nasal hairs – antibacterial enzymes (lysozyme)– convoluted passages – cilia and mucous lining of the trachea– mucous linings of the nasal turbinates – reflexes --coughing, sneezing, swallowing – Normal flora of nasopharynx and presence of normal flora
NORMAL MICROBIOTA OF THE RESPIRATORY SYSTEM
• UPPER respiratory tract :– Coagulase – negative staphylococci– Neisseria (Gram – negative cocci)– Corynebacterium spp. (Diphtheroids) (Gram – positive rods)– Anaerobic bacteria– Pathogens in low numbers (colonization vs. pathogenic) (S. aureus, Gram-negative rods (Enteric/Pseudomonas spp.)
• LOWER respiratory tract (trachea, bronchi, and bronchioles) is virtually free of microorganisms, mainly because of the efficient cleansing action of the ciliated epithelium
RESPIRATORY TRACT INFECTIONS
Bronchitis Pharyngitis
Epiglottitis Sinusitis
Pneumonia
PATHOGENS OF THE UPPER RESPIRATORY TRACT
THROAT:
•Streptococcus pyrogenes (Group A strep)•Viruses
SINUS:
•Moraxella catarrhalis (Gram – negative cocci)•Gram – negative rods•Staphylococcus aureus•Viruses
PATHOGENS OF THE LOWER RESPIRATORY TRACT
• Streptococcus pneumoniae• Hemophilus influenzae• Staphylococcus aureus• Moraxella catarrhalis (Gram – negative cocci)• Gram – negative rods (Klebsiella sp., Enterobacter sp., Serratia sp., E. coli, Pseudomonas sp.)• Fungi (Cryptococcus neoformans)• Viruses
The Respiratory System: Collecting Specimens & Diagnosis
• Upper respiratory tract: Mucus membrane (throat, sinus) is wiped with a special collection swab. – Culture for isolation, identification
and susceptibility testing of pathogens
• Lower respiratory tract: Sputum and or bronchial specimens are collected into a sterile container.
– Gram stain for white blood cells and microorganism and to check for the quality of the specimen
– Culture for isolation, identification and
susceptibility testing of pathogens.
MEDIASheep Blood agar:
•Enriched (contains 5% sheep blood with tryptic soy agar base)•Differential (used to detect hemolysis of microorganisms)
•Microorganisms produce toxins called hemolysins that can break down red blood cells causing a visual change in the media
IDENTIFICATION
•DIFFERENTIATION OF STAPHYLOCOCCI AND STREPTOCOCCI
– GRAM STAIN– CATALASE– HEMOLYSIS ON SHEEP BLOOD AGAR
GRAM STAIN
•STAPHYLOCOCCUS
•STREPTOCOCCUS
GRAM POSITIVE COCCI IN PAIRS ANDCLUSTERS
GRAM POSITIVE COCCI IN PAIRS ANDCHAINS
CATALASE TEST
Streptococci are Gram-positive, catalase negative (no bubbles)
Staphylococci are Gram-positive, catalase positive (bubbles)
Blood Agar - Hemolysins• Alpha-hemolysis – causes partial lysis of RBCs and
breakdown of hemoglobin which leads to a green color change in the agar.
• Beta-hemolysis – causes complete lysis of RBCs and breakdown of hemoglobin which leads to a colorless clearing in the agar.
• Gamma-hemolysis – an organism that does not cause hemolysis.
SHEEP BLOOD AGAR
BLOOD AGAR LAB PROCEDURE
BLOOD AGAR LAB PROCEDURE
Blood Agar Lab Procedure: RESULTS
GROUP A STREP (S. pyrogenes) – (causative agent of ‘Strep throat’)
Bacitracin (A) disk test
Group A beta strep is sensitive to bacitracin (zone of inhibition)
Other beta streps are resistant to bacitracin (no zone of inhibition)
Streptococcus pneumoniae• S. pneumoniae is an alpha
streptococci that is susceptible to the optochin disk (zone of inhibition)
• Most common cause of CAP
• Other alpha streptococci are resistant to optochin (no zone of inhibition)
Streptococcus pneumoniae• S. pneumoniae • Catalase negative• Gram – positive cocci in LANCET PAIRS• Frequently encapsulated
DICHOTOMOUS KEY FOR IDENTIFICATION OF STAPHYLOCOCCI AND STREPTOCOCCI
GRAM POSITIVECOCCI IN PAIRS, CHAINS OR CLUSTERS
CATALASE TEST CATALASE NEGATIVE = STREPTOCOCCI
CATALASE POSITIVE=STAPHYLOCOCCI
GAMMA HEMOLYSIS
HEMOLYSIS
BETA HEMOLYSIS – Bacitracin disk sensitive = S. pyrogenes (Beta hemolytic strep Group A)
ALPHA HEMOLYSIS – Optochin disk sensitive = S. pneumoniae
STAPHYLOCOCCI VS. STREPTOCOCCI
GRAM POSITIVE COCCIGRAM POSITIVE COCCIPAIRS AND CHAINS
PAIRS AND CHAINS
PAIRS AND CLUSTERSPAIRS AND CLUSTERS
CATALASECATALASENEGATIVENEGATIVE POSITIVEPOSITIVE
HEMOLYSIS
ALPHABETA
GAMMA
HEMOLYSIS
ALPHABETA
GAMMA
MSA PLATEMSA PLATEPOSITIVEPOSITIVE
NEGATIVENEGATIVE
Coagulase negative Staphylococci
Coagulase negative Staphylococci
S. aureusS. aureus