Celiac disease (CD) is a common autoimmune gastrointestinal disease. Several serological tests are available to screen for CD. Since CD can present with fatigue, serological screening was incorporated into a ‘tired all the time’ testing profile available to general practitioners. The utility of this inclusion was audited and the results are presented here.

by L. Hughes, Dr A. Ballantyne, Dr C. Ford, Dr A. Ekbote and Prof. R. Gama

Introduction
Celiac disease (CD) is an autoimmune disease caused by sensitivity to gliadin proteins, a component of gluten found in wheat, rye and barley. It primarily affects the small intestine, leading to villous atrophy and crypt hyperplasia, causing malabsorption. Common symptoms include diarrhea, abdominal bloating, weight loss, fatigue and faltering growth. CD is estimated to affect 1% of the population, although many of these are undiagnosed [1]. It mostly occurs in genetically pre-disposed individuals [2], particularly those with HLA-DQ2 and HLA-GQ8 genotypes, and approximately 90% of CD patients carry HLA-DQ2 [3]. Treatment of CD is primarily by elimination of gluten-containing foods such as pasta, cereals, cakes and sauces, therefore specialist dietitian input is often required. Supplements such as iron, folate, vitamin B12 and vitamin D may also be required while the digestive system repairs itself. Patients with CD frequently have antibodies to tissue transglutaminase, with the titre correlating well with the degree of muscosal damage [4]. Diagnosis of CD is important to inform patient lifestyle choices, and minimize villous atrophy, malabsorption, nutritional deficiencies and long-term adverse events.

Testing for CD
The gold standard for diagnosis of CD is histological evaluation of endoscopic duodenal biopsy specimens, which is an invasive and labour-intensive investigation. The National Institute for Health and Care Excellence (NICE), therefore, recommends serum serological tests as screening tests for endoscopic biopsy [5]. The first-line serological test for CD is IgA anti-tissue transglutaminase antibody (IgA-tTG) and if IgA-tTG is weakly positive then testing for IgA anti-endomysial antibody (IgA-EMA) is indicated.

One problem with IgA-tTG analysis is that 3% of patients with CD have co-existing total IgA deficiency, which may cause falsely negative IgA-tTG results. NICE guidance recommends concurrent measurement of IgA and if deficient, IgG-tTG, IgG-EMA and IgG anti-deamidated gliadin peptide antibody are suggested alternative serological tests to IgA-tTG.

Those with positive serological tests are then referred for endoscopic intestinal biopsy to confirm or exclude CD. Patients must be on a gluten-containing diet for at least 6 weeks before serological and histological tests [6, 7]. IgA-tTG and IgA-EMA, for example, have a sensitivity of below 50% in detection of villous atrophy in patients already following a gluten-free diet at time of testing [7]. NICE guidance does not recommend genetic testing in the diagnosis of CD, despite the strong genetic link, except in special circumstances [5].

The CD diagnostic pathway at Royal Wolverhampton NHS Trust (RWT), at the time of this study, (Fig. 1) differed from current NICE guidance, as all positive IgA-tTG results were reflexed for confirmatory IgA-EMA testing.

Inclusion of CD testing in a ‘tired all the time’ profile
The ‘tired all the time’ (TATT) profile is one of a number of condition-specific profiles that are available for electronic ordering by general practitioners (GPs) at RWT. The profiles are designed to provide appropriate first-line and reflex tests for certain conditions, and have been locally agreed.

The TATT profile includes a full blood count, urea and electrolytes, liver function tests, bone profile, glucose, hemoglobin A1c (HbA1c) and thyroid function tests. Further tests are reflexed dependent on first-line test results, for example ferritin is added if hemoglobin and mean cell volume are low. It was suggested that a CD screen should be added to the TATT profile as patients with CD may present with fatigue initially.
Effect of inclusion of CD in a TATT profile on diagnosis of celiac disease
A study was carried out to determine the utility of including the celiac screen in the GP TATT profile. Over a period of 34 weeks, there were 1468 TATT profile requests. Nineteen patients had a positive (+ve) IgA-tTG test (tTG>7 U/mL), of whom 13 were also IgA-EMA +ve. Of the 59 patients with an IgA-tTG concentration of 0 U/mL, one was IgG-tTG +ve (Fig. 2).

Ten IgA-EMA +ve patients were referred for small bowel biopsy, two had been previously diagnosed with CD and one patient had no recorded follow-up. Of those biopsied, CD was confirmed in six and excluded in one. Results were unavailable in three patients (biopsy delayed owing to pregnancy, private biopsy and did not attend for biopsy).

Four of the six IgA-EMA negative (−ve) patients were referred for small bowel biopsy, two had CD confirmed, one refused biopsy but had genotyping consistent with CD, and one patient had negative histology. Two patients had no recorded follow-up, and there was no recorded follow-up for the IgG-tTG +ve patient.

Aside from serological CD tests, CD patients can show other biochemical and hematological abnormalities, including anemia, hypocalcemia and raised liver enzymes. Of the 20 patients who were IgA/IgG-tTG +ve, none had low hemoglobin, three had low adjusted calcium, and one had low adjusted calcium and mildly elevated liver enzymes. This means that in 16 patients, the original tests in the TATT profile would not have indicated the need to investigate for CD.

Overall, 1.4% of the patients audited over the 9 month period had ≥1 positive serological tests for CD.
Discussion
The introduction of serological testing for CD as part of the TATT profile identified a minimum of nine previously undiagnosed patients. Other tests in the profile were not reliable indicators of the need to investigate for CD. Earlier detection of CD allows dietary intervention, repair of the intestinal mucosa and decreased long-term risks. Remaining on a gluten-containing diet may lead to multiple complications for CD patients, including an increased risk of small bowel lymphoma [8].

NICE guidance recommends that patients with positive serological test results should be referred for endoscopic biopsy to confirm or exclude CD [5]. We provided an interpretive comment advising endoscopic biopsy for patients who were IgA-tTG +ve/IgA-EMA +ve, and 10 of 11 (91%) previously undiagnosed patients were referred for endoscopic biopsy. Although not prompted, requesting clinicians referred four (67%) of the six patients who were IgA-tTG +ve/IgA-EMA −ve for endoscopic biopsy. This implies that laboratory comments may serve as useful prompts to appropriately follow up these patients. Of the four IgA-tTG +ve/IgA-EMA −ve patients, three were subsequently diagnosed with CD. Confirmatory EMA testing following an IgA-tTG +ve result may, therefore, reduce the number of Gastroenterology referrals, but increases the risk of leaving patients undiagnosed.

Conclusion
In conclusion, the study has shown value in adding a CD screen into a TATT profile, with at least 0.6% of patients requesting the profile being diagnosed with CD. Laboratory interpretative comments on positive serological results are useful prompts for endoscopic biopsy referral. Clinical follow-up for patients with a positive IgA-tTG and a positive IgA-EMA was generally carried out appropriately, but patients with a positive IgA-tTG and negative IgA-EMA, or positive IgG-tTG, received variable follow-up, and this may be due to lack of interpretive comments on these results. In IgA-tTG +ve patients, IgA-EMA testing was a poor discriminator between those with and without CD. IgA-EMA, therefore, is no longer reflexed for all positive IgA-tTGs, but is reserved for use in patients with equivocal IgA-tTG results. Our interpretative comment now advises that all IgA-tTG +ve and IgG-tTG +ve patients, not known to have CD, are referred for endoscopic biopsy.

References
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The authors
Lauren Hughes*¹ MSc, Angela Ballantyne² PhD, Clare Ford¹ PhD, Anjali Ekbote¹ MD, Rousseau Gama¹ MD
¹Clinical Chemistry, New Cross Hospital, Wolverhampton, UK
²Clinical Chemistry, Royal Infirmary of Edinburgh, UK

*Corresponding author
E-mail: Lauren.hughes2@nhs.net