Pertussis in Wisconsin CY 2004

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Pertussis in WisconsinCY 2004

• Dan Hopfensperger, Director
• WI Immunization Program
• 1 West Wilson Street
• P.O. Box 2659
• Madison, WI 53701-2659
• Ph 608-266-1339
• E-Mail hopfedj_at_dhfs.state.wi.us

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Outbreak Statisticsas of September 13, 2004

• 1910 Confirmed and Probable Cases
• Cases reported in 54 of 72 Counties
• gt9 fold increase in reported incidence
• 5 to 6 fold increase among children less than 10
• 4 to 9 fold increase among older children and
  adolescents (10-19)
• 17 fold increase among persons gt20 Y

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Number of Reported Pertussis Cases by County
Through 9/13/2004
Douglas
Bayfield
Iron
Ashland
Vilas
Saywer
Washburn
Burnett
Florence
Price
Oneida
Forest
Polk
Rusk
Barron
Marinette
Lincoln
Taylor
Langlade
Chippewa
St. Croix
Dunn
Menomonee
Oconto
Marathon
Pierce
Clark
Shawano
Eau Claire
Door
Pepin
Kewaunee
Waupaca
Buffalo
Wood
Portage
Brown
Outagamie
Jackson
Trempealeau
Winnebago
Manitowoc
Juneau
Waushara
Calumet
Monroe
Adams
LaCrosse
Marquette
Fond du Lac
Sheboygan
Green Lake
gt 50.0 22.1-50.0 9.1-22.0 lt 9.0 None reported
Vernon
Sauk
Columbia
Dodge
Washington
Richland
Crawford
Ozaukee
Dane
Jefferson
Iowa
Waukesha
Milwaukee
Grant
Racine
LaFayette
Green
Rock
Walworth
Kenosha
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Pertussis Rates in WI per 100,000 Population by
County Through 9/14/04
Douglas
Bayfield
Iron
Ashland
Vilas
Saywer
Washburn
Burnett
Florence
Price
Oneida
Forest
Polk
Rusk
Barron
Marinette
Lincoln
Taylor
Langlade
Chippewa
St. Croix
Dunn
Menomonee
Oconto
Marathon
Pierce
Clark
Shawano
Eau Claire
Door
Pepin
Kewaunee
Waupaca
Buffalo
Wood
Portage
Brown
Outagamie
Jackson
Trempealeau
Winnebago
Manitowoc
Juneau
Waushara
Calumet
Monroe
Adams
LaCrosse
Marquette
Fond du Lac
Sheboygan
Green Lake
gt 50.0 22.1-50.0 9.1-22.0 lt 9.0 None reported
Vernon
Sauk
Columbia
Dodge
Washington
Richland
Crawford
Ozaukee
Dane
Jefferson
Iowa
Waukesha
Milwaukee
Grant
Racine
LaFayette
Green
Rock
Walworth
Kenosha
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PertussisClinical Symptoms

• Cough of gt7 days duration
• Sleep disturbing cough
• Paroxysms
• Whoop
• Post-tussive vomiting
• Cyanosis
• Apnea
• Cough that does not respond to cough medicines
• Symptomatic patient knows a person with pertussis
  or works or attends school where pertussis is
  occurring

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Local Public Health Department (LPHD)
Investigation

• Verify demographic information
• Reporting Source
• Lab information
• Clinical signs and symptoms
• Complications
• Vaccine History
• Treatment
• Source
• Spread

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Pertussis 2004Communications

• May 7th - Letter to LPHDs in SE Region
• June 24th - Letter to Physicians Statewide
• July 23rd - Rec/Ed Camps
• August 18th - Schools
• September
• Colleges/Universities
• Day Care Centers

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PertussisLaboratories

• Prompt reporting of all suspect and confirmed
  cases to the LPHD
• Include all requested information on lab slip
• Ensure that kits are sent to SLH in timely manner

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Laboratory Diagnosis of Pertussis

• Dave Warshauer, PhD
• Chief Bacteriologist
• Wisconsin State Laboratory of Hygiene
• [phone]
• warshadm_at_slh.wisc.edu

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Bordetella species

• B. pertussis
• B. parapertussis
• B. bronchiseptica
• B. holmesii
• B. avium
• B. hinzii
• B. trematum

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Bordetella pertussis

• Small gram-negative coccobacillus
• Strictly aerobic
• Optimal growth at 35C, ambient air with humidity
• Most fastidious of the Bordetella
• Inhibited by constituents in media
• Fatty acids
• Metal ions
• Sulfides
• Peroxides

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B. Pertussis Gram Stain
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Specimen Collection

• Nasopharyngeal specimen
• Want ciliated epithelial cells
• Timing critical
• NO throat, sputum, or mouth specimens
• NP aspirates, washes, or swabs
• Dacron, rayon, or calcium alginate swabs
• NO cotton swaps

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Nasopharyngeal Specimen
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TRANSPORT

• B. pertussis extremely labile
• Direct culture optimal, not feasible
• Transport media
• If lt2hr----0.5-1.0 Casamino Acid Soln at room
  temp
• 2hr 24hr----Amies with charcoal, room temp
• gt24hr----Regan-Lowe or Jones-Kendrick at 4C

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Laboratory Diagnosis
CULTURE DFA PCR
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Isolation Media

• Bordet-Gengou
• Regan-Lowe
• Jones-Kendrick
• Stainer-Scholte synthetic medium
• Legionella buffered charcoal yeast-extract
• Incorporate antibiotics to suppress normal flora
• Incubate minimum of 7 days

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B. pertussis on Bordet-Gengou4 days incub. WSLH
Media
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B. pertussis on Bordet-Gengou7 days incub.
Commercial Medium
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Culture Sensitivity

• Considered no more than 50 sensitive
• Factors effecting sensitivity
• Type and quality of specimen
• Time specimen obtained in the course of illness
• Appropriate transport
• Choice of culture media
• Length of time cultures incubate

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Laboratory Diagnosis
CULTURE DFA PCR
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Direct Fluorescent Antibody

• In use since 1960
• Direct detection and ID of isolates

Courtesy of Mike Saubolle, PhD
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DFA

• Problems
• Poor sensitivity (18-78)
• Requires large numbers of organisms (gt104 /ml)
• Best when test early in course of illness
• Requires skilled and experienced microscopist
• Antibiotic therapy can affect binding of DFA
  reagent to cell wall
• Poor specificity (7-44 false positives)
• Advantage
• More rapid than culture

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Laboratory Diagnosis
CULTURE DFA PCR
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Pertussis PCR

• Optimal diagnostic test
• No prolonged asymptomatic carrier state
• If positive, considered diagnostic
• Recognized by CSTE as official laboratory
  confirmation of pertussis in addition to culture

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Pertussis PCR

• Nonstandardized
• Variety of targets
• Pertussis toxin (PT) promoter region
• Repeated insertion sequences (IS481)
• Adenylate cyclase toxin gene (in both B.
  pertussis and B. parapertussis)
• DNA region upstream from the porin gene
• Variety of platforms
• Conventional PCR with different detection methods
• Real-time PCR platforms

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Pertussis PCR

• Advantages
• Rapid
• Extremely sensitive
• lt1 CFU (5ul sample)
• Does not require viable organism
• Transport delays and antibiotics do not prevent
  laboratory diagnosis
• Positive longer than culture
• Specific
• Except for B. holmesii if using IS481

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Pertussis PCR

• Disadvantages
• Stringent requirements to perform PCR
• Not presently standardized
• No commercial FDA-cleared kits
• More expensive than culture or DFA
• PCR inhibitors
• Cross reaction with B. holmseii

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Pertussis PCR at WSLH

• Real-time PCR hybridization probe assay using
  LightCycler
• Primers target IS481
• Interpretation using crossing points
• No crossing point----------Negative
• Crosses at lt40 cycles----Positive
• Crosses at 40-45 cycles---Repeat
• If repeat crosses-------------Positive
• If repeat does not cross----Equivocal

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WSLH Pertussis Data
20 culture positive for B. parapertussis
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Summary

• SPECIMEN
• Nasopharyngeal aspirate/wash or 2 NP Swabs
• TESTS
• PCR test of choice
• Isolates for susceptibility testing and
  pulsed-field gel electrophoresis
• PCR performed at WSLH twice a week
• More frequently as required
• CHARGES
• PCR 75.00 CPT 87798
• Culture 21.11 CPT 87081
• Fee-exempt testing as directed by DPH

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References

• McCowen, K.L. 2002. Diagnostic tests for
  pertussis culture vs. DFA vs. PCR.
  Clin.Microbiol. Newsl. 24143-149.
• Loeffelholz, M.J. et al. 1999. Comparison of
  PCR, culture, and direct fluorescent-antibody
  testing for detection of Bordetella pertussis.
  J. Clin. Microbiol. 37 2872-2876.
• Reischl, U. 2001. Real-time PCR assay targeting
  IS481 of Bordetella pertussis and molecular basis
  for detecting Bordetella holmesii. J. Clin.
  Microbiol. 39 1963-1966.

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References

• Heininger, U. et al. 2000. Clinical validation
  of a polymerase chain reaction assay for the
  diagnosis of pertussis by comparison with
  serology, culture, and symptoms during a large
  vaccine efficacy trial. Pediatrics 105 E31.
• Katzko, G.M. et al. 1996. Extended incubation
  of culture plates improves recovery of Bordetella
  spp. J. Clin. Microbiol. 34 1563-1564.
• Black, S. 1997. Epidemiology of pertussis.
  Pediatr. Infect. Dis. J. 16 S85-S89.