MIDTERMS C2 - Catalase-Positive, Gram-Negative Cocci Flashcards by Mark James Mateo

All Cocci are Gram-Positive except

Veilonella, Neisseria, Moraxella

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Beta-Hemolytic Streptococci

Streptococcus pyogenes
Streptococcus agalactiae

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Viridans streptococci

Streptococcus mutans
Streptococcus salivarius
Streptococcus mitis
Streptococcus bovis
Streptococcus urinalis
Streptococcus anginosus
(Streptococcus milleri )

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Nutritionally variant streptococci

Abiotrophia defective
Granulicatella adiacens
Granulicatella balaenopterae
Granulicatella elegans

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Enterococci

Enterococcus faecalis
Enterococcus faecium
E. durans
E. mundtii
E. dispar
E. gallinarum
E. avium
E. hirae
E. raffinosus
E. casseliflavus

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6 Clinically Significant Streptococci and Enterococci

Streptococcus pyogenes
Streptococcus agalactiae
- Streptococcus pneumoniae
- Viridans streptococci
Enterococcus faecalis
Enterococcus faecium

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Other Catalase-Negative, Gram-Positive Cocci

Leuconostoc spp.
Lactococcus spp.
Globicatella sp.
Pediococcus spp.
Aerococcus spp.
Gemella spp.
Helcococcus sp.
Alloiococcous otitidis
Dolosicoccus paucivorans
Facklamia
Dolosigranulum pigrum
Ignavigranum ruoffiae
Tetragenococcus

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True Pathogen Streptococci

Streptococcus pyogenes, not part of normal microbiota

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Colonizer Streptococci

Streptococcus pneumoniae

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Alloiococcous otitidis Result

(+) in Catalase Test BAP (Blood Containing Media)
(-) In Catalase Test CAP (Whole Blood, lysed rbc)

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belong to the family Streptococcaceae
spherical to ovoid, catalase(-), gram (+) cocci arranged in pairs or chains when grown in liquid media

Streptococci (no copious bubble in addition of hydrogen peroxide kasi catalase negative)

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What is the color in gram stain of the Streptococci and Enterococci?

purple or blue

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non-spore-former and generally
non-motile except for the rare
motile strains of group D streptococci

Streptococci

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Streptococci are _____________ but some strains require added CO2 (***microaerophilic strains = Viridans streptococci)

facultative anaerobes

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generally non-encapsulated except for some strains of groups A, B, C andD
commonly found as part of normal human flora
growth enhanced by blood (BAP), serum or glucose

Streptococci

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Streptococci result in BAP

grayish, pinpoint, circular, and translucent to slightly opaque colonies while some have mucoid colonies

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Biochemical Test Result of Streptococci

(-) catalase, oxidase and gas production

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Classification of Streptococci based on temperature requirement

Academic or Bergey’s Classification

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neither grow on 10 C nor 45 C, grow only at 37 degree Celsius
produce pus; mostly β-hemolytic

Pyogenic Group

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Species of Pyogenic Group

S. pyogenes, group C and G streptococci (large
colony forming isolates)

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grow both at 45 oC and 37oC but not at 10oC
not part of the Lancefield group: resist Lancefield Precipitation Test

Viridans Group

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α-hemolytic or non-hemolytic
normal biota in URT in humans
some may have A, C, G, or N antigen

Viridans Group

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Species of Viridans Group

S. salivarius, S. mutans (dental plaque), S. mitis, S. sanguis, S. anginosus

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grow on 10 C and 37C but not at 45C
non-hemolytic with Lancefield N antigen
found in dairy products

Lactic Group

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Species of Lactic Group

S. lactis  normal coagulation and souring of milk

grow at 10C, 45C , and 37C; can withstand temperature above 60C normal of human intestine

Enterococcus Group

Species of Enterococcus Group

E. faecalis (part of normal fecal flora)

Classification based on hemolytic patterns on BAP

Smith & Brown's Classification

partial/incomplete hemolysis of RBC around colonies

Alpha-hemolytic Streptococci

 Culture: greenish or brownish discoloration around colony

Alpha-Hemolytic Streptococci

Species of Alpha-Hemolytic Streptococci

S. pneumoniae (green streptococci), some Enterococci spp., and Streptococcus bovis complex spp.

complete hemolysis of RBC around colony

Beta-Hemolytic Streptococci

 Culture: clear zone around colony

Beta-Hemolytic Streptococci

Species of Beta-Hemolytic Streptococci

S. pyogenes, S. agalactiae, S. dysagalactiae subsp. equisimilis and S. anginosus group and some enterococci spp

exhibit NO hemolysis of RBC around colony

Gamma-Hemolytic Streptococci

 Culture: RBC surrounding colony are not affected (no change)

Gamma-Hemolytic Streptococci

Species of Gamma-Hemolytic Streptococci

Enterococci and Streptococcus bovis complex

small area of intact RBCs around colony surrounded by a wider zone of complete hemolysis

Alpha-prime (α′) or wide zone

Hemolysis is enhanced by

stabbing the inoculating loop into the agar several times

Plates are always examined for hemolysis by

holding them in front of light source

Rebecca Lancefield based on antigenic characteristics of GROUP-SPECIFIC C SUBSTANCE, a cell wall polysaccharide

Lancefield Classification

extraction of C carbohydrate from the streptococcal cell wall by dilute acid and heating the suspension for ___ mins

10 mins

mostly significant in classifying and identifying beta hemolytic streptococci

Lancefield Classification

not considered as a part of indigenous flora = pathogenic acquired through contaminated droplets released through coughing and sneezing

Group A Streptococci: Streptococcus pyogenes

resistant to drying and can be recovered from swabs several hours after the collection colonizes throat and skin on humans

Streptococcus pyogenes

major virulence factor = resist phagocytosis and adherence of the bacterial cell to mucosal cells attached to peptidoglycan of cell wall and extends to cell surface

M Protein

most common in pharyngitis

M1 serotype

mediates post-streptococcal diseases: Rheumatic fever = Acute glomerulonephritis =

RF - class 1 M AG - class I or II

bind beta globulin factor H = regulatory protein involved in the degradation of C3b binds to fibrinogen blocking complement alternate pathway activation

M Protein

aka FIBRONECTIN-BINDING PROTEIN adhesion molecule that mediates epithelial cell attachment

Protein F

adhesion molecule present in the cell wall that is responsible for the adherence into respiratory epithelial cells

Lipoteichoic Acid

weakly immunogenic prevents opsonized phagocytosis by PMN or macrophages mask bacterial antigens

Hyaluronic Acid Capsule

causes lysis of fibrin clots binds plasminogen and activates the production of plasmin allows bacteria to move from clotted area (spread infecion)

Streptokinase

Streptokinase Application

treatment of pulmonary emboli, coronary artery, and venous thromboses

Types of Hemolysins

Streptolysin O Streptolysin S

oxygen labile; highly antigenic = induce antibody response responsible for subsurface hemolysis on BAP incubated anaerobically

Streptolysin O (SLO)

causes lysis of RBC, WBC, platelets, tissue cells inhibited by the cholesterol in skin lipids  absence of protective antibodies associated with skin infection

Streptolysin O

Serologic Test for SLO

Anti-Streptolysin O (ASO) Test

oxygen stable; nonimmunogenic surface hemolysis seen around colonies that have been incubated aerobically

Streptolysin S

causes lysis of RBC, WBC and platelets in the presence of room air inhibited by nonspecific inhibitor that is frequently present in sera of humans and animals

Streptolysin S

Oxygen Labile Antigenic (ASO Titer) Subsurface Hemolysis Lyse RBC, WBC, platelets, tissue cells

Streptolysin O

Oxygen Stable Nonimmunogenic Surface Hemolysis Lyse RBC, WBC, and Platelets

Streptolysin S

aka deoxyribonuclease) STREPTODORNASE (streptococcal lowers viscosity of exudates, giving pathogens more mobility

Deoxyribonuclease (Dnase)

aka SPREADING FACTOR solubilizes the ground substance of mammalian connective tissues (hyaluronic acid) antigenic and specific for each bacterial or tissue source

Hyaluronidase

serine protease capable of inactivating the chemotactic factor for neutrophils and monocytes (C5a)

C5a Peptidase

formerly called ERYTHROGENIC TOXINS cause ared spreading rash = SCARLET FEVER

Streptococcal Pyrogenic Exotoxins (SPEs)

heat labile and rarely found in group C and G act as SUPERANTIGENS helper cell

Streptococcal Pyrogenic Exotoxins

SPEs is associated with

STREPTOCOCCAL TOXIC SHOCK SYNDROME

Four Exotoxin Types:

SpeA, SpeB, SpeC, and SpeF

degrades protein mediates rashes that are caused by scarlet fever

Exotoxin B (Cysteine protease)

highly virulent strains can cause sharp outbreaks of sore throats and scarlet fever in schools and camps infants and small children = tendency to extend to the middle ear and mastoid

BACTERIAL PHARYNGITIS OR TONSILLITIS (STREP THROAT)

localized skin disease that begins as small vesicles that progress to weeping lesions that crust over after several days usually seen in young children (2 to 5 years)

Impetigo

acute spreading skin lesion that is intensely erythematous with a plainly demarcated but irregular edge rare infection of the skin and subcutaneous tissues observed frequently in elderly patients

Erypsipelas

diffuse, spreading infection of subcutaneous skin tissue characterized by defined area of redness (erythema) and the accumulation of fluid (edema) follows infection associated with mild trauma, burns, wounds, or surgical incisions

Cellulitis

Cellulitis may lead to

gangrene

Cellulitis is differentiated from erysipelas by two clinical findings:

lesion is not raised line between the involved and uninvolved tissue is indistinct

aka GALLOPING GANGRENE, FLESH-EATING BACTERIA SYNDROME, SUPPURATIVE FASCIITIS, HOSPITAL GANGRENE, or NECROTIZING ERYSIPELAS

NECROTIZING FASCIITIS

invasive infection characterized by rapidly progressing inflammation and necrosis of the skin, subcutaneous fat, and fascia

Necrotizing Fasciitis

acts as a superantigen, causing the immune system to contribute to the damage

Exotoxin A

Type 1 NF

polymicrobial infection from which aerobic and anaerobic bacteria are recovered

Type 2 NF

GAS

Type 3 NF

gas gangrene or clostridial myonecrosis

characterized by a precipitous drop in blood pressure, failure of multiple organs, and a very high fever caused by an invasive strep A that produces one or more of the streptococcal pyrogenic exotoxins

Streptococcal Toxic Shock Syndrome

initial streptococcal infection includes

pharyngitis, peritonitis, cellulitis, wound infections

play a major role in the pathogenesis of the disease = superantigens

Spe A

most common strains associated with streptococcal TSS

M1 and M3

punctate exanthem overlying a diffuse erythema that appears initially on neck and upper chest, 1 to 2 days following strep throat rash disappears over the next 5 to 7 days and is followed by desquamation

Scarlet Fever (Scarlatina)

communicable and spread by inhalation of infectious respiratory droplets results from a throat infection with a strain of S. pyogenes that carries lysgenic bacteriophage (T12)- cause by release of streptococcal pyrogenic exotoxin

Scarlet Fever (Scarlatina)

Cardinal Signs of Scarlet Fever

diffuse red rash on the upper chest and spreads to the trunk and extremities, and “strawberry-colored” tongue

susceptibility test for scarlet fever

Dick's Test

 Test arm: 0 .1 ml of Dick’s toxin (Eryhtrogenic Toxin)  Control arm: : 0.1 ml of Dick’s toxoid  Read reaction after 24 hours  (+) reaction: Eythema or redness in the test site  Interpretation: Susceptible to scarlet fever

Dick's Test

based on neutralization of erythrogenic toxins when anti-toxin is injected on the skin of patient with scarlet fever diagnose whether rashes of patient is due to scarlet fever or not

Schultz-Charlton Test

 (+) reaction: “BLANCHING PHENOMENON”– fading of the rashes

Schultz-Charlton Test

follows S. pyogenes pharyngitis autoimmune disease characterized by fever and inflammation of the heart, joints, blood vessels, and subcutaneous tissues

Rhematic Fever (Poststreptococcal sequelae)

mediated by antibodies produced against S. pyogenes M protein that cross-react with human heart tissue most serious result: chronic, progressive damage to the heart valves most common cause of permanent heart valve damage in children

Rhematic Fever (Poststreptococcal sequelae)

aka BRIGHT’S DISEASE inflammatory disease of the renal glomeruli

Acute Glomerulonephritis

develops 1–4 weeks after S pyogenes skin infection (pyoderma, impetigo) or respiratory infection deposition of antigen-antibody complexes, possibly involving the streptococcal M protein Type III hypersensitivity reaction

Acute Glomerulonephritis

Specimen for the Laboratory Diagnosis (S. pyogenes)

Pharynx and Tonsillar swabs (Throat Swabs)

Culture Medium for S. pyogenes

Blood Agar Plate

Result of BAP ( S. pyogenes)

colonies are transparent to translucent, convex or domed entire, circular, shiny an surrounded by wide zone of β hemolysis

presumptive identification of S. pyogenes (S) screening for GAS in throat cultures groups C and G are also susceptible

Bacitracin Susceptibility Test/Taxo A (0.02-0.04 U)

Result of Taxo A (S. pyogenes)

Susceptible

Result of Taxo A (S. agalactiae)

Resistant

detects L-pyrrolidonyl arylamidase more specific for S. pyogenes than bacitracin

Pyrrolidonyl-α-Naphthylamide Hydrolysis Test

is the only species of Streptococcus that is PYR positive

S. pyogenes

Other PYR (+)

Enterococcus, Aerococcus, and Gemella

result of (+) PYR

Cherry Red Color

Group A and B streptococci is _______ to SXT

resistant (positive result)

Group C is ________ to SXT

susceptible (negative result)

interfering respiratory microbiota will be inhibited by _____

SXT (Sulfamethoxazole and Trimethoprim Test)

positive result of SXT means

Resistance

Serologic Test for S. pyogenes

ASO Test

reciprocal of the highest dilution demonstrating no hemolysis expressed in TODD units

Titer

 Serum is added with measured amount of SLO reagent and incubated  Reagent RBC are added indicator  Enough antibody is present: SLO neutralized and no hemolysis occurs

ASO TEst

neutralize reagent DNase B, preventing it from depolymerizing DNA

Anti-DNase B Antibodies

DNase measured by its effect on a DNA methyl-green conjugate

Anti-DNase B Testing

GROUP B STREPTOCOCCI

Streptococcus agalactiae

have group B–specific antigen, acid-stable polysaccharide located in cell wall normal flora of female genital tract and lower gastrointestinal tract

Streptococcus agalactiae

MOT of S. agalactiae

Direct Contact

Causes infection of fetus during passage through the colonized birth canal and premature rupture of mother’s membrane

S. agalactiae

important virulence factor of S. agalactiae

Capsule

prevents phagocytosis but is ineffective after opsonization

Capsule

 most significant component of the capsule critical virulence determinant

Sialic Acid

AVIRULENT FACTORS (S. agalactiae)

Hemolysin CAMP factor Neuraminidase Dnase Hyaluronidase Protease

RELATED INFECTIONS AND DISEASES (S. agalactiae)

1. Most common etiologic agent of NEONATAL SEPSIS and MENINGITIS 2. Pneumonia 3. Postpartum Infection ENDOMETRITIS 4. Osteomyelitis 5. UTI 6. Puerperal Infection 7. Endocarditis (Tricuspid Valve Endocarditis) 8. Skin infection 9. Important etiologic agent of bovine mastitis

Neonatal GBS Disease

Early-onset infection (<7 days old)  pneumonia and sepsis Late-onset infection (7 days old- 3 months old) meningitis and sepsis

Specimen for S. agalactiae

Vaginal and Rectal Swab

BAP result of S. agalactiae

grayish white, mucoid, more translucent to opaque, soft, smooth colonies surrounded by smaller zone of β hemolysis

Todd-Hewitt broth with Colistin and Nalidixic acid = TRANSPORT MEDIUM

Lim Broth

Gentamicin and Nalidixic acid containing broth

TransVag broth

SELECTIVE BROTH orange or red pigment (6 hours incubation)

StrepB Carrot Broth

selective agar for vaginal or rectal swabs yellow to orange colonies

Granada Agar

act synergistically with β-hemolysin produced by S . aureus to cause enhanced lysis of RBC

CAPM FACTOR

presumptive identification of GBS:

S. agalactiae positive`

GBS are streaked perpendicular to a streak of S. aureus on ________ agar

sheep blood agar

result if positive in CAMO Test

Arrowhead-shaped hemolysis

place a drop of extracted β-lysin on area of confluent growth of suspected GBS

Rapid Camp Test or Spot Camp TEst

Incubation period for Rapid Camp Test

35° C for 20 minutes

positive result of RAPID CAMP TEST

enhanced hemolysis

S. agalactiae (+) has ______________ that hydrolyzes sodium hippurate to form sodium benzoate and glycine

hippuricase (hippurate hydrolase)

Ammonia + ninhydrin and hydrindantin =

PURPLE-Colored Complex

Coagglutination or Latex Agglutination

Serotyping

Serotyping Treatment

Penicillin

recovered from the upper respiratory tract, vagina and skin also posses M protein

GROUPS C AND G STREPTOCOCCI

are animal pathogens and the main source of STREPTOKINASE

Group C Streptococci

GROUPS CANDGSTREPTOCOCCI Species

S. dysagalactiae subsp. equisimilis S. equi subsp. zooepidemicus

Types of Group C and G Streptococci

1. Large Colony-forming Isolates 2. Small Colony-forming Isolates

isolates with groups A, C, G and L antigens belong to the ____________

pyogenic streptococci

large-colony–forming β-hemolytic isolates with group C and G antigens belong to

S. dysgalactiae subsp. equisimilis

small-colony–forming β-hemolytic isolates with group C and G antigens belong to the

S. anginosus group

all species (S. equi, S. equisimilis, S. zooepidemicus) are β-hemolytic except S. dysagalctiae which may be hemolytic or non-hemolytic differentiated by carbohydrate fermentation

Group C Streptococci

causes disease in horse

S. equi

may cause pharyngitis, puerperal sepsis, endocarditis, bacteremia, osteomyelitis, brain abcess, post-operative wound infection and pneumonia in humans

S. equisimilis

source of streptokinase used in Thrombolytic Therapy

S. equisimilis

 similar types of acute infections described for S. pyogenes and S. agalactiae, but usually involve compromised patients

Group C and G Streptococci

occur in patients with underlying malignancies

Group G streptococci

occasionally have been associated with acute pharyngitis

Group C organisms

ANTIMICROBIAL SUSCEPTIBILITY TESTS (Group C and G Streptococci)

 Group C streptococci  SUCEPTIBLE to Bacitracin and SXT  Group G streptococci may be bacitracin resistant or susceptible

aka α-prime streptococci that lack Lancefield group antigens normal microbiota of the upper respiratory tract, the female genital tract, and the gastrointestinal tract fastidious, with some strains requiring CO2 for growth

VIRIDANS Streptococci

most common cause of SUBACUTE BACTERIAL ENDOCARDITIS (SBE), condition associated with a transient bacteremia

VIRIDANS Streptococci

MOT of Viridans Streptococci

gain access to sterile site; most notably results from dental manipulations

S. mitis group SPECIES

S. mitis S. sanguis S. parasanguis S. gordonii S. cristatus S. infantis S. oralis S. peroris

S. mutans group SPECIES

S. mutans S. sobrinus

S. salivarius group SPECIES

S. salivarius S. vestibularis S. thermophilus

S. bovis group SPECIES

S. equinus S. gallolyticus S. infantarius S.alactolyticus

S. anginosus group SPECIES

S. anginosus S. constellatus S. intermedius

VIRIDANS STREPTOCOCCI can possess Lancefield group A,C, F, G, or N antigen and in some instances may not be groupable

S. anginosus group

VIRIDANS STREPTOCOCCI possess the group D antigen

S. bovis group

identified in some members of anginosus group

Polysaccharide Capsule and Cytolysin

adherence and colonization of these organisms in endocarditis

Extracellular Dextran and Adhesin (cell surface–associated proteins)

Low Virulence in VIRIDANS STREPTOCOCCI

oropharyngeal commensals and opportunistic pathogens

VIRIDANS STREPTOCOCCI RELATED INFECTIONS AND DISEASES

Major etiologic agent of Subacute Bacterial Endocarditis Major etiologic agent of Dental Carries(plaque) S. mutans Fulminant cardiovascular collapse or meningitis Gingivitis Sinusitis Cellulitis and Wound Infection Abscesses, osteomyelitis, and empyema Biliary or Intra-abdominal infections

abscess formation in the oropharynx, brain, and peritoneal cavity

S. anginosus group

S. constellatus subsp. pharyngis causes

pharyngitis

causes bacterial endocarditis in native valves and prosthetic valve infections

S. mitis group

causes bacteremia, endocarditis, and meningitis

S. salivarius

causes bacteremia, septicemia, and endocarditis

S. bovis group

presence has high correlation with gastrointestinal carcinoma

S. gallolyticus sbusp. gallolyticus

primary contributor to dental caries and also associated with bacteremia Most commonly isolated spp. of viridans streptococci

S. mutans

Specimen for Laboratory Diagnosis of Viridans Streptococci

Blood, Gingival Scrappings, Pus Secretions

BAP result of Viridans Streptococci

small and are surrounded by a zone of α-hemolysis; some isolates are β-hemolytic or nonhemolytic

in pure culture or in high concentration = sweet odor of honeysuckle or butterscotch

S. anginosus

peptidase that hydrolyzes peptide bonds adjacent to a free amino group

Leucine Aminopeptidase (LAP) Test

Leucine Aminopeptidase (LAP) Test Substrate: End Product: Reagent: (+) result:

Substrate: Leucine-β-naphthylamide End-product: β-naphthylamine Reagent: Paradimethylaminocinnamaldehyde (DMACA) (+) result: Red color

(+) LAP Bacteria

Viridans Streptococci, Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus pneumoniae, Enterococcus, and Pediococcus

(-) LAP Bacteria

Aerococcus and Leuconostoc spp.

distinguish small-colony–forming β-hemolytic anginosus group containing groups A or C antigens from large-colony–forming pyogenic strains

Voges-Proskauer Test

(+) VP Bacteria

S. anginosus, S. bovis, and S. mutans groups

detects action of β-D-glucuronidase

β-D-Glucuronidase

Result of β-D-Glucuronidase

Large-colony–forming β-hemolytic groups C and G streptococci (+) Small-colony–forming β-hemolytic anginosus group (-)

Note!!! VIRIDANS STREPTOCOCCI

All members are PYR (-) and LAP (+)

Diagnostic test for S. bovis group (Group D Streptococci)

a. Bile Esculin Test b. Salt Tolerance c. PYR Test d. Penicillin Test

Bile Esculin Test Reagent: (+) result

Reagent: Esculin and 40% Bile Salt (+) result = Blackening of the agar

(+) in Bile Esculin Test

Group D streptococci and Enterococcus spp.

(-) in Bile Esculin Test

(-): S. pyogenes and Viridans Streptococci

Salt Tolerance = 6.5% NaCl (+): (-):

(+): Enterococci (-): Non-enterococci = S. bovis

PYR TEST (+) (-)

(+): Enterococcus (-): S. bovis

Penicillin Test Susceptible:

S. bovis

previously classified as group D streptococci all species produce the cell wall–associated group D antigen natural inhabitants of the intestinal tracts of humans and animals

Enterococcus

most are nonhemolytic or α-hemolytic, some are β hemolytic ability to grow under extreme conditions—presence of bile or 6.5% NaCl or at 45° C or alkaline pH not highly pathogenic but frequent causes of nosocomial infection

Enterococcus

Enterococcus SPECIES

E. faecalis, E. faecium, E, avium, E. gallinarum, E. durans, E. raffinosus

can grow in extreme conditions resistant to multiple antimicrobial agents

Enterococcus

Virulence Factors of E. faecalis

 Extracellular surface adhesin proteins, extracellular serine protease, and gelatinase  Cytolysin

colonization and adherence to heart valves and renal epithelial cells

Extracellular surface adhesin proteins, extracellular serine protease, and gelatinase

 two-subunit toxin similar to bacteriocins produced by gram (+) bacteria and is expressed by a quorum sensing mechanism

Cytolysin

RELATED INFECTIONS AND DISEASES (ENTEROCOCCUS)

a. Urinary Tract Infections (UTIs) most common b. Endocarditis  elderly patients with prosthetic valves or valvular heart disease c. Bacteremia d. Intraabdominal or Pelvic Wound Infection e. CNS and Respiratory Tract infections rare

identified by its ability to grow in the presence of TELLURITE

E. faecalis

Specimen for Enterococcus

Specimen: Blood, Urine, or Wound

Culture Medium of Enterococcus

TSB or BHI with 5% sheep blood Grow well at 35° C in the presence of CO2

Selective Media for Enterococcus

Bile Esculin azide, CAN, PEA, Cephalexin-Aztreonam-Arabinose Agar

Enterococcus is identified based on their:

1. Ability to produce acid in carbohydrate broth 2. Ability to hydrolyze arginine 3. Tolerance of 0.04% tellurite 4. Utilization of pyruvate 5. Ability to produce glucopyranoside acid from 6. Growth around 100-μg efrotomycin acid disk 7. Motility

ENTEROCOCCUS BIOCHEMICAL TESTS AND RESULTS

a. Bile Esculin Test (+) b. PYR test (+) c. LAP test (+) d. Growth in 6.5% NaCl (+) e. Acid Production (+) f. Penicillin = Resistant g. Vancomycin = Resistant h. 100-μg efrotomycin acid disk = Resistant

E. faecalis requires ______ as growth factor

E. faecalis

Other organisms Bile Esculin (+) and 6.5% NaCl:

Leuconostoc Pediococcus Globicatella S. urinalis Lactococcus

Molecular Typing Methods for Enterococcus

Pulsed-field gel electrophoresis Contour-clamped homogeneous electric-field electrophoresis Ribotyping PCR-based typing methods

Antimicrobial Resistance of Enterococcus

Intrinsic or acquired resistance to aminoglycosides, β-lactams, and glycopeptides Vancomycin-resistant

aka PNEUMOCOCCUS and DIPLOCOCCUS encapsulated, characteristically lancet-shaped which occurs singly, on pairs and short chains

Streptococcus penuomoniae

contains antigen referred to as C substance which reacts with CRP facultative anaerobe requiring an increase CO2 tension (Candle Jar)

Streptococcus pneumoniae

Virulence Factors of S. pneumoniae

1. Polysaccharide Capsule 2. Adherance 3. Enzymes 4. Pneumolysin O 5. Autolysin 6. C-substance 7. Hemolysin

Enzyme that degrades surface structures of host tissue

Neuraminidase

enzyme that facilitate bacterial colonization on mucosal surfaces eliminating Ig

Protease

oxygen-sensitive toxin that is cytolytic forcells

Pneumolysin O

facilitate the release of pneumolysin O and other toxic proteins or inflammatory substance from cells

Autolysin

component of cell wall which is teichoic acid that reacts with CRP resulting in the activation of some nonspecific host immune response

C-substance

characterized by the presence of voluminous fluid which hastens in the spread of bacteria in the lungs sudden onset with chills, dyspnea, and cough

Lobar Pneumonia

most common cause of bacterial pneumonia in elderly and immunocompromised individual

Lobar Pneumonia

In Lobar Pneumonia, Microscopy of Sputum shows

large number of S. pneumonia cells and WBC; absence of oropharyngeal microbiota

follows other S. pneumonia otitis media or pneumonia most common cause of meningitis in adults

Meningitis

most common isolate in children under 3 years old with recurrent otitis media

most common isolate in children under 3 years old with recurrent otitis media

Specimen for S. pneumoniae Laboratory Diagnosis

Sputum, Swabs, Pus, CSF and Blood

Sputum that may indicate S. pneumonia

Rust-Tinged Sputum

Gram-Stain (S. pneumoniae)

Gram(+) cocci in pairs = DIPLOCOCCI cells are slightly pointed = LANCET SHAPE

Culture Medium (S. pneumoniae)

BHIA, TSA with 5% sheep’s RBC and CAP

NOTE!!!! Streptococcus pneumoniae

BAP:  Young colonies: circular, dome glistening, shaped, wet, mucoid  As the colonies become older, AUTOLYTIC CHANGES result in a collapse of each colony’s center, giving it umbilicate or doughnot appearance, Dimple-shaped (CHECKER or NAILHEAD COLONIES)

BAP (S. pneumoniae)  Colonies incubated aerobically produce __________.  Colonies incubated anaerobically produce __________ due to oxygen-labile PNEUMOLYSIN O

α-hemolysis; β-hemolysis

presumptive identification of S. pneumoniae Optochin (ETHYLHYDROCUPREIN HYDROCHLORIDE) disk is added to the surface of an SBA plate inoculated with an α-hemolytic Streptococcus

Optochin Susceptibility Test/ TAXO P

Result of Optochin Susceptibility Test/ TAXO P (S. pneumoniae)

Result:  ZOI >14 mm with a 6-mm disk = SUSCEPTIBLE  ZOI >16 mm with a 10-mm disk = SUSCEPTIBLE

based on the presence of autocatalytic enzyme AMIDASE in S. pneumoniae Under the influence of BILE SALT OR DETERGENT, bacteria’s cell wall lyses during cell division

Bile Solubility Test

Reagent and Result for Bile Solubility Test

Reagent: SODIUM DEOXYCHOLATE Result:  S. pneumoniae = solution becomes CLEAR (+)  Other α-hemolytic = solution remains CLOUDY (-)  Negative control: Suspensions made in saline

most useful, specific and rapid method for the identification of S. pneumonia (+) and allows serotyping of isolates performed by mixing on a slide loopful of emulsified sputum or CSF with a loopful of ANTICAPSULAR SERUM and METHYLENE BLUE

Neufeld-Quellang Reaction/ Capsular Swelling Test

(+) reaction (Neufeld-Quellang Reaction/ Capsular Swelling Test)

capsule appears swollen due to change in refractive index which in turn due to serologic reaction (OIL IMMERSION OBJECTIVE)

SKIN TEST for determining the presence of ANTIBODIES against pneumococci

Francis Test

Based on the sensitivity of mouse to even small inoculum of pneumococci Sputum containing pneumococci is injected intraperitoneally to a mouse which eventually dies within 16-48 hours Heart blood of the mouse contain pure culture of pneumococci

Mouse Virulence Test

- uses particle-bound antibody to enhance the visibility of the agglutination reaction between Antigen and Antibody

Coagglutination Test

confirmatory test wherein cell wall antigens are extracted either physically by heating or chemical or enzymatic extraction of cell suspension grown overnight in Todd-Hewitt Broth

Lancefield Precipitin Test

SEROLOGIC TEST FOR STREPTOCOCCI

a. Lancefield Precipitin Test b. Direct Fluorescent Antibody Test c. Coagglutination (Phadebact) d. Enzyme-linked Immunosorbent Assay (ELISA) e. Latex Agglutination Test f. ASO Titer for Group A Streptococcal Infection

formerly known as the NUTRITIONALLY VARIANT STREPTOCOCCI aka pyridoxal-dependent or vitamin B6-dependent, thiol-dependent and symbiotic streptococci

Abiotrophia and Granulicatella

grow as “satellite colonies” around other bacteria and require sulfhydryl compounds for growth part of the human oral and gastrointestinal microbiota

Abiotrophia and Granulicatella

Abiotrophia and Granulicatella Related Infections:

Bacteremia, endocarditis, otitis media, osteomyelitis, endophthalmitis after cataract extraction, brain abscess, chronic sinusitis, septic arthritis, meningitis, and breast implant– associated infections

Abiotrophia and Granulicatella Microscopy:

Gram-variable and Pleomorphic forms

Abiotrophia and Granulicatella Species:

Granulicatella adiacens, Granulicatella elegans, and Granulicatella balaenopterae, Abiotrophia defective, and Abiotrophia adjacens

Abiotrophia and Granulicatella Biochemical Characteristics

α-galactosidase β-galactosidase β-glucuronidase, Hippurate hydrolysis Arginine hydrolysis Acid production from trehalose and starch

resemble viridans streptococci

Aerococcus, Gemella, Lactococcus, Leuconostoc, and Pediococcus

commonairborne bacteria resemble viridans streptococci on culture but microscopically similar to staphylococci (tetrads or clusters) growth in 6.5% NaCl; weak catalase or pseudocatalase

Aerococcus

bacteremia and endocarditis; bile esculin(+) and PYR(+)

Aerococcus viridans

UTI, endocarditis, lymphadenitis, and peritonitis; bile esculin(-) and PYR(-)

Aerococcus urinae

similar colony morphology and habitat to viridans streptococci α-hemolytic or nonhemolytic, gram-negative cocci in pairs, tetrads, clusters, or short chains endocarditis, wounds, and abscesses

Gemella

Specie of Gemella

Gemella haemolysins

Gram(+) cocci occur singly, in pairs, or in chains and physiologically similar to enterococci α-hemolytic or nonhemolytic previously classified as group N streptococci UTI and endocarditis Does not produce acid from carbohydrates

Lactococcus

Similar biochemical characteristics with enterococci and viridans streptococci found on plant surfaces and vegetables, and in milk products meningitis, bacteremia, UTIs, and pulmonary infections

Leuconostoc

Leuconostoc SPECIES

Leuconostoc citreum, Leuconostoc cremoris, Leuconostoc dextranicum, Leuconostoc lactis, Leuconostoc mesenteroides, and Leuconostoc pseudomesenteroides

Leuconostoc Microscopy: Biochemical Test:

Microscopy: Irregular coccoid Biochemical Test: (-) catalase , (-) PYR, and (-) LAP ; (+) Bile Esculin; growth 6.5% NaCl; and production of gas from glucose

Gram(+) cocci in pairs, tetrads, and clusters that can grow at 45° C bacteremia, abscess formation, and meningitis

Pediococcus

Pediococcus SPECIES

Pediococcus acidilactici, Pediococcus damnosus, Pediococcus dextrinicus, Pediococcus parvulus, and Pediococcus pentasaceus

Pediococcus Biochemical Test:

(+) bile esculin, (+) LAP, and (-), do not produce gas from glucose, some grow in 6.5% NaCl

sepsis, meningitis, bacteremia, and UTIs α-hemolytic, PYR(+), LAP(-), and vancomycin susceptible

Globicatella sanguinis

otitis media in children nonhemolytic but α-hemolysis after prolonged incubation PYR- and LAP-(+); grow slowly in 6.5% NaCl

Alloiococcus otitidis