Staphylococcus aureusBasic Identification in Applied Microbiology Exercise Dr.T.V.Rao MD Dr.T.V.Rao MD Travancore Medical College, 1 Kollam Kerala
Patient presented with • A 30 years male patient presented with an abscess left mandibular region since 3 days, with onset of fever. Dr.T.V.Rao MD Travancore Medical College, 2 Kollam Kerala
What are bacteriological agents can produce this pyogenic condition ? Dr.T.V.Rao MD Travancore Medical College, 3 Kollam Kerala
Most important bacteria areMost abscesses are due toinfection byStaphylococcus (as inthe boil abscess), butother bacteria aresometimes associatedStreptococci, Hemophilusspp. E.Coli Klebsiella sppPseudomonas spp andmany others. Dr.T.V.Rao MD Travancore Medical College, 4 Kollam Kerala
How you proceed to identify the etiological agent Dr.T.V.Rao MD Travancore Medical College, 5 Kollam Kerala
How to make a Definitive Diagnosis• To make a definitive diagnosis and to confirm that Staphylococcus is the bacteria causing the infection, Microscopic observation of the given specimen and a culture can be done. Once a bacteria is identified on a culture, the pattern on sensitivities to antibiotics can help to tell whether or not it is actually MRSA, routine staph aureus, or another bacteria. Dr.T.V.Rao MD Travancore Medical College, 6 Kollam Kerala
Gram staining showing Gram positive Cocci in clusters – Probably Staphylococcus spp Dr.T.V.Rao MD Travancore Medical College, 7 Kollam Kerala
What are Staphylococci: Gram positive cocci ( from Greekstaphyle, means bunch of grapes ) that occur singly and in pairs, short chains and irregular grape-like clusters. Dr.T.V.Rao MD Travancore Medical College, 8 Kollam Kerala
Structure and Physiology of Staphylococcus• Gram-positive cocci, nonmotile, facultative anaerobes• Cells occur in grapelike clusters because cells division occurs along different planes and the daughter cells remain attached to one another• Salt-tolerant: allows them to tolerate the salt present on human skin• Tolerant of desiccation: allows survival on environmental surfaces (fomites) Dr.T.V.Rao MD Travancore Medical College, 9 Kollam Kerala
Staphylocoous appear as• Gram stained smear showing Gram positive cocci with cluster formation indicates presence of Staphylococci Dr.T.V.Rao MD Travancore Medical College, 10 Kollam Kerala
Virulence factors of Staphylococcus aureus III. Dr.T.V.Rao MD Travancore Medical College, 11 Kollam Kerala
Staphylococcus aureus grown on Nutrient agar Dr.T.V.Rao MD Travancore Medical College, 12 Kollam Kerala
General Cultural Characteristics• Some Staphylococcus species, most notably S. aureus, produce a hemolysin that completely lyses red blood cells of humans and some other mammals (sheep blood). This is referred to as “beta” hemolysis. 13
What are your observation on Blood Agar Dr.T.V.Rao MD Travancore Medical College, 14 Kollam Kerala
What are the commonly performed in vitro pathogenicity tests? Dr.T.V.Rao MD Travancore Medical College, 15 Kollam Kerala
Staphylococcus aureus: is thestaphylococcus which has the ability to clot plasma, which is coagulase positive. More than 80% ofStaphylococcus aureus strains produce beta-lactamases.
Other S. aureus CharacteristicsMannitol fermentation is another usefulcharacteristic – it is unique to, and consistentamong S. aureus strains. Virtually all strains ofS. aureus ferment mannitol.Bright yellow colonies on a yellow backgroundindicates mannitol fermentation on mannitolsalt agar. 17
Coagulase test for Identification of Staphylococcus Normal coagulation of plasma Thrombin Fibrinogen Fibrin (soluble ) (insoluble) Staphylococcus aureus coagulation of plasma* “Free” staphylocoagulase Fibrinogen + CRF Fibrin*EDTA Rabbit plasma is preferred for the “free” or tube coagulasetest because it contains a large amount of CRF 18
Slide Coagulase Test• The most important distinction among staphylococci is whether or not they produce the enzyme coagulase. S. aureus the most common pathogen among the catalase positive gram positive cocci and is differentiated from other staphylococci by the coagulase test. Here the bacterial cells have been suspended in a drop of rabbit plasma. Coagulase bound to the cell wall acts on fibrinogen and causes the clumping of the bacteria (right). Coagulase is an important virulence factor of S. aureus Dr.T.V.Rao MD Travancore Medical College, 19 Kollam Kerala
Tube Coagulase Test • The tube coagulase test detects both free and cell bound coagulase of S. aureus. Bacteria are incubated in plasma for 2-4 hours and the tubes turned on their sides as illustrated. Free coagulase acts on prothrombin and fibrinogen in plasma and forms a fibrin clot (left). In manylaboratories staphylococci are simply differentiated as coagulase positive or coagulase negative without speciation Dr.T.V.Rao MD Travancore Medical College, 20 Kollam Kerala
Mention commonly used Antibiotic agents for Staphylococcus aureus• Penicillins• However Penicillin developed early resistance• Other Beta lactam antibiotics like Ampicillin• Macrolide group• Vancomycin, Linezolid ( Only in MRSA strains ).• Commonly used oral anti staphylococcal antibiotics include the first-generation cephalosporins like cephalexin and cefadroxil. Dr.T.V.Rao MD Travancore Medical College, 21 Kollam Kerala
Plate 1a: Staphylococcus aureus resistant to penicillin (P0.5) only. Note the annular radius ofthe zone of inhibition of 9.5 mm around the cefoxitin (FOX 10) disc.
What is the importance of Drug resistance in this isolate?• MRSA is an acronym for Methicillin Resistant Staphylococcus aureus, a bacteria that has become resistant to many antibiotics, including methicillin, penicillin, amoxicillin, and cephalosporins. It is routinely pronounced M.R.S.A. Dr.T.V.Rao MD Travancore Medical College, 23 Kollam Kerala
What are the Other Diseasesproduced by Staphylococcus aureus Dr.T.V.Rao MD Travancore Medical College, 24 Kollam Kerala
Other Staphylococcal Diseases–Systemic Diseases • Toxic shock syndrome-TSS toxin is absorbed into the blood and causes shock • Bacteremia-presence of bacteria in the blood • Endocarditis-occurs when bacteria attack the lining of the heart • Pneumonia-inflammation of the lungs in which the alveoli and bronchioles become filled with fluid • Osteomyelitis-inflammation of the bone marrow and the surrounding bone Dr.T.V.Rao MD Travancore Medical College, 25 Kollam Kerala
How you Prevent Staphylococcal Infections• Prevention• Staphylococcus is a important cause of Hospital infection – Hand antisepsis is the most important measure in preventing nosocomial infections – Also important is the proper cleansing of wounds and surgical openings, aseptic use of catheters or indwelling needles, an appropriate use of antiseptics Dr.T.V.Rao MD Travancore Medical College, 26 Kollam Kerala
• The programme is created forUndergraduate students in Medical Colleges as a part of Applied Microbiology exercises Email email@example.com Dr.T.V.Rao MD Travancore Medical College, 27 Kollam Kerala
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