Staphylococcus and Micrococcus

Staphylococcus lugdunensis on blood agar

Colonies resemble S. epidermidis colonies; however, they tend to be cream colored

Ornithine decarboxylase test for ID of Staphylococcus lugdunensis

Unlike most other CNS species, S. lugunensis is ornithine decarboxylase positive The decarboxylase medium containing 1% ornithine is inoculated and incubated overnight Since some strains of S. epidermidis can also be positive at 24 h, the specimen should be examined at 8 h, a time at which S. lugdunensis is positive but S. epidermidis is negative Negative = yellow, fermentation of glucose only Positive = violet color, resulting from the alkalization of the medium

Staphylococci colonial morphology

Upon incubation in air at 35 C for 24 to 48 h, staphylococci grow rapidly on a variety of media, with colonies that range from 1 to 3 mm in diameter On blood agar, staphylococci produce white to cream opaque colonies S. aureus colonies typically are cream in color but occasionally have a yellow or golden pigement; can be beta-hemolytic; it is not uncommon to see both large and small colonies in the same culture, a phenotypic characteristic shared by several MRSA strains CNS, in particular S. epidermidis, produce white colonies; however, other CNS strains and species can have colonies with a slight cream pigment CNS strains are nonhemolytic; however, some produce a small zone of beta-hemolysis on blood agar

Tests to differentiate Micrococcus

Urease Acid production from carbohydrates Esculin Gelatin Bacitracin, lysostaphin, and furazolidone have been used to differentiate Staphylococcus from Micrococcus Satphylococcus is resistant to bacitracin (0.04-U disk); Micrococcus is susceptible Micrococcus is resistant to furazolidone (100-ug disk) and lysostaphin (200-ug disk); Staphylococcus is susceptible

Staphylococcus aureus on blood agar

Colonies are cream colored and opaque and have a smooth entire edge A zone of beta-hemolysis surrounds the colony

Boils

A boil is a skin infection that starts in a hair follicle or oil gland At first, the skin turns red in the area of the infection, and a tender lump develops After four to seven days, the lump starts turning white as pus collects under the skin The most common places for boils to appear are on the face, neck, armpits, shoulders, and buttocks When one forms on the eyelid, it is called a sty If several boils appear in a group, this is a more serious type of infection called a carbuncle

Coagulase tube test

A common method used to distinguish S. aureus from other Staphylococcus spp. S. aureus - positive CNS - negative Colonies of the isolate to be identified were emulsified in 0.5 ml of rabbit plasma The tube was incubated at 35 C for 4 h and tipped gently to look for clot formation

Micrococcus luteus on blood agar

A distinguishing feature of M. luteus is the vivd yellow colonies it produces Micrococcus is slower growing than Staphylococcus

Mannitol salt agar containing oxacillin

Can be used to screen for the presence of MRSA in nasal specimens since the 7.5% salt and 6 ug of oxacillin in this medium inhibit most other organisms that normally colonize the nares MRSA turns the medium yellow as a result of the fermentation of mannitol

Bacitracin susceptibility

Can differentiate Staphylococcus spp,,, which are bacitracin resistant, from Micrococcus spp., which are susceptible Bacitracin disks (A) 0.04-U

Modified oxidase test

A modified oxidase test, the Microdase test, is available for differentiating Micrococcus from Staphylococcus Micrococcus spp. posess cytochrome c, which is essential for producing a positive oxidase reaction, whereas clincally relevant Staphylococcus spp. are microdase negative since they lack cytochrome c Organism is rubbed onto a disk impregnated with tetramethyl-p-phenylenediamine (TMPD) dissolved in dimethyl sulfoxide Development of a purple-blue color within 2 min indicates a positive test due to the reaction of the enzyme oxidase with cytochrome c and TMPD

Mannitol salt agar

A selective and differential medium used for the isolation and presumptive ID of S. aureus The high salt concentration inhibits the growth of many organisms that inhabit the skin and mucosal membranes The phenol red indicator incorporated into the medium detects acid production (yellow) resulting from the fermentation of mannitol by S. aureus

Testing for MRSA

Can be difficult due to heteroresistance, in which the resistance is expressed to a different extend among subpopulations A molecular test to directly detect the mecA gene and rapid assay formats employing monoclonal abs to the altered PBP2a protein have been used to circumvent the problems of in vitro susceptibility testing for MRSA Screening agar plates incorporating 6 ug of either oxacillin or vancomycin are commercially available to screen for MRSA and VISA strains Selective chromogenic agars have facilitated the detection of MRSA from nares specimens

S. aureus infections

Can produce disease mediated by toxins or by direct invasion and destruction of tissues Infections range from superficial skin infections to fatal systemic infections that can occur when the integrity of skin is damaged, thus giving this pathogen access to sterile sites More common infections are boils, folliculitis, cellulitis, and impetigo Immunocompromised hosts are at particular risk of infection Systemic infections include septicemia, which can result in the seeding of distant sites, producing osteomyelitis, pneumonia, endocarditis, scalded-skin syndrome, and TSS Scalded-skin syndrome and TSS are caused by toxigenic strains S. aureus is also capable of producing food poisoning due to the elaboration of enterotoxins in foods such as potato salad, ice cream, and custards Intense vomiting and diarrhea usually occurs within 2 to 8 h after ingestion of food containing the toxin

Golden pigment of Staphylococcus aureus

Capable of producing golden pigment Strains with this degree of pigment are not frequently isolated from clinical specimens When left at RT or refrigerated following incubation, isolates tend to develop more intense pigment

Staphylococcus and Micrococcus characteristics

Catalase positive Gram-positive cocci in pairs and clusters

Cellulitis

Cellulitis is a common skin infection that happens when bacteria spread through the skin to deeper tissues Most cases are mild and last several days to a couple of weeks But cellulitis can sometimes progress to a more serious infection, causing severe illness that affects the whole body (sepsis) or other dangerous problems At first, the infected area will be warm, red, swollen, and tender If the infection spreads, you may have a fever, chills, and swollen lymph nodes

Endocarditis

Endocarditis is inflammation of your heart's inner lining, called the endocardium This condition is also called infective endocarditis It's usually caused by bacteria Endocarditis is uncommon in people with healthy hearts

Folliculitis

Folliculitis is an inflammation of the hair follicles Each hair on your body grows out of a tiny pouch called a follicle You can have folliculitis on any part of your body that has hair But it is most common on the beard area, arms, back, buttocks, and legs Folliculitis usually looks like red pimples with a hair in the center of each one The pimples may have pus in them, and they may itch or burn When the pimples break open, they may drain pus, blood, or both

Micrococcus microscopic and colonial morphology

Form pairs and clusters, can appear as tetrads Can be easily grown in the lab and can be recovered from a variety of media They are slower growing with smaller colonies present after 24 h of incubation at 35 C in comparison to staphylococci Depending on the species, the colony color can range from cream to yellow (M. luteus) or rose red

Gram stain of Staphylococcus aureus

Gram-positive cocci in grape-like clusters

Impetigo

Impetigo is a bacterial skin infection It causes red sores that can break open, ooze fluid, and develop a yellow-brown crust These sores can occur anywhere on the body. Impetigo is one of the most common skin infections in children It can occur in adults but is seen far more often in children Impetigo is contagious and can be spread to others through close contact or by sharing towels, sheets, clothing, toys, or other items Scratching can also spread the sores to other parts of the body

Thermostable endonuclease activity

In addition to DNase, S. aureus produces a thermostable endonuclease that can also cleave DNA To test for this organism, a heavy suspension of the organism is boiled and then used to fill a well that is cut in the DNA plate containing toluidine blue If the DNA is degraded there is a change in the color of the agar from blue to pink

MRSA

In the majority of MRSA strains, this is due to an alteration in penicillin binding protein PBP2a, with is encoded by the mecA gene Overproduction of beta-lactamase accounts for a smaller percentage of MRSA strains

Size variation of Staphylococcus aureus colonies

It is not uncommon for strains of S. aureus, in particular MRSA strains, to produce colonies that are heterogeneous in size and the degree of hemolysis

Latex test for the identification of Staphylococcus aureus

Latex particles have been coated with antibody that can recognize bound coagulase as well as immunoglobulin that will bind to protein A present on the surface of most S. aureus strains As with the slide coagulase test, some strains of MRSA may be negative and some strains of CNS, namely, S. lugdunensis and S. schleiferi strains, may be positive

Gram stain of Micrococcus luteus

M. luteus organisms are gram-positive cocci that, like S. aureus, can appear in pairs and clusters However, they also tend to form tetrads

Coagulase test

Measures the ability to clot plasma by converting fibrinogen to fibrin Useful in distinguishing S. aureus from other bacteria that appear similar A suspension of the organism to be identified is made in rabbit plasma containing EDTA and incubated at 35 C for 4 h The tube is tilted gently, and the presence or absence of clot formation is noted If the test is negative at 4 h, the suspension is incubated for up to 24 h The 4-h reading is important because some strains produce fibrinolysin, which can dissolve a clot upon prolonged incubation, causing a false-negative result Bound coagulase (clumping factor) can be detected by a slide agglutination test, in which a suspension of the organism is emulsified on a slide with a drop of plasma If bound coagulase is present, the organisms agglutinate Of the CNS, S. lugunensis and S. schleiferi can also test positive for bound coagulase but can be differentiated from S. aureus by a negative tube coagulation test

Micrococcus infections

Micrococcus spp. are common inhabitants of the skin and have a fairly low pathogenic potential Infections with these organisms have occurred in immunocompromised hosts M. luteus and related organisms have been implicated in a variety of infections, including meningitis, CNS shunt infections, endocarditis, and septic arthritis

Tests used to differentiate species of Staphylococcus

Novobiocin resistance Phosphatase Production of acetoin Polymyxin susceptibility Pyrrolidonyl arrylamidase activity (PYR) Acid production from carbohydrates

Osteomyelitis

Osteomyelitis is an infection of the bone, a rare but serious condition Bones can become infected in a number of ways: Infection in one part of the body may spread through the bloodstream into the bone, or an open fracture or surgery may expose the bone to infection.

Staphylococcus epidermidis on blood agar

Produces a white colony with little or no pigment

Slide coagulase test

Rapid assay that tests for clumping factor on the surface of the organisms The test is performed by emulsifying the organism to be IDed in both saline, which serves as a control for autoagglutination, and rabbit plasma S. aureus, S. lugdunensis, and S. schleiferi are positive

Coagulase-negative staphylococci (CNS)

Recognized as the leading cause of nosocomial infections, with immunocompromised hosts at increased risk Because CNS are members of the normal skin and mucosal membrane flora, they are frequently considered a contaminant when isolated from clinical specimens and therefore may be overlooked as a cause of infection An important virulence property of CNS is their ability to form a biofilm on the surface of indwelling or implanted foreign bodies, making them frequent agents of intravascular infections

Staphylococcus organisms most frequently associated with human infections

S. aureus S. epidermidis S. saprophyticus S. haemolyticus S. lugdunensis S. schleiferi S. aureus is the most virulent, being a major cause of morbidity and mortality

DNase plate to differentiate Staphylococcus aureus from DNS

S. aureus produces DNase, which can degrade DNA This property is used to aid in the differentiation of S. aureus from CNS This is particularly useful for ID of S. aureus strains that produce a small amount of coagulase, thus giving an equivocal or weakly positive coagulase test The only CNS species that shares this property with S. aureus is S. schleiferi In this test, a heavy inoculum of the organism is used to spot an agar plate that contains DNA and toluidine blue If the organism produces DNase, the DNA is degraded, resulting in the agar turning pink in the area surrounding the inoculum due to the metachromatic qualities of toluidine blue

CNS infections

S. epidermidis has been implicated as a cause of endocarditis and is associated with right-side endocarditis in intravenous drug users S. saprophyticus is a leading cause of non-complicated UTIs in young, sexually active females S. lugdunensis and S. schleiferi have been implicated in serious infections including endocarditis, septicemia, arthritis, and joint infections S. lugdunensis, which at times can behave more like S. aureus than CNS, has been associated with aggressive infections that have a high mortality rate

Novobiocin susceptibility

S. saprophyticus can be differentiated from other clinically significant CNS isolates by its resistance to the antibiotic novobiocin Mueller-Hinton agar was inoculated with a suspension equivalent to a 0.5 McFarland standard Novobiocin disks (5 ug) (NB 5) were placed on the agar surface, which was incubated for 24 h at 35 C Zones of inhibition measuring <16 mm indicate novobiocin resistance

Scalded-skin syndrome

Scalded skin syndrome is a skin infection caused by bacteria in which the skin becomes damaged and sheds Scalded skin syndrome (SSS) is caused by infection with certain strains of Staphylococcus bacteria The bacteria produce a toxin that causes the skin damage The damage creates blisters, as if the skin were scalded These blisters can occur;at areas of the skin away from the initial site SSS is found most commonly in infants and children under the age of 5

Spectra MRSA

Selective and differential agar that can e used to detect MRSA When the chromogenic substrate incorporated into the inhibitory agar is degraded by the enzymatic action of MRSA, the colony takes on a denim blue color

Lysostaphin susceptibility

Several species of Staphylococcus are susceptible to the endopeptidase lysostaphin, which cleaves the glycine-rich pentapeptide that is essential for cross bridging the cell wall Cleavage of these basic units weakens the cell wall, making it susceptible to lysis Depending on the makeup of this pentapeptide, specifically the glycine content, susceptibility to lysostaphin can vary S. aureus is very susceptible S. saprophyticus is less susceptible due to the serine content of its pentapeptide bridge Micrococcus and related species are not susceptible to lysostaphin Test is performed by making a heavy suspension of the unknown organism in saline and then adding an equal volume of lysostaphin reagent Clearing of the suspension after 2 h at 35 C indicates lysis of the organisms

S. aureus agar

Since S. aureus is frequently isolated in mixed cultures, selective and differential media are used to facilitate the detection of these organisms in clinical material particularly in nasal swabs Mannitol salt agar - the high concentration of salt (7.5%) inhibits many other organisms Mannitol, along with the phenol red indicator in the medium, facilitates the discrimination of S. aureus, which can ferment mannitol, from most CNS

DNase or thermostable-endonuclease test

Strains of S. aureus that produce a weak coagulase reaction can be further tested by DNase or a thermostable-endonuclease test S. aureus and S. schleiferi possess enzymes that can degrade DNA, a DNase and a thermostable endonuclease Both tests use the same basic medium containing agar that incorporates DNA and the metachromatic dye toluidine blue O A heavy suspension of organisms is spotted onto the plate; after 24 h of incubation at 35 C, a pink haze appears around the colony, in contrast to the azure blue of the medium When testing for the thermostable endonuclease, a suspension of the organism is boiled before being placed on the DNA plate

Assays to detect PBP2a protein found in MRSA

The produce of the mecA gene, which results in methicillin resistance, is an altered penicillin binding protein, PBP2a Monoclonal antibody to this altered protein has been used to coat latex particles, which are then used in the Oxoid agglutination assay to detect PBP2a Alere PBP2a lateral flow assay - also utilizing monoclonal antibodies for the detection of PBP2a protein Both formats are rapid tests that are used once the organism is isolated on solid media

Staphylococcus and Micrococcus commonly colonize...

the surface of skin and mucosal membranes of mammals and birds