Management of Streptococcal Pharyngitis: Role of the Laboratory and POC Testing Arthur E. Crist, Jr., Ph.D. Director, Clinical Microbiology York Hospital.

Management of Streptococcal Management of Streptococcal Pharyngitis: Role of the Pharyngitis: Role of the

Laboratory and POC TestingLaboratory and POC Testing

Arthur E. Crist, Jr., Ph.D.Arthur E. Crist, Jr., Ph.D.

Director, Clinical MicrobiologyDirector, Clinical Microbiology

York HospitalYork Hospital

OutlineOutline

OrganismOrganism NomenclatureNomenclature Pathogenesis & Spectrum of DiseasePathogenesis & Spectrum of Disease Clinical DiagnosisClinical Diagnosis Laboratory DiagnosisLaboratory Diagnosis

– Culture Culture – Rapid Antigen TestingRapid Antigen Testing– Molecular MethodsMolecular Methods

Gram Stain – Blood CultureGram Stain – Blood Culture

StructureStructure

Extracellular products:Pyrogenic exotoxin = erythrogenic toxin scarlet fever, toxic shock lysogenic bacteriophage, Spe-A,B,C,F, Ssa.

Hemolysins streptolysin O streptolysin S O2 sensitive O2 resistant antigenic (AST) non antigenic

EnzymesDNases A-D Hyaluronidase Streptokinase NADase

ClassificationClassification

Beta hemolysis on SBABeta hemolysis on SBA

Ignaz P. SemmelweissIgnaz P. Semmelweiss

Joseph ListerJoseph Lister

Why Diagnose Streptococcal Why Diagnose Streptococcal Pharyngitis? Pharyngitis?

Treatment Prevents Sequelae Treatment Prevents Sequelae -- Rheumatic Fever Rheumatic Fever

- Scarlet Fever - Scarlet Fever

- Glomerulonephritis - Glomerulonephritis

- Invasive Streptococcal Disease - Invasive Streptococcal Disease

- (“Flesh-eating Strep”)- (“Flesh-eating Strep”)

Local InfectionsLocal Infections

PharyngitisPharyngitis- Winter-Spring months- Winter-Spring months- School aged child, esp. 3-10 years- School aged child, esp. 3-10 years- Abrupt onset- Abrupt onset

Pyoderma (Impetigo)Pyoderma (Impetigo)

Otitis MediaOtitis Media

SinusitisSinusitis

Streptococcal Sore ThroatStreptococcal Sore Throat

ImpetigoImpetigo

Invasive InfectionsInvasive Infections ErysipelasErysipelas CellulitisCellulitis Necrotizing FasciitisNecrotizing Fasciitis Puerperal FeverPuerperal Fever SepsisSepsis EndocarditisEndocarditis Scarlet FeverScarlet Fever Toxic Shock SyndromeToxic Shock Syndrome

ErysipelasErysipelas

CellulitisCellulitis

Necrotizing FasciitisNecrotizing Fasciitis

Nonsuppurative SequelaeNonsuppurative Sequelae

Acute Rheumatic FeverAcute Rheumatic Fever– 1 to 5 weeks following acute pharyngitis1 to 5 weeks following acute pharyngitis

– 6 to 15 years of age6 to 15 years of age

– Attack rate 0.4% (civilian) to 3% (military)Attack rate 0.4% (civilian) to 3% (military)

– M types 1,3,5,6,12,18,19,24M types 1,3,5,6,12,18,19,24

– Inflammation of the heart (heart valves), joints, Inflammation of the heart (heart valves), joints, blood vessels, and subcutaneous tissuesblood vessels, and subcutaneous tissues

– Probably immunological (autoimmune?)Probably immunological (autoimmune?)

– Recurrences are commonRecurrences are common

Nonsuppurative SequelaeNonsuppurative Sequelae

Acute GlomerulonephritisAcute Glomerulonephritis– Approx. 10 days following acute pharyngitis; Approx. 10 days following acute pharyngitis;

(3 weeks following pyoderma)(3 weeks following pyoderma)– Winter/Spring (Summer/Fall-pyoderma)Winter/Spring (Summer/Fall-pyoderma)– 6 to 15 years of age (preschool-pyoderma)6 to 15 years of age (preschool-pyoderma)– Attack rate 10-15% with known neph. strainAttack rate 10-15% with known neph. strain– Limited M types involvedLimited M types involved– Probably immunological (immune complex)Probably immunological (immune complex)– Recurrences are rareRecurrences are rare

Which Streptococci Cause Which Streptococci Cause Sore Throat? Sore Throat?

Group A streptococcus (Group A streptococcus (S. pyogenesS. pyogenes))

Group C streptococcus (Group C streptococcus (S. dysgalactiae S. dysgalactiae subsp. subsp. equisimilis equisimilis) )

Group G streptococcus (Group G streptococcus (S. dysgalactiaeS. dysgalactiae subsp. subsp. equisimilisequisimilis) )

Signs of Streptococcal Signs of Streptococcal TonsillopharyngitisTonsillopharyngitis

Characteristic signsCharacteristic signs-Tonsillopharyngeal erythema -Tonsillopharyngeal erythema -Tonsillopharyngeal exudate -Tonsillopharyngeal exudate -Soft-palate petechiae (“doughnut” lesions) -Soft-palate petechiae (“doughnut” lesions) -Beefy red, swollen uvula -Beefy red, swollen uvula -Anterior cervical lymphadenitis -Anterior cervical lymphadenitis -Scarlatiniform rash -Scarlatiniform rash

Uncharacteristic signsUncharacteristic signs-Conjunctivitis -Conjunctivitis -Anterior stomatitis -Anterior stomatitis -Discrete ulcerative lesions-Discrete ulcerative lesions

Symptoms of Streptococcal Symptoms of Streptococcal TonsillopharyngitisTonsillopharyngitis

Characteristic symptomsCharacteristic symptoms-Sudden onset of sore throat -Headache -Sudden onset of sore throat -Headache -Pain on swallowing -Pain on swallowing -Abdominal pain -Abdominal pain -Fever -Fever -Nausea and -Nausea and

vomitingvomiting

Uncharacteristic symptomsUncharacteristic symptoms-Coryza -Coryza -Cough -Cough -Hoarseness -Hoarseness -Diarrhea -Diarrhea

(Bisno, NEJM)

Clinical Diagnosis of Clinical Diagnosis of Streptococcal PharyngitisStreptococcal Pharyngitis

Based on signs and symptomsBased on signs and symptoms Not very accurateNot very accurate 50% of patients with pharyngitis will be 50% of patients with pharyngitis will be

treated but will not be infected with Group treated but will not be infected with Group A StreptococciA Streptococci

30% of patients with pharyngitis will not be 30% of patients with pharyngitis will not be treated but will be infected with Group A treated but will be infected with Group A StreptococciStreptococci

Microbial Causes of Acute Pharyngitis (Bisno, NEJM)

PathogenEstimated

Percentage ofCases

ViralRhinovirus (100 types and 1 subtype) 20Coronavirus (3 or more types) 5Adenovirus (types 3, 4, 7, 14, and 21) 5Herpes simplex virus (types 1 and 2) 4Parainfluenza virus (types 1-4) 2Influenzavirus (types A and B) 2Coxsackievirus A (types 2, 4-6, 8, and 10) <1Epstein-Barr virus <1Cytomegalovirus <1Human immunodeficiency virus type 1 <1

Microbial Causes of Acute Pharyngitis (Bisno, NEJM)

Pathogen Estimated

Percentage of Cases

Bacterial Streptococcus pyogenes (Group A -hemolytic streptococci)

15-30

Group C -hemolytic streptococci 5

Neisseria gonnorrhoeae <1 Corynebacterium diphtheriae <1 Arcanobacterium haemolyticum 1

Chlamydial Chlamydia pneumoniae Unknown

Mycoplasmal Mycoplasma pneumoniae <1

Laboratory DiagnosisLaboratory Diagnosis

Culture Culture

– Specimen Collection (10% FN)Specimen Collection (10% FN)– ““Gold Standard” ??Gold Standard” ??– Selective vs. non-selective mediumSelective vs. non-selective medium– Aerobic, 5-10% COAerobic, 5-10% CO22, anaerobic atmosphere, anaerobic atmosphere– 1 or 2 day incubation1 or 2 day incubation– Broth enhancementBroth enhancement

CP1069709-1

Total positive

cultures (%)

Total positive

cultures (%)

DaysDays

SXT plus blood agarsSXT plus blood agars

Blood agar plate onlyBlood agar plate only

Aggregate antigen detection testsAggregate antigen detection tests

n=261n=261

n=189n=189

n=106n=106n=151n=151

n=187n=187

0

20

40

60

80

100

0 1 2

Cockerill. 2002. Mayo Clinic Data. AACC Internet PresentationCockerill. 2002. Mayo Clinic Data. AACC Internet Presentation

Culture MethodsCulture Methods

Roddey, et al. 1995. JAMA 274: 1863-1865Roddey, et al. 1995. JAMA 274: 1863-1865

Presumptive IdentificationPresumptive Identification

Definitive IdentificationDefinitive Identification(Biochemical or Serological)(Biochemical or Serological)

Positive PYR TestPositive PYR Test

Rapid Antigen TestsRapid Antigen Tests

Extract (acid) cell wall antigen from Extract (acid) cell wall antigen from organism on throat swaborganism on throat swab

Detect presence of extracted antigen byDetect presence of extracted antigen by– Latex agglutination (LA)Latex agglutination (LA)– Enzyme immunoassay (EIA)Enzyme immunoassay (EIA)– Optical immunoassay (OIA)Optical immunoassay (OIA)

Rapid Antigen TestRapid Antigen Test

FDA Approved KitsFDA Approved Kits

40 CLIA High Complexity (1993-1999) 40 CLIA High Complexity (1993-1999) Effective DateEffective Date

– Abbott TestPack Plus Strep AAbbott TestPack Plus Strep A 01-11- 01-11-19991999

– BD Directogen 1-2-3 Group A StrepBD Directogen 1-2-3 Group A Strep 10-27-1995 10-27-1995

– J & J CDI SureCellJ & J CDI SureCell 10-06-1995 10-06-1995

– Gen-Probe Group A Strep DirectGen-Probe Group A Strep Direct 06-10-1994 06-10-1994

Almost all are used to type organisms from cultureAlmost all are used to type organisms from culture Gen-Probe most frequently used direct specimen test in Gen-Probe most frequently used direct specimen test in

this groupthis group

FDA Approved KitsFDA Approved Kits 77 CLIA Moderate Com. (1993-2002) 77 CLIA Moderate Com. (1993-2002) Effective DateEffective Date

– Abbott Signify Strep AAbbott Signify Strep A 03-08-200203-08-2002

– Quidel Quickvue Strep A TestQuidel Quickvue Strep A Test 03-05-200203-05-2002

– Abbott TestPack Plus Strep A w/OBC IIAbbott TestPack Plus Strep A w/OBC II 01-11-199901-11-1999

– Binax Strep A TestBinax Strep A Test 04-19-199604-19-1996

– J & J CDI SureCellJ & J CDI SureCell 10-06-199510-06-1995

– Abbott TestPack Plus Strep A Abbott TestPack Plus Strep A 07-26-199307-26-1993

– Baxter MicroScan Cards O.S.Baxter MicroScan Cards O.S. 07-26-199307-26-1993

– BD Directogen 1-2-3 Grp A StrepBD Directogen 1-2-3 Grp A Strep 07-26-199307-26-1993

– Binax Equate Strep A Binax Equate Strep A 07-26-199307-26-1993

– BioStar Strep A OIABioStar Strep A OIA 07-26-199307-26-1993

– Hybritech ICON Strep A Hybritech ICON Strep A 07-26-199307-26-1993

– Meridian Diagnostics ImmunoCardMeridian Diagnostics ImmunoCard 07-26-199307-26-1993

– Quidel Group A StrepQuidel Group A Strep 07-26-199307-26-1993

– Wampole Bactigen Group A StrepWampole Bactigen Group A Strep 07-26-199307-26-1993

FDA Approved KitsFDA Approved Kits

24 CLIA Waived (1996-2002) 24 CLIA Waived (1996-2002) Effective Date Effective Date – Genzyme OSOM Ultra Strep A TestGenzyme OSOM Ultra Strep A Test 07-19-200207-19-2002– Quidel Quickvue In-Line Strep AQuidel Quickvue In-Line Strep A 07-08-200207-08-2002– Acon Strep A Rapid Test StripAcon Strep A Rapid Test Strip 12-18-200112-18-2001– Beckman Coulter ICON DS Strep ABeckman Coulter ICON DS Strep A 12-18-200112-18-2001– Quidel Quickvue Dipstick Strep A Quidel Quickvue Dipstick Strep A 07-26-200107-26-2001– ICON DS Strep AICON DS Strep A 07-17-200107-17-2001– Beckman Coulter ICON FX Strep ABeckman Coulter ICON FX Strep A 03-16-200103-16-2001– Wyntek Diagnostics OSOM Ultra Strep AWyntek Diagnostics OSOM Ultra Strep A 07-06-200007-06-2000– Fisher Sure -Vue Strep AFisher Sure -Vue Strep A 09-22-199909-22-1999– Meridian ImmunoCard STAT Strep AMeridian ImmunoCard STAT Strep A 05-19-199805-19-1998– Abbott Signify Strep A TestAbbott Signify Strep A Test 09-22-199709-22-1997– Biostar Acceava Strep A TestBiostar Acceava Strep A Test 07-09-199707-09-1997– Binax NOW Strep A TestBinax NOW Strep A Test 01-10-199701-10-1997

Factors Affecting ADT SensitivityFactors Affecting ADT Sensitivity

Culture methods selected in a study influence the Culture methods selected in a study influence the calculated ADT sensitivitiescalculated ADT sensitivities

Studies are likely to report higher ADT sensitivities Studies are likely to report higher ADT sensitivities when cultures are performed in physician officeswhen cultures are performed in physician offices

When culture methods result in identification of very When culture methods result in identification of very small numbers of GABHS, the calculated sensitivity small numbers of GABHS, the calculated sensitivity of an ADT will be lower than that calculated when a of an ADT will be lower than that calculated when a less rigorous culture method is used because ADTs less rigorous culture method is used because ADTs are likely to fail to detect low numbers of GABHSare likely to fail to detect low numbers of GABHS

Comparison of a CLIA Waived Comparison of a CLIA Waived & Moderately Complex Test& Moderately Complex Test

Mod. Complex ADTMod. Complex ADT Sens. (%) Spec. (%)Sens. (%) Spec. (%)

Waived ADTWaived ADT 9191 93 93

Gold StandardGold Standard

Sens. (%) Spec. (%)Sens. (%) Spec. (%)

Waived ADTWaived ADT 8080 95 95

Gold StandardGold Standard

Sens. (%) Spec. (%)Sens. (%) Spec. (%)

Mod. Com ADTMod. Com ADT 8282 100 100

Comparison of the Abbott Signify Group A Test to Comparison of the Abbott Signify Group A Test to Culture – York Hospital Dec. 1999 to Mar. 2000Culture – York Hospital Dec. 1999 to Mar. 2000

(n=397)(n=397)

StudyStudy Sens. (%) Spec. (%) PVP (%) Sens. (%) Spec. (%) PVP (%) PVN (%) PVN (%) ____________________________________________________________________________________________________________________________________

YH*YH* 82.9 82.9 97.8 97.8 90.0 90.0 96.0 96.0

PI**PI** 96.0 96.0 97.8 97.8 ND ND ND ND

•*York Hospital*York Hospital•** Package Insert** Package Insert

Sensitivity and Specificity of the OIA

Study Sensitivity (%)

Specificity (%)

Harbeck 1993 97.4 95.6Harbeck 1993 98.9 98.4Dale 1994 81 97.5Roe 1995 83 89Heiter 1995 92.3 95.4Fries 1995 94 98Harris 1995 96 94 Baker 1995 78 90Gerber 1997 84 93Hart 1997 77 62Pitetti 1998 79.5 96.5Kuhn 1999 89 96.5Chapin 2002 86.1 97.1

Rapid Group A Strep Tests

1.0E+00

1.0E+01

1.0E+02

1.0E+03

1.0E+04

1.0E+05

1.0E+06

Test

Mea

n s

ensi

tivi

ty (

cfu

/sw

ab)

Error bars = 1 standard deviation

Comparison of Test SensitivityComparison of Test Sensitivity

Cockerill. 2002. Mayo Clinic Data. AACC Internet PresentationCockerill. 2002. Mayo Clinic Data. AACC Internet Presentation

Conclusions: Rapid Antigen Kit Analytical Sensitivity Comparison

Detection of different isolates by different tests showed no differences

Analytical Sensitivities were best for the OIA (3.3 x 104 CFU) and worst for Directigen (5 x 105 CFU)

Cockerill. 2002. Mayo Clinic Data. AACC Internet PresentationCockerill. 2002. Mayo Clinic Data. AACC Internet Presentation

Molecular Tests for Detection of Group A Streptococcus from Throat Swabs -

Not Point-of-Care

• Group A Streptococcus Direct Test (Gen-Probe)

• PCR – Conventional or Real Time

Gen-Probe Group A Streptococcus Direct Test

• Test detects rRNA sequences using a chemiluminescent single-stranded DNA probe

• Enhanced sensitivity vs. Antigen detection methods

Percent

Test Sensitivity Specificity Positive

predictive value

Negative predictive

value

Heiter 1993 93.5 99.7 99.0 97.8Pakorski 1994 85.7 97.8 83.9 98.1Chapin 2002 94.8 100 100 96.9

Comparison Of The Gen-Probe Test and Culture

Polymerase Chain ReactionPolymerase Chain Reaction

UnamplifiedDNAUnamplifiedDNA

Denatureand annealprimers

Denatureand annealprimers

PrimerextensionPrimerextension

Denatureand annealprimers

Denatureand annealprimers

Cycle 0Cycle 0

Cycle 1Cycle 1

Targeted sequenceTargeted sequence

Cycle 2Cycle 2

Cycle 3Cycle 3

Denatureand annealprimers

Denatureand annealprimers

PrimerextensionPrimerextension

PrimerextensionPrimerextension

Cycles 4-25Cycles 4-25

At least 105-fold increase in DNAAt least 105-fold increase in DNA

Comparison of PCR Comparison of PCR to Other Methodsto Other Methods

TestTest Sensitivity (%) Specificity (%) PVP (%) PVN (%) Sensitivity (%) Specificity (%) PVP (%) PVN (%)________________________________________________________________________________________________________________________________________

OIAOIA 76 76 92 92 85 85 87 87

Agar cultureAgar culture 79 79 100100 100 100 90 90

Both-enhanced cult. 86 Both-enhanced cult. 86 100100 100 100 93 93

Combined cult.Combined cult. 89 89 100100 100 100 94 94

OIA & agar cult.OIA & agar cult. 89 89 92 92 86 86 94 94

PCRPCR 96 96 100100 100 100 98 98

Kaltwasser, et al. 1997. Pediatr. Infect. Dis. J. 16: 748-753Kaltwasser, et al. 1997. Pediatr. Infect. Dis. J. 16: 748-753

Management of Streptococcal PharyngitisManagement of Streptococcal Pharyngitis

Hofer, et al. 1997. Arch. Pediatr. Adolesc. Med. 151: 824-829.Hofer, et al. 1997. Arch. Pediatr. Adolesc. Med. 151: 824-829.

Management of Streptococcal PharyngitisManagement of Streptococcal Pharyngitis % Physicians Ordering Test% Physicians Ordering Test

__________________________________________________________________________________________________________________________________________

Private HMO Residents Private HMO Residents Total Total (n=50) (n=50) (n=50) (n=150) (n=50) (n=50) (n=50) (n=150)________________________________________________________________________________________________________________________Case ACase A Throat culture Throat culture 100 100 100 100 100 100 100 100 20-minute ADT 20-minute ADT 74 74 90 88 84 90 88 84 4-hour ADT 4-hour ADT 32 46 40 39 32 46 40 39

Case BCase B Throat cultureThroat culture 80 64 66 80 64 66 70 70 20-minute ADT20-minute ADT 54 54 54 54 54 54 54 54 4-hour ADT 4-hour ADT 18 30 20 23 18 30 20 23

Berwick, et al. 1987. Pediatr. Infect. Dis. J. 6: 1095-1102Berwick, et al. 1987. Pediatr. Infect. Dis. J. 6: 1095-1102

Management of Streptococcal PharyngitisManagement of Streptococcal Pharyngitis % of Physicians Who “Believe” ADT% of Physicians Who “Believe” ADT

__________________________________________________________________________________________________________________________________________

Private HMO Residents Private HMO Residents Total Total (n=40) (n=43) (n=36) (n=119) (n=40) (n=43) (n=36) (n=119)________________________________________________________________________________________________________________________Case ACase A Positive Positive 98 93 98 93 94 94 95 95 Negative Negative 42 42 42 50 45 42 50 45

(n=44)(n=44) (n=45)(n=45) (n=43) (n=132) (n=43) (n=132)Case BCase B PositivePositive 89 93 72 89 93 72 85 85 NegativeNegative 73 73 82 79 78 82 79 78

Berwick, et al. 1987. Pediatr. Infect. Dis. J. 6: 1095-1102Berwick, et al. 1987. Pediatr. Infect. Dis. J. 6: 1095-1102

Management of Streptococcal PharyngitisManagement of Streptococcal PharyngitisEffect of ADT Availability on TreatmentEffect of ADT Availability on Treatment

Patients Treated (per100)Patients Treated (per100)__________________________________________________________________________________________________________________________________________

Private HMO Residents Private HMO Residents Total Total (n=50) (n=50) (n=50) (n=150) (n=50) (n=50) (n=50) (n=150)________________________________________________________________________________________________________________________Case ACase A None None 92 84 82 86 92 84 82 86 20-minute ADT20-minute ADT 70 70 59 67 65 59 67 65 4-hour ADT 4-hour ADT 82 68 75 75 82 68 75 75

Case BCase B NoneNone 19 6 8 11 19 6 8 11 20-minute ADT20-minute ADT 17 17 16 16 16 16 16 16 4-hour ADT 4-hour ADT 19 10 11 13 19 10 11 13

Berwick, et al. 1987. Pediatr. Infect. Dis. J. 6: 1095-1102Berwick, et al. 1987. Pediatr. Infect. Dis. J. 6: 1095-1102

Conclusions - Rapid Testing for Streptococcal Infection

Rapid Antigen Detection (LA, EIA, or Rapid Antigen Detection (LA, EIA, or OIA-based, point-of-care) OIA-based, point-of-care) - Culture if Rapid Antigen Result Negative- Culture if Rapid Antigen Result Negative

Molecular methods, e.g. Real-time PCR, may equal or exceed sensitivity provided by culture

(American Academy of Pediatrics) (American Academy of Pediatrics) (Infectious Disease Society of America)(Infectious Disease Society of America)

Any Questions?Any Questions?