How do we diagnose Clostridioides (C.) difficile in stool?

Diagnosis of Clostridioides difficile in Stool

Use a two-step testing algorithm starting with a high-sensitivity screening test (GDH or NAAT) followed by toxin detection, rather than NAAT alone, to distinguish active infection from asymptomatic colonization. 1, 2

Pre-Test Requirements

Before ordering any laboratory test, ensure the patient meets clinical criteria:

• Only test patients with clinically significant diarrhea (≥3 unformed stools in 24 hours that take the shape of the container) 1, 2
• Do not test patients who received laxatives within the previous 48 hours, as this confounds results 1
• Reject formed stool specimens at the laboratory level to improve specificity 1
• Clinical context matters: recent antibiotic use, hospitalization, advanced age, abdominal pain, fever, or leukocytosis increase suspicion 1, 2

Recommended Testing Algorithms

Two-Step Algorithm (Preferred Approach)

Step 1 - High Sensitivity Screening:

• Glutamate dehydrogenase (GDH) enzyme immunoassay OR Nucleic acid amplification test (NAAT/PCR) for toxin genes 1, 2
• GDH is sensitive but cannot differentiate toxigenic from non-toxigenic strains 1
• NAAT has excellent sensitivity (91%) and specificity (98%) but may detect asymptomatic colonization if used alone 2

Step 2 - Confirmation:

• Toxin A/B enzyme immunoassay (EIA) to detect free toxins in stool 1, 2
• This step distinguishes active infection from colonization 2, 3

Alternative Algorithm Options

• GDH + Toxin EIA, arbitrated by NAAT: If GDH positive but toxin negative, perform NAAT to resolve discordance 1
• NAAT + Toxin EIA: Use NAAT for screening, confirm with toxin detection 1

Individual Test Characteristics

Reference Standards (Not for Routine Use)

• Cell culture cytotoxicity assay (CCNA): Detects toxin directly, highly specific but time-consuming (24-48 hours), requires specialized facilities 1
• Toxigenic culture: Culture on selective media (cycloserine-cefoxitin-fructose agar) followed by toxin testing of isolates; most sensitive but slow (≥48 hours) 1
• These methods are reserved for epidemiological typing and strain characterization, not routine diagnosis 1

Rapid Tests (Components of Algorithms)

• Toxin A/B EIA alone: Fast and inexpensive but NOT recommended as standalone test due to low sensitivity (32-98%, often 70-80%) despite high specificity (84-100%) 1, 2
• GDH EIA alone: Sensitive but cannot distinguish toxigenic from non-toxigenic strains (20% of C. difficile are non-toxigenic) 1
• NAAT alone: Excellent sensitivity and specificity but increases detection of asymptomatic colonization when used without toxin confirmation 1, 2

Critical Pitfalls to Avoid

• Never test asymptomatic patients or those with formed stools - this detects colonization (up to 7% of hospitalized patients), not infection 1, 2
• Do not perform "test of cure" - patients shed spores for up to 6 weeks after successful treatment 2
• Avoid repeat testing during the same diarrheal episode unless there is high clinical suspicion during an outbreak or epidemic situation 1
• Do not rely on toxin EIA alone - suboptimal sensitivity will miss cases 2

Managing Discordant Results

When screening test is positive but toxin test is negative:

• Cannot differentiate active CDI from asymptomatic colonization 3
• Base management decisions on clinical context: symptom severity, risk factors, and overall presentation 3
• Implement isolation precautions promptly even before confirmatory testing is complete to prevent transmission 3
• Consider the institutional prevalence of C. difficile when interpreting results 3

Special Considerations

• Perirectal swabs may be acceptable alternatives to stool specimens in selected patients unable to provide stool (sensitivity 95.7%, specificity 100%) 1
• Multiplex NAATs can simultaneously detect C. difficile along with other enteric pathogens, improving turnaround time 1