Department of PathologyRawalpindi Medical College

ByProf. Dr. Abbas Hayat Balouch

PYOGENIC COCCI

Gram Positive cocci:Staphylococci (Catalase Positive)Streptococci (Catalase Negative)

Streptococci pneumoniae Gram Negative cocci:

Niesseria Species

Common Pyogenic Cocci

STAPHYLOCOCCUS

STAPHYLOCOCCUSImportant Classification

Species Catalase CoagulaseNovobioci

n Sensitivity

DNAse

Staph. aureus

Positive Positive Sensitive Positive

Staph. epidermidis

Positive Negative Sensitive Negative

Staph. saprophyticus

Positive Negative Resistant Negative

Staph. lugdunensis

Positive

Positive (Slide)

Sensitive NegativeNegative

(Tube)

Important Features• Except for Staph. aureus, all other

staphylococci are coagulase negative. • They are salt tolerant and often

hemolytic.• Staph. lugdunensis can gives a

positive slide-coagulase test (bound coagulase) but a negative tube-coagulase test (free coagulase). DNAse negative. Commensal . Can cause osteomyelitis, septicaemia, and aggressive endocarditis.

• Final Identification requires biotype analysis.

Structure: Staphylococci are Gram-positive cocci 1 µm in

diameter. They form clumps/clusters.

►S. aureus colonizes the nasal passage and axillae.

►S. epidermidis is a common human skin commensal.

►S. saprophiticus is a commensal of female reproductive tract

►Other species of staphylococci are infrequent human commensals. Some are commensals of other animals.

►Gram positive cocci ►Morphologically striking clusters, due to

division in 3 successive planes- grape-like clusters (Greek: grape = staphylos)

► Facultative anaerobes- grow better aerobically - some better in presence of CO2

►Golden pigment- produced by many strains (Latin: gold = aureus)- especially with extended incubation

►S. aureus very resistant to drying, certain disinfectants (HgCl2, phenol), salt (7.5 - 10.%),

Important Characteristics

Three Important Species ►S. epidermidis

- usually coagulase negative, disease in compromised hosts only- minor abscesses- postoperative endocarditis

►S. saprohyticus- community acquired urinary tract infections in females

►S. aureus- coagulase positive     * skin infections (impetigo), boils, styes- serious disease possible    * deep abscesses, wound infections, osteomyelitis    * purulent arthritis, pneumonia, meningitis, septicemia, endocarditis    * exfoliative skin disease   

* infection of spermatic cord stump following castration     * cattle: mastitis, favored by automatic milking machines - also causes intoxications / food posioning

Golden Yellow growth of Staph. aureus with Beta-hemolysis

White growth of Staph. epidermidis

Pyogenic cocci Gram stain under the

microscope.

TOXINS & ENZYMES►(a) a-toxin (b) ß-toxin

(c) d-toxin (d) g-toxin (e) Leukocidin

►Superantigens: enterotoxins and toxic shock syndrome toxins

►Epidermolytic (exfoliative) toxin (ET) This toxin causes the scalded skin syndrome

in neonates, with widespread blistering and loss of the epidermis

Other Extra cellular Proteins: Coagulase Staphylokinase

Many strains of S aureus express a plasminogen activator called Staphylokinase.

As with coagulase there is no evidence that staphylokinase is a virulence factor, although it seems reasonable to imagine that localized fibrinolysis might aid in bacterial spreading.

Pathogenesis►S aureus expresses many potential

virulence factors.

(1) Surface proteins that promote colonization of host tissues.

(2) Factors that probably inhibit phagocytosis (capsule, immunoglobulin binding protein A).

(3) Toxins that damage host tissues and cause disease symptoms. Coagulase-negative staphylococci are normally less virulent and express fewer virulence factors.

S. epidermidis readily colonizes implanted devices.

Superficial Folliculitis Deep Folliculitis Furuncle

Carbuncle Impetigo Scalded Skin Syndrome

A) Abscess/Furuncle:  (``Phoora``)

Localized, begins when organism enters hair follicle. Organism spreads to surrounding tissues. Tiny lesion becomes larger and swollen, inflamed. The localized lesion walled off by deposition of fibrin by the tissue, walling off is to prevent the Staph. infection from going further.  There is a lot of pus (organism is Pyogenic). The main Pyogenic organisms are Staph. Aureus.

Treatment: Lancing: Allows drainage to "get rid of the infection". This gets rid of the material present so that antibiotics can work.

(Antibiotics cannot diffuse through non-living tissue.)

... Drainage does get rid of some of the organisms, but more importantly gets rid of the fluid in which they reside. After lancing, the abscess is treated topically with penicillin, orally with penicillin, or some other suitable antibiotics.

►B) Septicemia: S. aureus gains access to the blood. This may result from a cutaneous infection, contamination during surgery, or from the organism (S.a. is a skin resident) entering via a scratch on the skin.   Blood poisoning is a chronic septicemia usually caused by Staphylococcus aureus.

C. Impetigo:  Occurs in neonates (impetigo of the newborn) and in addition there are many cases in older children.

In newborns: The organism overgrows the skin rapidly, owing to the lack of other resident bacteria. The organism may then enter the dermis through an abrasion or a hair follicle. Causes clustered lesions anywhere on the body, which crust over, then break open. The disease is spread further by scratching and rubbing against sheets. Prevented in nurseries by using lotions containing hexachlorophene.

Treat with penicillin. In children: Similar, but not as aggressive spreading in older

children because the resident skin bacteria are well established. Is considered contagious, spread easily by children wrestling and fighting. Occurs mostly on face.

Impetigo

►D) Staphylococcal Scalded Skin Syndrome (SSSS): Usually in neonates, when a strain of S.a. which excretes exfoliatin (a pathogenic factor) invades the skin. Rapid invasion of the skin occurs in neonates because competing bacteria have not yet become established.

* The skin becomes reddened and sloughs off rapidly, causing causing drastic fluid loss.* May be fatal.

* Treat with penicillin.

Pt. Name: Abbas HayatDate:6TH March 2013

E) Minor Skin Infections: ►Pimples-localized and minor. Infection of

single hair follicle.

►Sty--minor Staph. skin infection. ►Follicle involved is the eyelash.

“Acne and pimples areembarrassing”

► F) Toxic Shock Syndrome (TSST-1): Super-absorbent types of tampon or wound packing material can harbor Staph. aureus and bind Mg. Low Mg availability to Staph. aureus triggers

production of TSST-1. Labile B/P, increase nausea, fever, rash, finger tips peel and swell. May be fatal.

Penicillin is treatment.

►G) Staphylococcal Food Poisoning: Factor is enterotoxin F. It grows in certain foods. (Cream filled and starchy.) e.g. Potato salads. Organism gets in easily by handling potatoes and

eggs. It is a food toxicity, not infectious disease. It lasts 24 hours and the incubation period is two hours. There is no treatment.

►G) Staphylococcal Food Poisoning: Factor is enterotoxin F. It grows in certain foods. (Cream filled and starchy.)

Potato salad-organism gets in easily by handling potatoes and eggs. It is a food toxicity, not infectious disease. It lasts 24 hours and the incubation period is two hours.

There is no treatment.

►H. Osteomyelitis: Staph. aureus gains access to bone and proliferates. It colonizes surface and deteriorates its. Becomes chronic. Can be surgically removed but surgery damages bone tissue. Treatment is penicillin.

►I. Pyelonephritis: Staph. aureus infection of the kidneys.

Be mindful that most cases of Pyelonephritis are caused by Escherichia coli.  Whatever organism causes this infection typically ascends from bladder infection (cystitis) or hematogenous spread

Identification of Staphylococci in the Clinical

laboratory

►Laboratory Methods In addition to the several lab tests (Catalase, glucose fermentation, coagulase, etc.) in lab for identification of  G+ cocci, there are a few useful media used in isolation and identification of Staphylococcus:

►The catalase test is important in distinguishing streptococci (catalase-negative) from staphylococci which are catalase positive.

►The test is performed by flooding an agar slant or broth culture with several drops of 3% hydrogen peroxide. Catalase-positive cultures bubble at once.

►The test should not be done on blood agar because blood itself will produce bubbles.

► Blood Agar: Routinely used also differentiates between S. aureus and epidermitis, colonies > 1 mm in diameter

► Mannitol-Salt Agar:MSA agar is useful in selectively isolating Staphylococcus species from clinical specimens, as well as skin, etc. It contains 7.5% NaCl to inhibit all bacteria except Staph.

► DNAase Agar: This medium contains DNA. Colonies of Staph. aureus produce DNAase, and the area around the colonies clears, while the other species don't produce this enzyme and no clearing occurs.

► Phage Typing: Used to identify different strains of Staph. aureus. Often used to trace source of a Staph. aureus strain which has caused a rash of infections in particular O.R.

Antibiotic Resistance

►Multiple antibiotic resistance is increasingly common in S aureus and S epidermidis.

►Methicillin resistance is indicative of multiple resistance. Methicillin-resistant S aureus

►(MRSA) causes outbreaks in hospitals and can be epidemic.

Resistance of Staphylococci to Antimicrobial Drugs

►Hospital strains of S aureus are often resistant to many different antibiotics. Indeed strains resistant to all clinically useful drugs, apart from the glycopeptides vancomycin and teicoplanin, have been described. The term MRSA refers to methicillin resistance and most methicillin-resistant strains are also multiply resistant to other antibiotics.

Prevention & Control

► Prevention & Control.1. Hand washing.2. Universal Infection control

methods .

►Patients and staff carrying epidemic strains, particularly MRSA, should be isolated. Patients may be given disinfectant baths or treated with a topical antibiotic to eradicate carriage of MRSA.

►Infection control programs are used in most hospitals.

MRSA

Methicillin-resistant Staphylococcus aureus

(MRSA)►Methicillin-Resistant

Staphylococcus aureus (MRSA) is a bacterium responsible for several difficult-to-treat infections in humans.

►MRSA is any strain of Staphylococcus aureus that has developed resistance to beta-lactam antibiotics, which include the penicillins (methicillin, dicloxacillin, nafcillin, oxacillin, etc.) and the cephalosporins.

►mecA gene is responsible for resistance to Methicillin and other β-lactam antibiotics.

►Troublesome in hospitals and nursing homes

►Patients with open wounds, invasive devices, and weakened immune systems are at greater risk of infection than the general public.

►Treatment of MRSA:• Vancomycin , teicoplanin are

glycopeptide antibiotics used to treat MRSA infections.• Linezolid is now considered drug of

choice.

Vancomycinintermediate-resistant Staphylococcus aureus (VISA)

►Linezolid,quinupristin/dalfopristin (synercid), daptomycin, and tigecycline are used to treat severe infections that do not respond to glycopeptides such as vancomycin.

THANK YOUFOR

MRSA & PVL

►Hospital Associated MRSA (HA-MRSA)►Community Associated MRSA (CA-

MRSA) Has Panton-Valentine Leucocidin (PVL)

Toxin.

Panton Valentine Leucocidin

Panton-Valentine leukocidin (PVL) is a cytotoxin - one of the β-pore-forming toxins.

The presence of PVL is associated with increased virulence of certain strains (isolates) of Staphylococcus aureus.

Present in the majority of community-associated Methicillin-resistant Staphylococcus aureus (CA-MRSA) isolates.

Cause of necrotic lesions involving the skin or mucosa, including necrotic hemorrhagic pneumonia.

PVL creates pores in the membranes of infected cells. PVL is produced from the genetic material of a bacteriophage that infects Staphylococcus aureus, making it more virulent

The methicillin-resistance gene (mecA) can be passed from one bacterial cell to another as a transposable element (a piece of DNA that inserts itself into the bacterial chromosome). The pvl gene is normally present in the genome of the S. aureus bacteriophage and encodes a

toxin known as the PVL protein. Upon infection, the phage can insert its DNA into the bacterial chromosome transforming a non-toxic bacterial cell into a bacterium capable of producing the PVL toxin. The mecA gene can be acquired by both CA-MRSA and HA-MRSA.

The pvl gene, on the other hand, is found normally in CA-MRSA but not in HA-MRSA.