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STAPHYLOCOCCUS

STAPHYLOCOCCUS. Part 2 Dr. Salma. LAB DIAGNOSIS: Specimens collected : Depends on the type of infection. Suppurative lesion- Pus, Respiratory infection- Sputum, Bacteremia & septicemia- Blood, Food poisoning- Feces, vomit & the remains of suspected food,

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STAPHYLOCOCCUS

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  1. STAPHYLOCOCCUS Part 2 Dr. Salma

  2. LAB DIAGNOSIS: Specimens collected: Depends on the type of infection. Suppurative lesion- Pus, Respiratory infection- Sputum, Bacteremia & septicemia- Blood, Food poisoning- Feces, vomit & the remains of suspected food, For the detection of carriers- Nasal swab.

  3. Methods of examination: I) Direct microscopy: Direct microscopy with Gram stained smear is useful in case of pus, where cocci in clusters are seen.

  4. II) Culture: a) Media used: b) Cultural Characteristics: c) Gram staining: Smears are examined from the culture plate and reveals Gram positive cocci(1μm in diameter) arranged in grape like clusters.

  5. Catalase test; Although microscopic examinations of stained smears presumptively permit distinction between Staphylococci and streptococci, a definitive classification can be made on the basis of the presence or absence of the enzyme catalase. Staphylococci contain this enzyme, streptococci do not.

  6. Hemolysis on blood agar Some bacteria produce hemolysins, exotoxins that cause red blood cells (RBC's) to burst open (hemolyse).  When these bacteria are cultured on blood agar, this hemolysis is visible as an area of clearing around the colony (zone of hemolysis).  If the organism produces enzymes that completely lyse the RBC's, this is termed beta hemolysis (β-hemolysis).  Partial destruction of the RBC's produces a greenish color to the zone of hemolysis and is termed alpha hemolysis (α-hemolysis).  Organisms lacking hemolysins cause no change in the color or opacity of the media and are termed gamma hemolytic or none hemolytic. 

  7. Production of staphyloxanthin (pigmentation) Staphyloxanthin is an orange pigment produced by Staphylococcus aureus that contributes to its virulence. It is the main pigment of this pathogen. Milk agar provides a white background to visually observe the colonies of Staphylococcus aureus

  8. Mannitol fermentation on Mannitol Salt agar (MSA) Staphylococci are able to tolerate the high salt concentration found in Mannitol Salt agar and thus grow readily. If mannitol is fermented, the acid produced turns the phenol red pH indicator from red (alkaline) to yellow (acid).

  9. Production of deoxyribonuclease (DNase) on DNase agar DNase agar contains 0.2% DNA. To detect DNase production, the plate is inoculated and incubated. After growth, the plate is flooded with 1N hydrochloric acid (HCl). DNase positive cultures show a distinct clear zone around the streaked area, where the DNA in the agar was broken down by the bacterial DNase. DNase negative cultures appear cloudy around the growth where the acid caused the DNA in the agar to precipitate out of solution.

  10. d) Biochemical reactions: III) Antibiotic sensitivity tests done as a guide to treatment. IV) Bacteriophage typing is done for epidemiological purposes. V) Serological tests are not useful.

  11. Agar plates with a novobiocin (NB) disc Novobiocin is an antibiotic to which Staphylococcus sp. are either resistant or sensitive. The appearance of a zone of inhibition > 16 mm indicates sensitivity. Filter paper discs impregnated with the appropriate chemical are placed on an agar surface. The chemical diffuses through the agar. Organisms that are susceptible to the chemical will not grow on the agar containing the chemical. The size of the zone of growth inhibition determines the organisms' susceptibility to the chemical.

  12. Production of coagulase The staphylococcal enzyme coagulase will cause inoculated citrated rabbit plasma to gel or coagulate. The coagulase converts soluble fibrinogen in the plasma into insoluble fibrin. . In the laboratory, it is used to distinguish between different types of Staphylococcus isolates.

  13. The Staphyloslide Latex Test for cell-bound coagulase (clumping factor) and/or Protein A: The Staphyloslide Latex Test is an agglutination test that detects cell-bound coagulase (clumping factor) and/or Protein A. This test uses blue latex particles coated with human fibrinogen and the human antibody IgG. Mixing of the latex reagent with colonies of the suspectedS. aureus having coagulase and/or Protein A bound to their surface causes agglutination of the latex particles.

  14. Gelatinase test. Nutrient gelatin is a differential medium that tests the ability of an organism to produce an exoenzyme, called gelatinase that hydrolyzes gelatin. The gelatinase test can be used to differentiate between Staphylococcus aureus and Staphylococcus epidermidis. 

  15. TREATMENT: Drug resistance is common. Benzyl penicillin is the most effective antibiotic, if the strain is sensitive. Cloxacillin or Methicillin is used against beta-lactamase producing strains. Methicillin Resistant Staphylococcus aureus (MRSA) strains have become common. Vancomycin is used in treatment of infections with MRSA strains.

  16. EPIDEMIOLOGY: Staphylococci are primary parasites of human beings and animals. Hospital infections caused by staphylococci deserve special attention because of their frequency & they are caused by strains resistant to various antibiotics. Staphylococci are the common cause of postoperative wound infection and other hospital cross infections.

  17. PREVENTION: Isolation & treatment of MRSA patients. Detection of carriers among hospital staff, their isolation & treatment. Avoid indiscriminate usage of antibiotics.

  18. Coagulase Negative Staphylococci( CoNS ): Two species of coagulase negative Staphylococci can cause human infections- 1. Staphylococcus epidermidis 2. Staphylococcus saprophyticus

  19. S. Epidermidis: It is a common cause of stitch abscesses. It has predilection for growth on implanted foreign bodies such as artificial valves, shunts, intravascular catheters and prosthetic appliances leading to bacteraemia. In persons with structural abnormalities of urinary tract, it can cause cystitis. Endocarditis may be caused, particularly in drug addicts.

  20. S.saprophyticus: It causes urinary tract infections, mostly in sexually active young women. The infection is symptomatic and may involve the upper urinary tract also. Men are infected much less often. It is one of the few frequently isolated CoNS that is resistant to Novobiocin.

  21. Novobiocin sensitivity test

  22. Table summarized the point of differences between 3 imp. Staphylococci species

  23. THANK YOU

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