Pitfalls in the diagnosis of celiac disease: Bridging gaps from serology to clinical practice

Celiac disease is an immune-mediated enteropathy triggered by gluten in genetically predisposed individuals (HLA-DQ2/DQ8), with a global seroprevalence of approximately 1.4%. However, only one in four cases is clinically recognized, and diagnostic delays often exceed five to ten years. Classic presentations include diarrhea and malabsorption, but up to 50% of adults exhibit non-classical or extraintestinal manifestations (osteoporosis, iron-deficiency anemia, dermatitis herpetiformis, neuropathy). This review identifies key pitfalls in adult celiac disease diagnosis—from serology to histology and genetics—and proposes a stepwise algorithm to enhance accuracy.

Serologic Pitfalls
Serology is the first-line diagnostic step, but multiple factors compromise accuracy:

• Gluten restriction before testing: Reduces antibody titers and may normalize histology. Patients must remain on a gluten-containing diet during work-up.

• Selective IgA deficiency: Occurs in 2–3% of celiac patients. tTG-IgA and EMA-IgA are falsely negative; total IgA must be checked, and IgG-based tests (tTG-IgG, DGP-IgG) used.

• Low-titer/borderline antibodies: May reflect early disease, dietary fluctuation, or non-celiac autoimmunity (type 1 diabetes, thyroiditis). Repeat testing in 6–8 weeks on gluten is recommended.

• Assay variability: Inter-laboratory differences in calibration and antigen source can reclassify weak positives as negative. Use the same lab/assay longitudinally; laboratories should participate in external quality assessment (EQA).

• Isolated DGP-IgG in adults: Low positive predictive value; should not be used alone for diagnosis.

• Pre-analytic factors: Hemolysis can artificially lower tTG-IgA signals; repeat testing on non-hemolyzed specimens is warranted.

Histopathologic Pitfalls
Duodenal biopsy remains the gold standard but has inherent limitations:

• Patchy villous atrophy: Lesions may be confined to the duodenal bulb or skip segments. Guidelines recommend ≥5 biopsies (≥4 distal duodenum + 1 bulb) in separate jars to maximize yield. Ultrashort celiac disease (atrophy confined to bulb) is missed without dedicated bulb sampling.

• Marsh-Oberhuber subjectivity: Interobserver agreement is only 60–70%, especially for early lesions (Marsh I/II). Quantitative intraepithelial lymphocyte (IEL) counts (≥25 per 100 enterocytes) and CD3 immunostaining improve reproducibility.

• Histologic mimics: Conditions such as Giardia infection, tropical sprue, autoimmune enteropathy, drug-induced enteropathy (e.g., olmesartan), and H. pylori infection can produce similar histology. Integration with serology, clinical context, and stool studies is essential.

Genetic Testing Limitations
HLA-DQ2/DQ8 testing has a high negative predictive value (>99%) but a low positive predictive value, as 30–40% of the general population carries these alleles. Overuse in low-risk patients (e.g., IBS) leads to misinterpretation of genetic susceptibility as disease presence. HLA testing should be reserved for:

• Discordant serology and histology

• Patients already on a gluten-free diet without prior testing

• Inconclusive biopsy findings

Special Clinical Scenarios

• Seronegative celiac disease: Villous atrophy with negative celiac-specific serology. Requires: (1) adequate multisite biopsies on gluten, (2) exclusion of mimics, (3) supportive HLA, (4) clinical/histologic response to gluten-free diet (GFD) or gluten challenge.

• Potential celiac disease: Positive serology with normal histology. Approximately 30–40% progress to overt disease within five years; surveillance rather than routine GFD for all.

• Overlap syndromes: Celiac disease coexists with microscopic colitis, small intestinal bacterial overgrowth (SIBO), and IBS. Persistent symptoms on GFD warrant evaluation for these conditions.

• Refractory celiac disease and enteropathy-associated T-cell lymphoma (EATL): May present with negative serology and nonspecific symptoms. Evaluation requires cross-sectional imaging, capsule endoscopy, device-assisted enteroscopy, and immunophenotyping. Splenic atrophy and cavitating mesenteric lymph nodes are key imaging clues.

Emerging Biomarkers

• Intestinal fatty acid-binding protein (I-FABP): Released upon enterocyte injury; detects early epithelial damage before villous atrophy. Normalizes with GFD; potential for monitoring.

• MicroRNA (miRNA) panels: Differential expression of miR-192 and miR-21 correlates with active disease and normalizes with GFD. Stable in serum/plasma; requires validation.

• Microbiome signatures: Depletion of Bifidobacterium and overrepresentation of Proteobacteria are characteristic. Investigational, not yet routine.

Discussion and Proposed Algorithm
A stepwise, guideline-aligned approach optimizes diagnostic accuracy:

1. Pre-analytic: Test while patient consumes gluten; avoid trial GFD before work-up.

2. Serology: First-line tTG-IgA with total IgA; use EMA-IgA for confirmation in equivocal cases. For IgA deficiency, switch to tTG-IgG or DGP-IgG (but avoid isolated DGP-IgG in adults).

3. Biopsy: Obtain ≥5 biopsies (≥4 distal duodenum + 1 bulb) in separate jars; ensure proper orientation; report Marsh grade and IEL counts.

4. Genetics: Use HLA-DQ2/DQ8 to exclude disease in equivocal or historic cases; not for routine screening.

5. Special scenarios: For seronegative cases, exclude mimics, confirm HLA, and document response to GFD. For persistent symptoms on GFD, verify adherence, then evaluate for overlap syndromes (microscopic colitis, SIBO) before escalating to refractory work-up.

Health-system improvements include EHR alerts to delay GFD until diagnosis, lab stewardship to suppress legacy assays, and endoscopy quality metrics for biopsy adequacy.

Conclusions
Adult celiac disease diagnosis is optimized by on-gluten serology with explicit IgA-deficiency handling, bulb-inclusive biopsy protocols with adequate orientation, and a standardized framework for seronegative presentations that mandates exclusion of mimics, HLA support, and objective response to a GFD. Recognizing technical errors (hemolysis), ultrashort (bulb-only) disease, and imaging clues of complicated disease (splenic atrophy, cavitating mesenteric lymph nodes) is critical to avoid missed or delayed diagnoses and to triage patients appropriately for refractory disease and EATL work-ups. Emerging biomarkers such as I-FABP and miRNA panels may eventually serve as adjunctive tools, but further validation is required.

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The study was recently published in the Journal of Translational Gastroenterology.

Journal of Translational Gastroenterology (JTG) dedicates to improving clinical diagnosis and treatment, advancing understanding of the molecular mechanisms, and promoting translation from bench to bedside of gastrointestinal, hepatobiliary, and pancreatic diseases. The aim of JTG is to provide a forum for the exchange of ideas and concepts on basic, translational, and clinical aspects of gastroenterology, and promote cross-disciplinary research and collaboration.

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