Staphylococcus (CH 14) PDF

CLINICAL BACTERIOLOGY_LECTURE STAPHYLOCOCCI (CH 14) COAGULASE GENERAL CHARACTERISTICS v Staphylocoagulase enzyme clots plasma. v Gram-positive cocci v Coagula...

CLINICAL BACTERIOLOGY_LECTURE STAPHYLOCOCCI (CH 14) COAGULASE GENERAL CHARACTERISTICS v Staphylocoagulase enzyme clots plasma. v Gram-positive cocci v Coagulase-positive staphylococci Ø Catalase positive Ø S. aureus: human pathogen Ø Singly, in pairs, and in clusters Ø S. delphini, S. intermedius, S. lutrae, S. Ø “Bunches of grapes” hyicus: animal pathogens v Morphology v CoNS Ø Appear creamy, white, or, rarely, light gold Ø S. epidermidis: hospital-acquired infections Ø Buttery looking Ø S. saprophyticus: urinary tract infections Ø Some species produce β-hemolysis (UTIs) in young, sexually active females · S. aureus · Important in urine samples v Characteristics Ø Nonmotile CoNA Ø Non-spore forming Ø Non-encapsulated Ø Aerobic or facultative anaerobe · One exception S. saccharolyticus (obligate anaerobe) v Rare strains Ø Small colony variants · 1/10 size MICROCOCCUS v Micrococcus luteus Ø Gram-positive cocci Ø Catalase positive · Note: modified catalase containing mild detergent CLINICALLY SIGNIFICANT STAPHYLOCOCCI Ø Coagulase negative v S. aureus v Morphology Ø Habitat: anterior nares (carriers) (20% to Ø Distinct yellow pigmented colony 30% of humans) v Generally considered a nonpathogen Ø Primary pathogen of the genus Ø Can be recovered from almost any clinical DIFFERENTIATING STAPHYLOCOCCI AND specimen MICROCOCCI v Produce superficial to systemic infections Ø Skin Ø Bacterial sepsis VIRULENCE FACTORS OF S. AUREUS v Enterotoxins Ø Heat-stable exotoxins that cause diarrhea and vomiting v Toxins A-E and G-I (eight total) Ø Resistant to gastric acid and associated with food poisoning · A, B, and D are associated with staphylococcal food poisoning. Ø Toxic shock syndrome toxin-1 (TSST-1) Ø TSST-1 and B, C, G, and I are superantigens. Ø Causes toxic shock syndrome Ø Produced by phage group I  CLINICAL BACTERIOLOGY_LECTURE VIRULENCE FACTORS OF S. AUREUS (CONT.) EPIDEMIOLOGY OF S. AUREUS v Exfoliative toxin (epidermolytic toxin) v Primary reservoir Ø Causes the epidermal layer of the skin to Ø Nares slough off and is known to cause Scalded v Other reservoirs Skin Syndrome or Ritter’s disease Ø Axillae, vagina, pharynx, and other skin · Also associated with bullous impetigo surfaces v Hospital outbreaks CYTOLYTIC TOXINS OF S. AUREUS Ø Nurseries v Hemolysins Ø Burn units Ø Three main types α, β, δ (there is a γ but Ø Surgical patients generally not important) v Transmission · α hemolysin destroys platelets and Ø Unwashed hands, fomites tissues. · β hemolysin shows enhanced activity by INFECTIONS CAUSED BY S. AUREUS acting on the sphingomyelin of red blood v Skin and wound infections cell (RBC) membranes, causing lysis. Ø Pus formers - Hot-cold lysin because it works best Ø Furuncle (boil) at 37°C and very wellwhen stored at · A painful inflammation of the skin and 4°C subcutaneous tissue · δ hemolysin causes injury to cells and Ø Carbuncles leukocytes but is less lethal. · Boils that have multiple lesions and may CYTOLYTIC TOXINS OF S. AUREUS (CONT.) progress into deeper tissues v γ-hemolysin INFECTIONS CAUSED BY S. AUREUS (CONT.) Ø Also called Panton-Valentine leukocidin v Skin and wound infections (PVL) Ø Folliculitis · Exotoxin kills polymorphonuclear · Infection of the hair follicle leukocytes Ø Bullous impetigo · Helps prevent phagocytosis · Large pustules surrounded by small zone Ø Often associated with community-acquired of erythema (redness) infections · Spread by direct contact and fomites; highly contagious VIRULENCE FACTORS OF S. AUREUS (CONT.) v Sometimes occur due to blocked follicles, v Enzymes sebaceous glands, and sweat glands Ø Coagulase · Very diagnostic but importance in SCALDED SKIN SYNDROME virulence is NOT completely understood v Extensive exfoliative dermatitis Ø Hyaluronidase Ø Staphylococcal scalded skin syndrome (SSSS) · Hydrolyzes the hyaluronic acid present · More likely to occur in renal failure in connective tissues, helping spread of patients infection · Immunocompromised Ø Lipase v Severity ranges from mild to severe · Breakdown of the fats and oil created Ø Localized lesion to large generalized area by the sebaceous glands on skin surfaces with profuse peeling of the epidermal layer v Protein A v Lasts about 2 to 4 days Ø Bind the fragment crystallizable (Fc) portion Ø Spontaneous recovery in children of antibodies to avoid phagocytosis Ø Adult cases can lead to mortality. · Masking of its immunogenic proteins with host proteins to look like “self” · Assists in blocking phagocytosis · Negates the protective effects of immunoglobulin G (IgG)  CLINICAL BACTERIOLOGY_LECTURE TOXIC SHOCK SYNDROME S. EPIDERMIDIS v Association with super-absorbent tampons v Predominantly nosocomial infections v Clinical presentation Ø Skin flora gets introduced in catheters, heart Ø High fever valves, cerebrospinal fluid (CSF) shunts Ø Rash of trunk spreading to extremities Ø Produces a slime layer (biofilm) that helps Ø Watery diarrhea adherence to prosthetics and avoidance of Ø Vomiting phagocytosis · Dehydration Ø Common cause of hospital-acquired UTIs v Leads to hypotension v Disseminated intravascular coagulation (DIC) S. SAPROPHYTICUS v Increase in blood urea nitrogen (BUN) and v UTIs in young, sexually active women creatinine Ø Due in part to increased adherence to v Fatal in 2% to 5% of cases due to multiorgan epithelial cells lining the urogenital tract system failure v Rarely present in other skin areas or mucous membranes TOXIC EPIDERMAL NECROLYSIS (TEN) v Urine cultures v Multiple causes Ø If present in low amounts (less than Ø Drug induced 10,000), it is still considered significant. Ø Infections Ø Vaccines OTHER CoNA · Cause officially NOT known v S. lugdunensis v Similar to SSSS Ø Can be more virulent than S. aureus and Ø Treatment with steroids helpful can mimic S. aureus infections · Unlike SSSS · Known to carry mecA v High mortality rate Ø Important pathogen in infective endocarditis, septicemia, meningitis, skin and soft tissue FOOD POISONING infections, UTIs, and septic shock v Toxin, NOT bacterial growth, causes disease. · Aggressive in endocarditis, causing valve v Enterotoxin A, B, D (A and D most common) replacement Ø From enterotoxin-producing strains Ø Clumping factor can mimic positive contaminating the rich foods (mayonnaise) coagulase slide test. Ø Inadequate refrigeration OTHER CoNA (CONT.) v Symptoms v Opportunistic pathogens Ø Appear rapidly about 2 to 8 hours after Ø S. warneri, S. capitis, S. simulans, S. hominis, ingesting food and S. schleiferi Ø Usually resolve in 24 to 48 hours, sometimes · Normal flora but can be involved in less endocarditis, septicemia, and wound Ø Nausea, vomiting, abdominal pain, and infections cramping v S. haemolyticus Ø Common CoNS OTHER INFECTIONS Ø Can be found in wounds, bacteremia, v Secondary pneumonia endocarditis and UTIs Ø After influenza A infection but relatively · May have resistance to vancomycin RARE · High mortality rate SPECIMEN COLLECTION AND HANDLING v Bacteremia and endocarditis v Transport to the laboratory without delay Ø Intravenous (IV)-drug addicts presenting Ø Prevents drying with fever Ø Maintains the proper environment Ø Enter through injection site Ø Minimizes the growth of contaminating v Osteomyelitis organisms Ø Occurs secondary to bacteremia and results v Requires NO special procedures in bacteria invading the bone Ø Specimens should be taken from the site of Ø Fever, chills, swelling, and pain around the infection. infected area Ø Cleanse surrounding area to avoid v Arthritis if bacteria in the joint contamination  CLINICAL BACTERIOLOGY_LECTURE MICROSCOPIC EXAMINATION IDENTIFICATION METHODS v Numerous gram-positive cocci v Oxidative-fermentative (O/F) glucose medium v Polymorphonuclear leukocytes Ø Most staphylococci ferment glucose. (PMNs) · Micrococci fail to ferment glucose v Purulent exudates, joint fluids, anaerobically. aspirated secretions - However, does NOT work for all v Aspirate is best. staphylococci v If swabs, clinicians should send two. v Modified oxidase Ø Microdase disk will be positive for Micrococci. Ø Most staphylococci are negative. v See Table 14-3 STAPHYLOCOCCUS SPP. IDENTIFICATION TESTS: CATALASE ON GRAM STAIN v Principle: tests for enzyme catalase Ø 2 H2O2 in presence of catalase becomes 2 H2O + O2 v Drop H2O2 onto smear v Bubbling = positive (most bacteria, O2 generated) v No bubbling = negative (streptococci and other ISOLATION OF STAPHYLOCOCCI lactic acid bacteria, no O2 generated) v Grow easily on blood agar plates and thioglycolate COAGULASE v If heavily contaminated, they can be selected v Clumping factor (formerly cell-bound with the following. coagulase) Ø Mannitol salt agar Ø Causes agglutination in human, rabbit, or pig Ø Columbia colistin-nalidixic acid agar (CNA) plasma Ø Phenylethyl alcohol agar v Slide test method Ø CHROMagar Staph aureus Ø Mix water or saline suspension of organism with a small amount of rabbit plasma · Selective media Ø Check for clumping (if clumping, then positive) v If clumping is negative, a tube test should be performed. Ø Why? · 5% do NOT produce clumping factor COAGULASE (CONT.) v Extracellular free coagulase Ø Extracellular enzyme secreted that clots plasma Ø Tube test · Check for coagulation 4 hours after inoculation and 24 hours after - Prevents autolysis and false-negative result v Hallmark test for S. aureus v Other staph can produce positive coagulation Ø Pyrrolidonyl arylamidase (PYR) test is NEGATIVE for S. aureus but POSITIVE for the other coagulase-positive staphylococci.  CLINICAL BACTERIOLOGY_LECTURE DIFFERENTIATING CoNS v If it is a urine sample Ø Test for S. saprophyticus v Presumptive identification can be done using novobiocin (5 μg). Ø Streak organism onto plate and add novobiocin disk to heavy growth quadrant v If resistant, S. saprophyticus v If susceptible, likely S. epidermidis Ø Can be others, would use tests in Table 14-5 Staphylococci-Producing Clumping Factor S. AUREUS v Mode of transmission: traumatic introduction Ø Needle stick Ø Destruction of skin layers (burns, road rash) Ø Medical procedures v Predisposing conditions Ø Chronic infections Ø Indwelling devices Ø Skin injuries Ø Immune response defects SEPARATING MICROCOCCUS v Bacitracin disk test Ø Micrococcus · Susceptible (Micrococcus luteus) · Lemon yellow color of colony does not hurt -. Do NOT produce acid under anaerobic conditions in glucose O/F media Ø CoNS · Resistant (S. saprophyticus, S. epidermidis, or others)  CLINICAL BACTERIOLOGY_LECTURE v Health care–associated/community onset MRSA (HACO-MRSA) v Hospital-associated MRSA (HA-MRSA) Ø Pose serious threats to health institutions METHICILLIN-RESISTANT STAPHYLOCOCCI (CONT.) v Infection control Ø Barrier protection Ø Contact isolation Ø Handwashing v Treat with vancomycin Ø Test for susceptibility with cefoxitin disk v Borderline oxacillin-resistant S. aureus (BORSA) Ø Can separate from MRSA on oxacillin salt agar plate RAPID IDENTIFICATION v CLSI document M100 v BBL staphyloslide · Recommends cefoxitin to be used to v Staphaurex® determine methicillin resistance v BactiStaph® v mecA gene Ø Figure 14-9 · Encodes penicillin-binding proteins v Plasma-coated carrier particles detect both (PBPs) clumping factor and protein A. · Only a small fraction of the population Ø Particularly useful for identification of express the phenotype despite having the methicillin-resistant S. aureus (MRSA), which genetic potential. are weakly positive or negative in slide v Gold standard coagulase test · mecA gene detected by PCR RAPID IDENTIFICATION (CONT.) v Real-time polymerase chain reaction (PCR) VANCOMYCIN-RESISTANT STAPHYLOCOCCI Ø For MRSA and methicillin-sensitive S. aureus v Vancomycin-resistant S. aureus (VRSA) and (MSSA) vancomycin-intermediate S. aureus (VISA) v Qualitative nucleic acid hybridization assays Ø Isolated in United States 2002 targeting rRNA sequences for staphylococcal identification from blood cultures MACROLIDE RESISTANCE v Matrix-assisted laser desorption-ionization v Resistance to clindamycin may not be obvious time-of-flight (MALDI-TOF) mass spectrometry Ø Erythromycin and clindamycin should have Ø Continues to be integrated as U.S. Food and SAME resistance patterns. Drug Administration (FDA) approval occurs · If NOT, perform D-zone test v D-zone test ANTIMICROBIAL SUSCEPTIBILITY Ø Use erythromycin and clindamycin disks v Standard guidelines issued by Clinical and · Growth between disks but NOT on side of Laboratory Standards Institute (CLSI) clindamycin disk v Most S. aureus are resistant. - Inducible clindamycin resistance Ø Thus, do β-lactamase test v Serious infections Ø Require susceptibility testing METHICILLIN-RESISTANT STAPHYLOCOCCI v MRSA v Methicillin-resistant S. epidermidis Ø MRSE v Community-acquired MRSA (CA-MRSA) Ø Can be greater than 50% of isolates in some areas