Staphylococci & Other Catalase-Positive, Gram-Positive Cocci

Micrococcus and CoNS

How are infections caused by gram-positive cocci spread? What type of infection does GPC cause?

• through direct contact with individuals or objects

• pyogenic infection - accumulation of WBCs, bacteria, and fluid at the site of infection

genus Micrococcus

• GPC in tetrads or pairs

• colonizes the skin, but rarely causes infection

• can cause skin infection in immunocompromised individuals

How is Micrococcus identified in the lab? (colony morphology, catalase, oxidase)

• produces pigmented, circular, smooth, convex colonies

• M. luteus is bright yellow in color

• catalase positive

• oxidase positive

general Staphylococcus lab identification (oxygen requirements, catalase, motility, morphology)

• facultative anaerobes

• catalase positive

• can be isolated on MSA

• non-motile

• colony morphology

  • blood agar: raised, convex, creamy colonies

  • CNA: white, cream, or yellow pigment

Where can Staphylococcus sp. be found?

• animals, the environment, and human skin and mucous membranes

• can be a part of normal flora on the skin, nares, axillae, inguinal, and perianal areas

• important human pathogen

What test is used to differentiate S. aureus from other species of Staphylococcus?

• coagulase test

  • S. aureus is positive

  • all other Staphylococci are negative (coagulase-negative staphylococci, CoNS)

Colony morphology of S. aureus on SBA

• white-yellow, raised, or creamy colonies with a smooth surface

• medium to large

• narrow zone of beta hemolysis

What is the most reliable test to ID S. aureus?

Coagulase test

• most strains of S. aureus produce bound coagulase (aka clumping factor)

• detected with the slide coagulase test

• latex agglutination tests can also detect coagulase

Difference between slide and tube coagulase tests

• slide coagulase test

  • bacterial colony is mixed with rabbit plasma

  • in a positive reaction, agglutination occurs (indicating presence of bound coagulase)

• tube coagulase test

  • performed after negative slide tests to determine if free coagulase is present

  • organism is incubated with rabbit plasma at 37 C and then observed for a clot (indicates free coagulase)

  • false-negatives may occur due to fibrinolysin, which can break up the clot (test needs to be read periodically)

S. aureus is DNase:

• positive

  • agar contains DNA and dye

  • S. aureus produces DNase, which breaks down DNA, allowing for a color change in the media

S. aureus can be isolated on chromogenic agar. What is this agar, and what does S. aureus look like when grown on it?

• agar with chromogens (an artificial substrate) added

  • some organisms have enzymes that utilize the chromogens and form colored colonies

• S. aureus: deep pink-fuchsia on HARDYchrome agar

S. aureus is a common HAI and can also cause CAIs. What type of infections does this organism cause?

• skin and soft tissue infections

  • boils, folliculitis, bullous impetigo, scalded skin syndrome, acne, and acute or post-op wound infections

  • usual specimens: wounds, sputum, blood, urine, abscesses

List the 4 virulence factors of S. aureus

• Protein A

  • binds to antibody and inhibits complement activation

• Capsules

• Peptidoglycan and teichoic acids

  • helps with attachment to mucus membranes and withstanding environmental stress

• Penicillin-binding protein 2

  • alters how penicillin-type molecules bind to the bacterium

What 3 exotoxins does S. aureus release to increase its virulence?

• hemolysin

  • lyses host RBCs

• clumping factor/staphylocoagulase

  • aka coagulase, induces fibrin formation, fibrin binds to neutrophils, inhibiting phagocytosis

• staphylokinase

  • dissolves fibrin clots

What enterotoxins does S. aureus produce?

• hemolysin

• heat-stable proteins

  • A, B, C, and D are associated with foodborne illness

  • B and C are associated with enterocolitis

superantigens

• A class of antigens that cause non-specific activation of T-cells, leading to an exaggerated immune response

• they are produced by certain bacteria, including S. aureus

What superantigens does S. aureus release?

• enterotoxins

• toxic-shock syndrome toxin 1 (TSST-1)

  • promotes cytokine release and progression of TSS

  • risk factors of TSS: menstrual products, childbirth, miscarriage, surgical wounds, skin infection, retained foreign objects

Describe the following S. aureus exotoxins: lipase, exfoliative toxins A and B, hyaluronidase, and leukocidin and Panton-Valentine Leukocidin

• lipase

  • hydrolyzes lipids in the skin, allowing staph to colonize and cause infection

• exfoliative toxins A and B

  • proteolytic, hydrolyzes tissues, responsible for scalded skin syndrome

• hyaluronidase

  • lyses hyaluronic acid in CT, helping spread infection

• leukocidin and Panton-Valentine Leukocidin (PVL)

  • enzymes toxic to neutrophils and macrophages, inhibiting phagocytosis

List some S. aureus infections

• prosthetic joint infections

  • forms biofilms around implants

• pneumonia (opportunistic)

• bone infections

  • osteomyelitis

• toxic shock syndrome

antibiotic resistance patterns of S. aureus

• can produce beta-lactamase, causing resistance to the penicillins and other beta-lactams

  • such as methicillin and vancomycin

• can also be resistant to clindamycin

How is MRSA detected in the lab?

• cefoxitin disk screen test

• chromogenic agars

• latex agglutination

• molecular methods

  • detects mecA and mecC genes

What are the 3 strains of S. aureus that are susceptible and resistant to vancomycin?

• vancomycin-susceptible S. aureus (MIC of < 2 mg/mL)

• vancomycin-intermediate S. aureus (VISA) (MIC of 4-8)

• vancomycin-resistant S. aureus (VRSA) (MIC of 16+)

S. aureus clindamycin resistance

• resistance may only be detected if the S. aureus is also exposed to erythromycin

  • inducible-clindamycin reistance (ICR)

• ICR of S. aureus can be detected by a D test

  • lawn of bacteria is grown on Mueller-Hinton agar

  • clindamycin disk and erythromycin disk are placed next to each other

What are the Coagulase-Negative Staphylococci?

• S. epidermidis, S. saprophyticus, S. lugdunensis, S. hominis, and S. haemolyticus

• once considered contaminants and normal flora, but are now opportunistic pathogens

  • elderly, infants, chronically ill, those with prosthetics

• they can cause blood infections, prosthetic biofilms, UTIs, and endocarditis

general lab identification of coagulase-negative staphylococcus

• GPC in clusters

• catalase positive

• coagulase negative

• DNase negative

• grows on MSA, but does not ferment mannitol

How is S. epidermidis identified in the lab?

• colony morphology

  • medium colonies that are white-gray, creamy, raised, and gamma-hemolytic

• coagulase-negative, catalase-positive

• DNase negative

• novobiocin susceptible

What infections does S. saprophyticus cause, and how is it identified in the lab?

• contaminant unless found in urine

  • causes UTIs (cystitis, pyelonephritis, catheter-associated UTIs)

  • can induce UTIs with low colony counts

• lab ID

  • coagulase negative

  • DNase negative

  • resistant to novobiocin

  • grows on MSA with variable fermentation

S. lugdunensis

• opportunistic pathogen that causes aggressive infections with a high mortality rate

• associated with catheter-associated bacteremia, prosthetic joint infections, shunt infections, endocarditis, and UTIs

• positive PYR and ODC

CoNS species: S. haemolyticus, S. hominis, and S. intermedius

• S. haemolyticus

  • BETA hemolytic, associated with endocarditis and bacteremia

• S. hominis

  • rare cause of septicemia in immunocompromised hosts

• S. intermedius

  • common veterinary pathogen - dog bite wounds

S. schleiferi subsp. coagulans and S. warneri

• S. schleiferi

  • veterinary pathogen

  • endocarditis and septicemia in humans

• S. warneri

  • human and animal pathogen

  • can cause spontaneous abortions, UTIs, meningitis, or endocarditis

penicillin binding protein (PBP)

A set of proteins found in bacterial cell membranes that play a crucial role in building the cell wall, making them key targets for beta-lactam antibiotics

• PBP2 and PBP2a

  • allowing cross-linking of peptidoglycan layers

  • targets for beta-lactam binding