MDL 251 CLINICAL MICROBIOLOGY
The Gram-Positive Cocci – Part I
Staphylococcus species
We encounter at least 3 clinically important species of staphylococci in the medical laboratory (refer
to chapter 14 for background and description of the various species of Staphylococci)
 Staphylococcus aureus – Usually considered a pathogen when encountered, except when
found in respiratory specimens where it is usually regarded as normal flora (reservoir in the
nasal vestibule)
 Staphylococcus epidermidis – Highly opportunistic, but generally not considered a pathogen
unless found in obvious association with disease (reservoir: skin surface and mucous
membranes), Coagulase negative Staph. (CNS)
 Staphylococcus saprophyticus – Also opportunistic, and considered pathogenic when found
in conjunction with urinary tract infections (UTI)/ pyelonephritis in pregnant women
(reservoir: skin, urogenital tract), Coagulase negative Staph. (CNS)
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Many other species exist that are not further identified in the clinical lab due to lack of clinical significance.
Identification of Staphylococcus
 Blood Agar
 Creamy, white or light gold; buttery-looking
 May be beta hemolytic (species dependent)
 Chocolate Agar
 Similar to growth on blood agar (no hemolysis)
 Mannitol Salt Agar - Growth
 MacConkey Agar - NO GROWTH!!
 Selective agars: PEA, CNA, CHROMagar
Staphylococcus species
Identification schema: The catalase and coagulase tests are key for identification of the
staphylococci. Study these on pages 239-241
 Catalase- Production of bubbles when organism mixed with Hydrogen Peroxide
(H2O2). See Page 201 for procedure
 Coagulase- presence of enzyme causes fibrin clot formation in presence of S. aureus.
See Page 203 for procedure.
Staphylococcus aureus
 Reservoir is in the anterior nares; colonizes damaged skin
 Presence usually suggests pathogenicity, except in respiratory cultures
 Resistant to heat, desiccation, high salt concentration and alcohol
 Recovery strategy
 Grows very well on blood agar (BA) and chocolate agar
 Also grows on polyethyl alcohol (PEA) agar
 Also CNA, PEA agar and mannitol salt agar (high salt concentration)
 Growth at 35°C in CO2 in 18- 24 hours.
 Specimens of choice: Blood, Urine, Wounds
 Acquired resistance to methicillin – MRSA
 Methicillin resistant Staph aureus (MRSA) increasing in frequency, due to highly
prescribing methicillin and other penicillins in 1960’s.
 2005- 60% of S. aureus isolates were MRSA, some resistant to up to 20 different
antibiotics
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 May use CHROMagar
 Staph. aureus
 Identification strategy
 Recover large white colonies from BA (often beta hemolytic)
 Gram stain= gram positive cocci in clusters
 Catalase test – positive (streptococci are negative)
 Coagulase test – positive for S. aureus; negative for S epidermidis and other CNS
 If slide test for bound coagulase is negative, must then perform a tube test for
free (unbound) coagulase
 Produces a variety of enzymes and bioactive proteins
 Presence of these may determine pathogenicity and virulence
 Tend to vary among different strains
S. aureus - produces localized suppurative (pus-forming) infections facilitated by a variety of
enzymes and bioactive substances (see examples below)
 Protein A
 Major antigenic component, located in cell wall of S. aureus
 Has anti-phagocytic and anti-complementary properties
 Lowers host resistance to other organisms
 Coagulase
 Produces a fibrin clot at site of infection, trapping additional organisms
 Neutralizes bactericidal effects of normal serum
 Alpha toxin and hemolysins
 Breaks down RBCs
 Inactivates leukocytes
 Increases tissue cell permeability
 May be dermonecrotic
 Polysaccharide capsule – slime layer, biofilm
 Toxins
 Leukocidins and hemolysins inactivate antiphagocytic cells
 Enterotoxins cause disruption in Na/K pump in bowel producing diarrhea and
vomitting
 Exfoliative toxin causes skin depolymerization and peeling
 TSST-1- Toxic Shock Syndrome Toxin
 Aggressins – a class of enzymes which collectively act on host tissue to establish bacterial
infections and relate to virulence
 Hyaluronidase – loosens tissue via depolymerization; related to septicemia
 Lipase – acts on oils and fats in skin; related to furuncles and carbuncles
 DNAse – degrades host DNA
 Staphylokinase – desolves fibrin clots; inactivates plasminogen (may be utilized as an
anti-clotting medication)
 Biofilms
Diseases produced by S. aureus (see p. 234-235)
 Toxic Shock Syndrome (TSS) TSST-1- Toxic Shock Syndrome Toxin- produced by
organism
 Causes fever, vomiting, diarrhea, desquamation, hypotension
 May be life-threatening via progression to uncontrolled shock and sepsis, death
 Furruncles Boils; limited subcutaneous involvement
 Carbuncles- Focus of deep-seated abscess, often with multiple drainage sites
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 Associated with genetic predisposition and/or immune compromise (George
Washington suffered endless misery from these)
 Common in diabetic patients
 Scalded Skin Syndrome (Ritter’s Disease)
Mostly in children/ neonates and facilitated by exfoliative toxins
 May manifest as blister, or desquamation of skin, which may be painful; strain
specific
 Lesions are sterile – cannot recover organism from these; classic intoxication
 MRSA INFECTION
 Bacteremia
 Presence in blood but no reproduction in blood
 May cause endocarditis; fatality is about 50%
 Petechiae and positive blood cultures
 Septicemia
 Bacterial reproduction in blood
 Usually involves neonates or the debilitated elderly
 Osteomyelitis
 From trauma, fractures, bruises; may involve marrow
 Most common in children <12 years of age
 Usually found at ends of long bones or in spinal column
 The Staphylococci (cont.)
 Pyelonephritis
 From kidney stones, previous bacteremias
 Impetigo
 Bullous (blister), exfoliative bacterial dermatitis
 Usually nosocomial in origin
 Produces septic lesions; do not confuse with scalded skin syndrome
 Food Poisoning
 From enterotoxins; produces sudden onset of nausea, diarrhea and vomiting; NO
fever
 Types A and B enterotoxins
 Common 2-6 hours after picnics (eggs, seafood, pastries)
 Other Staphylococcal infections
S. epidermidis, S.haemolyticus and S.lugdunensis infections:
Associated with medically implanted devices, nosocomial spread of resistant isolates,
opportunistic
S. saprophyticus infection
UTI-females
Micrococcus species
 Gram positive cocci in clusters
 Colonies: small, opaque, pigmented (yellow), nonhemolytic
 Catalase=positive
 Coagulase=negative
 Microdase=positive (see p. 215)
 Usually a contaminant. Rarely implicated in infections
 Treatment
 Resistance to every therapeutically useful antimicrobial has been described.
 Detailed study of specific antimicrobials and their mechanisms will be explored in Micro 2.
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Key Terms
Catalase - Bacterial enzyme that breaks down peroxides with liberation of free oxygen
Coagulase – protein enzyme that enable conversion of fibrinogen to fibrin, used to differentiate S.
aureus
Microdase – Modified oxidase test used to differentiate Micrococcus spp. from catalase positive
staph spp.
Novobiocin – differentiates S. saprophyticus
MRSA – Methicillin-resistant S. aureus
TSST – Toxic shock syndrome toxin, S. aureus
Hyaluronidase - It is often speculated that Streptococcus and Staphylococcus pathogens use
hyaluronidase as a virulence factor to destroy the polysaccharide that holds animal cells together,
making it easier for the pathogen to spread through the tissues of the host organism.
Exotoxins – toxin secreted by bacteria
Enterotoxins – protein exotoxin released by a microorganism that targets the intestines
Aggressins – Any substance produced in the body by a pathogenic bacterium that enhances the
virulence of the bacteria
Scalded Skin syndrome - The syndrome is caused by S. aureus and causes detachment within the
epidermal layer.
Impetigo – highly contagious staph infection (aureus or less commonly pyogenes), causing lesions
and scabbing often seen on arms, legs, and face.
Furuncles – another S. aureus infection resulting in boils at a hair follicle
The Gram-Positive Cocci – Part II
Streptococcus and Enterococcus species
Streptococcus species
Streptococci are ubiquitous in clinical specimens, and are notorious opportunistic pathogens. Scan
chapter 15, but do not become intimidated with the myriad of details presented in this chapter. We
will begin by focusing on
Four clinically relevant streptococci and their close cousins the enterococci:
 Streptococcus pyogenes (Group A strep; Beta-hemolytic strep)
 Streptococcus agalactiae (Group B strep; Beta-hemolytic strep)
 Streptococus pneumoniae (Alpha-hemolytic strep; the pneumococcus)
 Streptococcus - Group D (Alpha or non-hemolytic strep)
 Enterococcus faecalis (Alpha,non-hemolytic or beta)
Description
 Partial lysis of red cells around colony; greenish discoloration around colony
 Complete lysis of red cells around colony; clear area around colony
 No lysis of red cells around colony; no change in agar
 Small area of intact red cells around colony surrounded by a wider zone of complete
hemolysis
 Identification schema: The catalase test separates streptococci (catalase negative) from the
staphylococci (catalase positive)
 Hemolysis: The more pathogenic species often display beta-hemolysis (see page 87, Photos
p. 94)
 Other biochemical tests include: growth in 6.5% NaCl (p. 226) and growth in bile-esculin
media (p. 199 )
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Most strains of streptococci are fastidious and have particular nutritional requirements. Identical
strains may cause more than one disease, while a single disease may be caused by immunologically
different strains. Also, one antigenic type may show association with more than one single infection
 Streptococci may be classified on the basis of
 Hemolysis (alpha= green, beta= clear, gamma= none)
 Biochemical reactions
 Ability to tolerate NaCl and/or bile salts
 Sensitivity to Optochin & Vancomycin
 Lancefield groupings
 Immunologically (ASO, Anti-DNase, Anti-streptokinase, anti-hyaluronidase)
 Source of recovery
 Streptococcus species (cont.)
 Most can be grouped according to Lancefield’s scheme (except pneumococci) and certain
other non-pathogenic strains); Grouped according to carbohydrates (C-substances) in cell
wall. Of the Beta Hemolytic strains
 Group A – S. pyogenes (anginosus group
 Group B – S. agalactiae
 Group C – opportunistic pathogens found in humans and horses
 Group D – alimentary canal of man and animals (usually non hemolytic)
Note: Enterococci – originally members of the Group D strep; now a separate genus- may be alpha,
beta or non-hemolytic
 Many species of alpha or non-hemolytic streptococci inhabit the normal skin, alimentary
canal, mucous membranes and oral region; presence does not necessarily imply infection
 Most beta-streptococci do not usually occupy healthy tissue; presence almost always
suggests pathogenicity
Streptococcus pyogenes
(Group A streptococci) Identification
 Beta-hemolytic and causes acute human infections; most beta-hemolytic clinical isolates will
be S. pyogenes; one of most aggressive pathogens encountered.
 PYR test – positive ( pg 225)
 Often identified serologically (Lancefield precipitin test originally, now latex
agglutination or coagulation) rather than biochemically (see page 136 and 258)
 Presence always suggests pathogenicity
 Relatively unresponsive to most biochemicals; sensitive to vancomycin
 Specimens of choice: Wounds, Respiratory, Urine
 Streptococcus pyogenes (cont.)
Many strains produce a variety of virulence factors (see p. 250)
 M-protein – a major virulence factor
 Shared by all strains of S. pyogenes
 May be lost if repeatedly subcultured in lab
 Has anti-phagocytic properties via hyaluronidase in their capsules
 Facilitates attachment to tissues
 Hemolysins –
 Streptolysin S: Not produced by all strains; produces hemolysis of RBCs and is
leukocidal; non-antigenic
 Streptolysin O: Damaged by oxygen and is reversibly deactivated; is cardiotoxic and
leukocidal; is antigenic; basis for ASO test used to detect rheumatic fever
 Streptokinase 5
 Fibrinolysin; destruction of fibrin clots
 Plasmin deactivator
 May be administered medically- used as “clot buster” for strokes/ MIs
 Erythrogenic toxin – Streptococcal pyrogenic exotaxin (SPE)
 Scarlet fever toxin, result of Streptococcal pharyngitis: causes rash of face & upper
trunk. Toxin is immunogenic and immunity ensues against the toxin, not the
organism. Also Toxic Shock Syndrome- renal & respiratory failure, rash & diarrhea.
S. pyogenes (cont.) – Disorders produced by Group A streptococci include watery, spreading lesions
(suppurative) which are highly communicable and infectious
 Scarlet fever –
Pharyngeal involvement with rash on face & upper trunk
 Necrotizing fasciitis and myositis – “flesh-eating bacteria”
 Acute Pharyngitis –
 Classic sore throat in children; may spread to other organ systems if not treated
 Most classic bacteria causing sore throats, skin infections. Also “flesh-eating
bacteria”- deeper tissue & organ involvement
 Perinasal sinusitis –
 From unresolved pharyngitis
 Also leads to otitis media, mastoiditis (ear canals), cervical adenitis (inflamed lymph
nodes)
 Erysipelas – localized inflammation of skin & hot, red lesions on face & head
 Diffuse pyoderma produced by streptococcal toxin
 Impetigo –
 Diffuse maculopapular rash; staphylococci may be a predisposing factor
 Rheumatic fever – post-streptococcal disease
 Follows unresolved pharyngitis and/or scarlet fever
 Detected by ASO test
 Glomerulonephritis – post stretococcal disease
 Follows unresolved pharyngitis
 Deposition of Ag-Ab complexes in kidney resulting in loss of renal function
Streptococcus agalactiae
Often referred to as Group B streptococci, and are the second of two frequently
encountered beta-hemolytic streptococci (the first was S. pyogenes);
see pp 248-251 for a discussion of Group B streptococci
 S. agalactiae is a frequent cause of neonatal sepsis and may be isolated from the normal
vagina; its presence is always assumed to denote pathogenesis, but otherwise may be part of
the normal vaginal flora in the non-pregnant female
 S. agalactiae (along with E. coli and Listeria monocytogenes) is the most frequent cause of
fatal sepsis and meningitis in the newborn infant
 Prospective mothers are routinely tested for S. agalactiae as a precaution;
 Unfortunately pregnant women may carry the organism, but be culture-negative
 May also cause rare cases of UTI, pneumonia, endocarditis, arthritis, osteomyelitis and other
skin & soft tissue infections in immunocompromised patients
 Capsular material interferes with phagocytic activity and complement cascade activation
 Specimen of choice – vaginal; blood; CSF
 Streptococcus agalactiae (cont.)
 Beta hemolytic, Morphologically similar to Group A streptococci. Usually a smaller beta
zone. PYR test = negative
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 Demonstrates at lease three different types of carbohydrate envelope antigens – I, II and III,
and may be identified on the basis of serological tests (latex agglutination testing)
 Also may be differentiated from Group A streptococci on the basis of the CAMP test
(Christie, Atkins, Munch-Peterson), which is an old test, but still performed in certain smaller
laboratories (see p. 200):
 Latex Agglutination testing for
Lancefield grouping of beta hemolytic Streptococcus
 Other Beta Hemolytic Streptococci
 Groups C, F and G
 Identified serologically (Lancefield precipitin test originally, now latex agglutination or
coagulation) rather than biochemically
 Recovered from wounds, respiratory ,blood.
Streptococcus pneumoniae
Streptococcus pneumoniae is often referred to as one of the viridans streptococci
due to its production of alpha hemolysis, which denotes an incomplete clearing of
red blood cells when cultured on blood agar; also referred to as the pneumococcus
 A causative agent of subacute bacterial endocarditis (SBE) and/or septicemia and
frequently gains entrance to the bloodstream from virtually any portal, especially the
nasopharyngeal region where it may be regarded as normal flora. Common following
rheumatic fever (valves damaged), or in abnormal heart.
 Specimen of choice is blood
 Causative agent of bacterial (lobar) pneumonia, occurring as a sequellum to previous viral
infection – especially in alcoholics, smokers, and debilitated
 Specimen of choice is deep cough and/or transtracheal aspirate
 Pneumococcal meningitis – may initially involve middle ear (otitis media); also from
pharyngitis, sinusitis
 CSF is specimen of choice; sensitive to cold, therefore keep specimen warm
 Specimens may be collected from other opportunistic sites/infections
 Gram stain appears as gram positive cocci in pairs (lancet shaped)
 Colonies tend to dip down in the center and resemble a doughnut (umbilicated) as they age
 Colonies may be mucoid if polysaccharide capsule present (Quellung test- determine
serologic testing)
 Auto-hemolysis produces characteristic colonial appearance on blood agar, resembling
volcano - concave centers
 ID based on optochin( pg. 222) and bile solubility (pg. 199) tests
 Sensitive to vancomycin
 Vaccine available to prevent infection from S. pneumoniae (recommended for elderly,
aspleenic, sickle cell, diabetes, HIV, immunocompromised)
Enterococcus and Group D Streptococcus
Composed of a large number of saprophytes and opportunistic pathogens; their presence in clinical
specimens does not necessarily suggest pathology (except blood). May be either alpha, or nonhemolytic .
Enterococcus spp. may be alpha, beta or gamma
 Enterococcus may be separated from Group D streptococci, and these from other
Streptococcus spp. (viridans ) on the basis of bile-esculin hydrolysis, and growth in 6.5%
NaCl broth.
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Enterococcus testing
Salt tolerance (pg 200)
Bile Esculin (pg 226)
Enterococcus and Grp D Strep
Responsible for UTI (specimen – urine); or SBE (specimen – blood)
Most strains considered normal flora in respiratory and alimentary canal
Enterococci were once counted among the Group D streptococci; now comprises the separate
genus Enterococcus (E. faecalis and E.faecium)
 Enterococcus spp. have acquired resistance to many commonly used antibiotics
 Enterococcus Vancomycin resistance has become more common --VRE
Key Terms
Hemolysis – visible breakdown of red blood cells on blood agar
Pyr test – Pyrrolidonyl arylamindase test. S. lugdunensis, S. pyogenes, and Enterococci spp are
positive.
Lancefield groups – groups beta hemolytic bacteria based on carbohydrate composition of
bacterial antigens found on cell walls.
Camp test – used to identify beta hemolytic streptococci.
Bile esculin – used to identify Enterococci and Group D Strep.
Salt Tolerance – used to identify Enterococci
Optochin – used to identify S. pneumoniae
Bile Solubility – used to identify/confirm S. pneumonia
ASO - Anti-streptolysin O (ASO or ASLO) is the antibody made against streptolysin O, an
immunogenic, oxygen-labile hemolytic toxin produced by most strains of group A and many
strains of groups C and G streptococci. The main function of streptolysin O is to cause betahemolysis.
Streptokinase – enzyme secreted by several species of streptococci that activate human
plasminogen
SPE – exotoxins produced by S. pyogenes
Scarlet Fever – infection of S. pyogenes from erythrogenic toxin
Rheumatic fever – inflammatory disease post S. pyogenes infection.
Neonatal sepsis – systemic bacterial infection of newborn after exposure to flora of birth canal
VRE – Vancomycin-resistant Enterococci
Gram-Positive Rods: Bacillus, Listeria, Corynebacterium and Related
Organisms
The gram-positive rods are comprised of many genera, some of which are
aerobic spore-forming, and non spore-forming; many of which
produce serious disease . Several genera will be covered here, others in subsequent lectures.
 Bacillus species: Aerobic spore-forming rods (see Ch. 16)
 Listeria monocytogenes: Aerobic non spore-formers (see Ch. 17)
 Corynebacterium species and related organisms: Aerobic non spore-formers (see Ch. 17)
 Eripselothrix, Lactobacillus and Gardnerella : Aerobic (Ch. 18)
You are only responsible for the chapter sections that are related to the above organisms
Bacillus species
At least three B. species are encountered clinically: B. cereus, B. subtilis, and
 B. anthracis. Produce spores which are metabolically inactive and resistant to heat & chemicals.
Are aerobic or facultatively anaerobic, grow on sheep BA, and are large gram positive or gram
variable rods, with spores appearing as holes inside rods. Specific causative agent of anthrax
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 B. cereus: Recovered from soil and grains; Causative agent of food poisoning (intoxication)
producing a self-limiting, non-fatal nausea and vomiting, similar to that caused by S. aureus
Usually associated with partially cooked fried or boiled rice
May cause septicemia in drug abusers and/or those on hemodialysis
 B. subtilis: Recovered from soil: Rare opportunistic causative agent of wound and eye
infections; frequently overlooked and/or under-reported
Bacillus anthracis
Large boxy gram-positive spore-forming rods with subterminal endospores; forms large,
nonhemoltyic medusa-head colonies on BAP. Sporulates under aerobic conditions.
Production of capsules which impart anti-phagocytic properties
Spores formed only in air; highly resistant; ingestion/inhalation and germination
Forms “strings of pearls” (chains of spherical bacilli) when incubated for several hrs on agar
containing low concentration of penicillin
Lab personnel must work under Biosafety Level 3 conditions
Pathology
Pathogenicity due to overwhelming septicemia, capillary damage and death
Produces toxin composed of multiple components
Multiple clinical manifestations occur in man
Cutaneous anthrax – formation of papule to slack scar (Black Eschar) on skin at site of spore
penetration; usually from contaminated hides, wool or hair; 20% mortality if untreated (from
toxemia)
Characterized by generalized hemorrhage and edema
Rarely progresses to meningitis and/or pneumonia
Pulmonary anthrax (Wool Sorter’s Disease): Severe respiratory infection with nearly 100%
mortality whether treated or not; begins with malaise & mild fever & nonproductive cough,
progressing to respiratory distress, massive chest edema, cyanosis & death
Very rapid dissemination, initially producing bronchopneumonia
Severe mediastinitis (area separating the lungs); involving many lymph nodes
Hemothorax, septicemia and/or meningitis
Rapidly fatal multiple organ failure
Gastrointestinal anthrax: Rare form of Wool Sorter’s Disease produced by eating raw meat
and/or massive numbers of spores
Ingestion of spores; affects either oropharyngeal or abdominal area
Most patients die from toxemia & overwhelming sepsis.
Bioterrorism worry- Post Office scare of 2001-2002. Most likely bioterroristic organism to
use for agents of mass destruction.
The Corynebacteria
Loose knit group of irregularly staining gram-positive non-spore-forming pleomorphic rods, many
with fastidious growth requirements; most do not cause disease and are opportunistic pathogens.
“Coryne” is Greek for club- club-shaped rods. Gram positive short or slightly curved rods with
rounded ends- some with rudimentary branching- some in parallel rows (palisades) or resembling
Chinese letters (V, L & Y forms)
Main pathogen: C. diphtheriae
Other Corynebacteria may produce a variety of diseases, generally associated with bacteremias,
septicemias, endocarditis and/or meningitis
The term “diphtheroids” is used to describe non pathogenic corynebacterium (not diphtheriae)
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Corynebacterium diphtheriae
(Klebs- Loeffler’s bacillus)
Causative agent of diphtheria – leathery throat lesion (pseudomembranous “diphth”)
Man is sole habitat; mucous membrane of nose and throat; skin & vagina. Some are asymptomatic
carriers.
Pathology
Represents a true intoxication; toxin only is disseminated, acting on PNS and heart
Toxigenic strains infected by specific bacteriophage that carries toxic gene
Incubation 1-7 days; low fever, pallor, tachycardia, weakness
Produces a fibrinous exudate and necrotic tissue; may erode epiglottis
Can extend to larynx and close off trachea
Peripheral vascular collapse and death
Recovery does not impart immunity
Sequellae include myocarditis and chronic peripheral neuritis
Also Cutaneous form of diphtheria- nonhealing ulcers with membrane formation
Corynebacterium diphtheriae
Culture for C. diphtheriae is on:
Tinsdale’s agar (black colonies with dark brown halo)
Cystine-tellurite BA (black or gray colonies)
Also Loeffler’s medium- contains serum & egg- shows metachromatic granules in cells
Testing for diphtheria:
Biochemical testing, whole-cell fatty acid analyses, cell wall diamino acid analysis or 16S rRNA
gene sequencing- reference labs only
Toxin testing for diphtheria using guinea pig lethality testing, immunodiffusion testing using Elek
plate, tissue culture cell test, and PCR test
Treatment: antitoxin to neutralize diphtheria toxin plus penicillin or erythromycin to kill organism
About one person in 10 who gets diphtheria dies of it. Diphtheria is more severe for those under 5
and over 40 years of age.
Most cases have occurred in nonimmunized (or inadequately immunized) individuals.
Vaccine: multidose diphtheria toxoid (inactivated)- begun in infancy, boosters every 10 years
(Diphtheria & Tetanus)
Exposed persons: treat with single dose IM penicillin or 7-10 day course oral; follow-up throat
cultures at 2 weeks (repeat 10 day if positive); also be revaccinated.
Listeria monocytogenes
Causes profound neonatal sepsis and death; primarily disease of persons under 1 month, and over 40
yrs of age; organism easily recovered from soil, widely distributed in nature, occasionally found in
human GI tract.
Dangerous due the ability to survive inside phagocytes due to formation of Listeriolysin toxin; short
gram positive rod occurring singly or in short chains resembling streptococci
Infection by ingestion of contaminated food, such as meat and dairy products. Also colonized
mothers pass organism onto fetus. Entry usually from GI tract to blood, and sometimes, to
meninges.
A peculiar property that affects its food-borne transmission is the ability to multiply at low
temperatures. The bacteria may therefore grow and accumulate in contaminated food stored in the
refrigerator. So it is not surprising that listeriosis is usually associated with ingestion of milk, meat or
vegetable products that have been held at refrigeration temperatures for a long period of time.
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Initial symptoms may include flu-like symptoms, backache, vomiting diarrhea, headache.
Meningitis accounts for 75% of cases of Listerial infections; other clinical manifestations exist .
Often complicated by encephalitis, a pathology that is unusual for bacterial infections .
When listeria meningitis occurs, the overall mortality may be as high as 70%; from septicaemia
50%, from perinatal/neonatal infections greater than 80% in the fetus.
CNS listeriosis in pregnant mothers and otherwise healthy individuals may be asymptomatic
Septic military granulomatosis causes cardiorespiratory distress in military recruits
Vomiting, diarrhea, dark red skin papules, hepatosplenomegaly
Abscesses of liver, spleen, adrenals, lungs, pharynx, GI, CNS
Oculoglandular listeriosis
Severe conjunctivitis, corneal ulceration
May occur at any age
Most susceptible: elderly, immunocompromised or if underlying disease is present; (transplant
patients, diabetics, Hodgkin’s disease)
ISOLATION
Clinical specimens: CSF, blood, placenta, fetal tissue, stool
Good growth on 5% BAP (beta hemolytic)
Oxford media ( selective) black colonies – for non-sterile sources
IDENTIFICATION
Small, Gram-positive rods, which are sometimes arranged in short chains. In direct smears they
may be coccoid, so they can be mistaken for streptococci. Longer cells may resemble
corynebacteria.
Presumptive ID by observing motility by direct mount (end-over-end tumbling) in nutrient broth at
RT for 1-2 hrs. Alternate is umbrella-shaped pattern from overnight RT incubation in tube of semisolid media.
Catalase (+) , esculin( +)
Immunocompromised & pregnant women should avoid eating soft cheeses such as Mexican-style,
brie, feta, Camembert, & blue-veined cheese, and reheat hotdogs & cold cuts before eating
PCR assay available for testing food proucts
Ampicillin & penicillin are treatment options
Listeria species
Erysipelothrix rhusiopathiae
Carried in the intestinal tract of turkeys and swine ( also rabbits, sheep, birds)
Causes erysipeloid, localized cellulitis, following a bite in an individual handling animals or animal
products.
Butchers and veterinarians
May become disseminated and lead to endocarditis
Identification
-short gram pos rod
-growth on BAP
-catalase (-), H2S (+)
-commercial systems (API)
Penicillin is the treatment of choice
Lactobacillus
Normal flora of human mouth, GI tract and female genital tract
Usually a contaminant. May cause infections in the immunocompromised.
Gram stain: gram pos rods in chains (fig 18-1)
(called an aerotolerant anaerobe)
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Grows well on BAP(alpha), Choc, CNA
Cat.= neg
Mac=NG
Gardnerella
Normal vaginal flora
Causes Bacterial vaginosis, UTI
Growth on BAP,Choc,CNA, HBT (beta) (fig 18-2)
Gram stain: pleomorphic Gram variable or gram neg rods and coccobacilli
“clue cells” on saline wet mount or gram stain
Clue cells- stipled epithelial cells, indicating infection
1. Which simple test will separate Staphylococci from Streptococci?
Catalase positive organisms are Staphylococci sp., while negative would be Streptococci sp.
2. Which tests will separate S.aureus from all "coagulase negative staphylococci" like S.
epidermidis?
Coagulase. S. aureus is positive.
3. How can you differentiate Micrococcus from coagulase negative staphylococci like S.
epidermidis?
Micrococci are Microdase positive while Staphylococci are negative.
4. Staphylococci are considered resistant to Novobiocin if the zone is less than 16mm.
5. Which organism may give a pseudocatalase positive result?
Enterococci
6. Why is it useful to perform both a slide and tube coagulase?
Staphylococcus aureus contains both bound and free coagulase. The slide test confirms
bound coagulase and the tube test is for free coagulase. Negative slide tests should be confirmed
with the tube test.
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