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microbial diseases laboratory branch

Test​​​ Name

Adult Botulism Diagnostic Testing

Testing Methodology

Botulinum Neurotoxin detection by Mouse Bioassay (MBA); isolation and identification of neurotoxigenic species of Clostridium by anaerobic culture and MBA

Reflex Testing Criteria

None

​​Pre-Approval Required

Yes. Pre-approval for testing is given by the Disease Investigation Section, Infectious Disease Branch, CDPH. Refer to MDL Point of Contact for additional questions.

Supplemental Information and Required Form(s) for Specimen Submission

Testing is limited to suspect cases of botulism in adults due to wound, foodborne, intestinal colonization, iatrogenic, or bioterrorism sources.

Form 176 (PDF) “Adult C. Botulinum Toxin Detection” from the drop-down menu on the MDL General Electronic Submission Form

The MDL General Electronic Submission Form  should be downloaded and opened using a pdf viewer. To download, right-click the link and select “Save link as.” If you are unable to open the Form using a pdf viewer, please contact MDL at 510-412-3700 or MDL.Submissions@CDPH.ca.gov.

Instructions on completing this form

Required Specimen Types

Dependent on suspected source of disease:

Suspect wound botulism: Serum (required primary specimen), and wound specimen(s) if collected. Wound specimens may be either: 1) surgically collected tissues from a wound or abscess, 2) cultures of a primary wound specimen, or 3) cultures of a suspect isolate. Wound specimens and wound cultures will only be accepted for testing if the patient has an illness suggestive of botulism.

Suspect Foodborne botulism: Serum, epidemiologically implicated foods, feces (or enema), gastric aspirate/vomitus

Adult intestinal colonization: Serum and feces

Suspect Iatrogenic botulism: Serum

Suspect bioterrorism (inhaled or ingested toxin): Serum

Optimal Volume or Mass Required

Serum: 15 ml (derived from 30 ml of whole blood collection)

Feces: 10-25 grams solid stool – or – 20 ml of enema if solid stool cannot be obtained

Food: Entire food specimen, e.g., entire jar of olives, remaining portion of left-over food.

Gastric aspirate/vomitus: 20 ml

Wound: Entire piece of surgically debrided tissue or volume of wound aspirate; swab with anaerobic transport medium used for deep wound sampling is acceptable.

Cultures: ≥10 ml of enriched broth culture

Specimen Collection Instructions

Serum: Collect 30 ml of whole blood in clot tubes. Centrifuge clot tubes to separate approximately 15 ml of serum and aliquot into transfer tubes before shipping or storage.

Enema: Collect an enema if patient is profoundly constipated. Collect fecal material using sterile, non-bacteriostatic water per rectum with a tube and syringe, or digital rectal examination. Do not use glycerin suppositories. Use a sterile urine cup as the primary collection container.

Food: Place food in secondary, leak-proof bag or container. Collect only food items that are epidemiologically linked, and/or other foods known to be potential vehicles of foodborne botulism.

Wound: Cleanse surface of wound and collect tissue deep from within wound or abscess. Place surgically debrided tissue in the smallest container possible to limit air exposure. Coordinate with microbiology lab to place the container in an anaerobic bag (often used for culture plates) in as short a time as possible. Alternatively, an anaerobic wound swab may be used to sample wound exudate from deep wounds.

Gastric aspirate/vomitus: Do not collect stomach contents if activated charcoal has been used to neutralize possible poisoning.

Specimen Stability and Acceptance Criteria

Serum: 7 days from time of collection when stored refrigerated at 2 - 8°C

Feces: 2 weeks from time of collection when stored refrigerated at 2 - 8°C - or - ​​if solid stool cannot be obtained, 20 ml of rectal wash that must contain visible feces. Overly dilute specimens are subject to rejection. 

Food: Food should be sent as soon as possible after collection to aid in the timeliness of identifying and investigating a possible foodborne outbreak of botulism. Food samples are stable if stored at the same temperature in which they were collected for several days but must be expedited to the CDPH botulism laboratory for analysis without delay.

Gastric aspirate/vomit: Accepted if collected within 3 days of onset of illness and is stable for 4 days from time of collection when stored refrigerated at 2 - 8°C. Specimen should be received by CDPH no later than 7 days from onset of illness.

Wound: 3 days from time of collection when stored refrigerated at 2 - 8°C

Culture: Stable for 2 weeks from time of inoculation when stored refrigerated at 2 - 8°C, or incubated at 30 - 37°C

Rejection Criteria

  • Unlabeled or poorly labeled specimen container (see Specimen Labeling instructions)
  • Specimens collected or shipped outside of specimen stability or shipping criteria
  • Inadequate specimen volume or mass
  • Specimens collected from a patient not authorized for testing.

Specific rejection criteria:

  1. Gastric aspirate containing activated charcoal
  2. Plasma (instead of serum)
  3. Wound tissue containing formalin fixative, aerobic wound swabs
  4. Feces collected in fixative vials
  5. Wound specimens or isolates collected from patients who do not have clinical evidence of a botulism-like illness

Storage/Transport Conditions

Storage: Store all clinical specimens refrigerated at 2-8°C. Do not freeze. Store food at the same relative temperature in which it was found at the time of collection. Store surgically collected tissue in an anaerobic atmosphere.

Transport: Transport all clinical specimens refrigerated using frozen gel packs. Do not use wet or dry ice. Transport food samples at the same relative temperature in which it was found at the time of collection.

Transport Medium, if applicable

Anaerobic swabs must contain an anaerobic generating atmosphere or anaerobic transport medium.

Sample/Specimen Labeling

All specimens require two patient identifiers, including patient full name, and at least one other identifier (e.g., specimen accession number, DOB, MRN); and date and time of collection.

Shipping Instructions and Specimen Handling Requirements

Ship using appropriate DOT/IATA approved shipping procedures. The submitter is responsible for making sure that all specimens and samples are packaged and shipped according to the current federal and state packaging and shipping regulations for “Infectious substance, Category A” and/or “Biological substance, Category B”.

MDL does not accept shipments on weekends and holidays unless prior arrangements are made with the botulism lab POC (see below).

Ship to:

California Department of Public Health
Microbial Diseases Laboratory 
ATTN: HRP – Botulism Lab
850 Marina Bay Parkway, Specimen Receiving Room B106
Richmond, CA  94804

Turnaround Time

1 week for serum testing 

2 – 4 weeks for cultures 

Note: Some cultures may take longer if isolation in pure culture is difficult, which may be dependent on the primary specimen matrix submitted for testing. Preliminary positive results will be reported the same day as observed, followed by a final report once testing is complete.

Interferences & Limitations

  1. Activated charcoal may neutralize toxin in stomach contents
  2. Formalin will sterilize tissues 
  3. HBAT will neutralize toxin circulating in serum
  4. Collection of serum soon after administering the tensilon test for myasthenia gravis will interfere with MBA 

Reference Range

None

Additional Information

Communicate with MDL botulism lab point of contact

MDL Point of Contact

For submission pre-approval and questions about specimen submission, please email MDL.Submissions@cdph.ca.gov or call the MDL Main Line to contact the High-Risk Pathogens Section at (510) 412-3700

For consultation during business hours, contact:

Jason Barash (510) 414-6738 (Primary lab POC)

Jayakumar Poovassery (510) 510-621-8743

Mahtab Shahkarami (510) 221-8196

Call during Non-Business Hours

CDPH Duty Officer at 916-328-3605
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