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Table of Contents
REVIEW ARTICLE
Year : 2020  |  Volume : 8  |  Issue : 1  |  Page : 34-42

Diagnostic modalities used for celiac disease: Advancements and future prospects


1 Multi Disciplinary Research Unit, Guru Gobind Singh Medical College and Hospital, Faridkot, Punjab and Adarsh Vijendra Institute of Pharmaceutical Sciences, Shobhit University, Gangoh, Uttar Pradesh, India
2 Multi Disciplinary Research Unit, Guru Gobind Singh Medical College and Hospital, Faridkot, Punjab, India
3 School of Biological Engineering and Life Sciences, Shobhit Institute of Engineering and Technology, Meerut, India
4 Adarsh Vijendra Institute of Pharmaceutical Sciences, Shobhit University, Gangoh, Uttar Pradesh, India
5 University Centre of Excellence in Research, Baba Farid University of Health Sciences, Faridkot, Punjab, India

Date of Submission04-Feb-2020
Date of Decision01-Jun-2020
Date of Acceptance09-Jun-2020
Date of Web Publication30-Jun-2020

Correspondence Address:
Dr. Malika Arora
Research Scientist I, Multidisciplinary Research Unit, Guru Gobind Singh Medical College and Hospital, Baba Farid University of Health Sciences, Faridkot - 151 203, Punjab
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/JIHS.JIHS_3_20

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  Abstract 


Celiac disease (CD) is an autoimmune disorder that occurs generally in genetically predisposed individuals due to the ingestion of gluten and related proteins. Initially, the occurrence was reported only in few of the countries, specifically in predominant Caucasian populations, but nowadays, it is being reported from almost all parts of the world. The increased prevalence rate of CD worldwide is because of an increase in awareness and detection of the disease due to advancements in diagnostic modalities which were not available in earlier times. As per current scenario, the clinical demonstration of the disease vary greatly due to multiple and varying symptoms; hence, there is a great difficulty in designing of specific and sensitive diagnostic tests. All the available diagnostic tools are invasive in nature leading to increased patient compliance. Serological (antigliadin antibody, EMA, transglutaminase immunoglobulin A, and deamidated gliadin peptide) test results always need to be verified by biopsy test in order to confirm the disease. A step-by-step diagnostic approach should always be followed by the doctors as well as patients for proper diagnosis and management of the disease. Keeping the scarcity of patient-friendly, specific, and sensitive diagnostic tools in mind, the present manuscript has been compiled to highlight different diagnostic modalities to enable coeliac disease scientists to find out and develop novel diagnostic techniques for this disease. In addition, the article also highlights the advancements in the development of new techniques over advantages and disadvantages of existing techniques.

Keywords: Biopsy, celiac disease, diagnostic modalities, invasive, transglutaminase immunoglobulin A


How to cite this article:
Kumari M, Arora M, Kaur N, Singh R, Bansal P. Diagnostic modalities used for celiac disease: Advancements and future prospects. J Integr Health Sci 2020;8:34-42

How to cite this URL:
Kumari M, Arora M, Kaur N, Singh R, Bansal P. Diagnostic modalities used for celiac disease: Advancements and future prospects. J Integr Health Sci [serial online] 2020 [cited 2020 Jul 15];8:34-42. Available from: http://www.jihs.in/text.asp?2020/8/1/34/288687




  Introduction Top


Nowadays, celiac disease (CD) is becoming a common health problem issue as the disease is distributed worldwide throughout the globe, with considerable variation in prevalence even among geographically proximate populations. The prevalence of CD is about 1%–3% of the population, in most of the areas of the world [1] and its incidence has been reported to increase consistently during the past several decades.[2] It is an autoimmune disorder generally precipitated in genetically predisposed people on exposure to gluten a protein present in wheat, rye, barley, and oats.[3] The exposure of gluten leads to damage of their intestinal mucosa, leading to poor absorption as well as distribution of food and finally resulting in malnutrition. In fact, body of people with CD upon ingestion of gluten shows an immune response which attacks the small intestine. These attacks cause damage to the villi that are small finger like projections lining the small intestine responsible to promote nutrient absorption. When the villi get damaged, nutrients remain unabsorbed in the body. This interplay between gut microbiota and the mucosal immune system is supposed to play a major contribution in CD.[4],[5]

It has been observed that CD is considered to be genetic disease as it runs within families depending on their human leukocyte antigen (HLA) typing. In a family, if a person is suffering from CD, first-degree relatives such as parent, children, or sibling have a possibility of developing the disease and the probability of the same is 1 in 10. The major genetic risk factor involved in CD is found to be HLA-DQ genes. Around 90% of the affected individuals carry the HLA-DQ2 haplotype, 5% carry the DQ8 haplotype, and the other 5% carry at least one of the two DQ2 alleles.[6],[7] Although genetic predisposition plays an important role in the development of CD, but as per the previous reports, it has been revealed that the HLA gene itself is not sufficient for the development of disease.[8] As per the data available, it has been observed that around 30% of the general population carry the HLA-DQ2/8 CD susceptibility genes; however, only 2%–5% among them will develop CD, suggesting that additional environmental factors are also contributing to disease development.[9]

Overall, the prevalence rate of CD is increasing worldwide due to an increase in awareness and detection of the disease due to advancements in diagnostic modalities which were not available in earlier times.[10] As per the current scenario, the clinical demonstration of the disease varies greatly due to multiple and varying symptoms; hence, there is a great difficulty in designing of specific and sensitive diagnostic tests.[11]

Currently, serological testing using anti-transglutaminase immunoglobulin (Ig) A (tTG) antibody is quite famous test for diagnosis of the disease. It is helpful to confirm a great number of borderline cases in patients with mild intestinal lesions, etc.[12] Sometimes, rather most of the times, on the basis of the tTG reports, patients are advised to follow gluten-free diet (GFD) without confirmation with other tests which are not appropriate approach. The other diagnostic modality to be used is biopsy, which is considered to be the gold standard for diagnosis due to histological changes (most relevant feature of the disease). Hence, in today's scenario, the diagnosis of the disease is considered to be complete on the basis of combination of serological tests as well as confirmation by histological demonstration of duodenal mucosal damage. In recent years, a progressive decline in the use of this confirmatory diagnostic tool has been observed due to its invasive nature.[8],[13],[14] Keeping the scarcity of patient-friendly, specific, and sensitive diagnostic tools in mind, the present manuscript has been compiled to highlight different diagnostic modalities to enable coeliac disease scientists to find out and develop novel diagnostic techniques for this disease. In addition, the article also highlights the advancements in development of new techniques over advantages and disadvantages of existing techniques.


  Celiac Disease Development Top


CD is a systemic autoimmune disorder preferentially triggered by gluten peptides present in the grains including wheat, rye, and barley. More than 90% of people with CD carry one of the two major histocompatibility complex class-II molecules, namely HLA-DQ2 or DQ8.[15] These genes present gluten peptides to body's defense system in such a manner that it activates an antigen-specific T-cell response. DQ2 or DQ8 genes are the major factor which helps in the genetic predisposition to CD.[16] However, it is observed from various studies that being DQ2 or DQ8 positive is not only the essential reason for developing CD. The additional environmental or genetic factors contribute in together with genetic factor in order to develop CD.[17] The various factors for development of the disease are represented in [Figure 1].
Figure 1: Representation of various factors responsible for development of Celiac disease

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  Evolution of Diagnostic Tools for Celiac Disease Top


Diagnostic tools for CD evolved from primitive symptomatic features to latest sequencing techniques in the last 70 years.[18] Until the 1950s, CD was diagnosed based on clinical observations focused on malabsorptive features such as chronic diarrhea, abdominal distension, and weight loss (or poor weight gain) occurring in young children a few months after the introduction of solid food to their diet.[19] A substantial change in the diagnostic paradigm was produced by development of the peroral intestinal biopsy in 1955–1956. During that period of time, gluten-dependent enteropathy has been based on histological assessment of intestinal mucosa and it was the standard for diagnosis.[20] Ongoing research led to the development of sensitive and specific serologic tests for CD in the 1980s. Prior to the 1980s, a lack of tests other than biopsy testing for CD severely limited diagnosis rates. Serological tests are now used as the first step tool to clarify a suspicion of CD and to decide/identify patients who should undergo intestinal biopsy analysis. In the last few years, the rate of diagnosis in CD patients has been increased due to increase in awareness regarding disease as well as development in diagnostic tools.[21] [Table 1] represents chronology of evolution of diagnostic tools for CD.
Table 1: Chronology of evolution of diagnostic test for celiac disease

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  Diagnostic Tools Used for Celiac Disease Current State of Art Top


In the early 1950's, it has been observed that CD is an immune-mediated disorder that primarily affects the gastrointestinal (GI) tract which is caused due to intake of gluten.[28] Further investigations indicated that the mucosa regains its integrity once gluten is withdrawn from the diet.[29] This information revolutionized the diagnosis and management of CD. Keeping this in mind, various diagnostic tools were developed. As per the current status, biopsy is the gold standard in diagnosis, but serological tests hold significant situate in determining whether to undergo endoscopy/biopsy. Initially, blood antibodies were being used to mark the presence or absence of CD and they were termed as serological diagnostic markers of CD, but findings of serological tests were not enough for the diagnosis of CD. Hence, invasive pathological diagnostic modalities such as endoscopy/biopsy were introduced for further confirmation and clinical management of CD. The current state of art of various techniques is as follows:

Serological test

Nowadays four types of serological tests are commercially available in laboratories based on anti-gliadin IgA and IgG (AGA IgA and AGA IgG), anti-reticulin IgA, anti-Endomysial IgA (EMA), and anti-tissue transglutaminase (TTG) antibodies.[30] These tests particularly are helpful in preliminary diagnosis of CD patients using their serum sample. Moreover, these tests are being widely used in epidemiology studies for determining the seroprevalence of disease. Most commonly used serological tests include as follows.

Antiglidin antibodies (AGA immunoglobulin A and AGA immunoglobulin G)

It was the first serological marker established in the mid-1980s for CD screening.[31] Anti-gliadin antibodies (AGA) were supposed to be reasonably accurate in previous years. This test is widespread, simple, and informative test when AGA of both IgG and IgA classes is measured. Antigliadin antibodies (AGAs) are antibodies of the IgA and IgG classes found in the sera of CD patients as these antibodies mainly target gliadin-derived peptides, which are the main proteins of gluten.[32] In addition, the specificity and sensitivity of the test is under question as it also gives positive findings with other GI diseases such as gastritis, gastroenteritis, and inflammatory bowel disease (IBD). Although it is an enzyme-linked immunosorbent assay (ELISA)-based test, the positive predictive value in moderate-risk groups is <30%.[33],[34] Due to its lower sensitivity and specificity, this test has been replaced by other serological tests such as EMA, anti-tTG, and anti-deamidated gliadin peptide (DGP) tests etc.

Limitation of antigliadin antibody

  • AGA is an anti-food protein antibody; as such, they are not indicative of any autoimmune reactions. Thus, patients with true nonceliac gluten sensitivity, patients with IBS and no gluten sensitivity, as well as individuals who are totally healthy (perhaps a bit less commonly, but not significantly so) may all have positive (or negative) AGA. AGA should not be relied upon to prove or disprove the diagnosis of CD or nonceliac gluten sensitivity [34]
  • Their predictive value is quite significantly lower than that of EmA and tTG antibodies [35]
  • Time-consuming protocols and unsuitable for testing large numbers of samples
  • AGA displays a very low specificity, since their production is largely dependent on the increased permeability of the intestinal mucosa to gliadin, also occurring in several pathological conditions other than CD.[36]


Endomysial antibodies

This test became available in 1983, as a diagnostic tool for CD due to high sensitivity and specificity toward CD. Anti-endomysial antibodies (EMA) are IgA class auto-antibodies directed against endomysium, the collagen matrix of human and monkey tissues and is used as substrate over the last 30 years.[37] This test identifies anti-gluten circulating antibodies, gliadin, or reticulin, using immunofluorescence or the ELISA. The IgA EMA can also be performed using human umbilical cord as they are having antibody/substrate with pooled sensitivity and specificity.[38]

Limitations of endomysial antibody test [24],[37],[38],[39]

  • The sensitivities and specificities of these tests have varied from center to center and this variability has been a drawback to their diagnostic use
  • More time-consuming to perform and more expensive
  • Furthermore, the assay requires the laboratory technician to assess for Immunofluorescence, but the use was curtailed and deceased day by day due to cost and interpretability issues
  • More chances of interpretation error as interpretation are operator dependent
  • This test has more potential to give errors due to sensitivity and specificity in different age groups. The sensitivity and specificity range of test varies in various age groups (children, adults because of the difference in umbilical cord associated antibodies that may differ from person to person
  • The EMA test may be less accurate in children under 2 years of age and hence cannot be used as a sure shot indicator of CD as the diagnosis is based on the villous dystrophy, which is a variable condition. In addition, EMA cannot detect CD in its early stage. CD People with less villous atrophy cannot be detected by EMA.


Tissue transglutaminase autoantibody enzyme-linked immunosorbent-based assay

In 1997, tTG was discovered using guinea pig protein that is the antigen against which endomysial antibodies were being formed was the enzyme tTG.[24],[25] A tTG-IgA test may be done when patients has CD symptoms such as poor growth, belly pain, diarrhea, allergy, constipation, and vomiting. It is helpful to confirm a great number of borderline cases in patients with mild intestinal lesions and positive serology, diagnosing the so-called potential CD.[39],[40] It is likely that serology could identify CD in its early stages, before the appearance of a severe intestinal damage. Anti-tTG2 is auto-antibodies of class IgA and IgG produced by tTG2-specific B cells. Specifically, IgA auto-antibodies recognize the enzyme tTG2, making specific markers for CD tTG2. TTG is a calcium-dependent cytosolic protein possessing both intracellular and extracellular functions; it appears to play a critical role in controlling cell and tissue homeostasis. With recombinant technology, human tTG became available for commercial use and the cost of testing decreased considerably.[41] Most hospital laboratories now measure the tTG antibody instead of the anti-endomysial antibody. The currently available serologic tests for the diagnosis of CD remain within the province of the specialized diagnostic laboratory. Sensitivity of antigen DGP for antibody IgA by using ELISA-based test for diagnosis combined ranged is 88 (74%–100)% and specificity 80 (70%–95)%. Similarly for sensitivity of antigen gliadin for antibody IgG by using ELISA-based test for diagnosis, having the combined range is 90 (80%–95)% and specificity 98 (95%–100)%.[42]

Advantages

tTG is the serological test of choice for the detection of CD.[43]

Operating procedure for transglutaminase

TTG is used to diagnose weather a child is having CD or not. For the test result to be accurate, it is necessary that the person undergoing the test must be on a gluten containing diet until test is done.[44] [Figure 2] represents various steps involved in tTG.[45]
Figure 2: Steps involved in transglutaminase

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Limitations of tissue transglutaminase autoantibody enzyme-linked immunosorbent-based assay [46],[47],[48]

  • ”False positives” for tTG have also been observed in patients with IBD, food allergy, irritable bowel syndrome, giardiasis, other intestinal infections and autoimmune disorders
  • These “false positives” cannot always be resolved by Endomysial antibodies (EmA) because the results of EmA testing are reliable only in laboratories skilled in immunoflourescent assays
  • TTG cannot detect CD in its early stage. CD People with less villous atrophy cannot be detected by tTG
  • The results of tTG depend on the laboratory setup and the person performing the test, so its result cannot be reliable always.


Anti-deamidated gliadin peptide antibody test

This is the latest generation serologic test. It does not offer any considerable advantage over measurement of the tTG antibody as the primary screening test; however, the IgG-based DGP antibody is slightly more sensitivity and specificity that is substantially better than IgG-based tTG antibody and should be considered the test of choice in patients with selective IgA deficiency. Moreover anti-DGP tests have largely replaced AGA testing.[49],[50]

Limitation of anti-deamidated gliadin peptide antibody test

  • Expensive
  • Rarely predicts disease when tissue tTG is normal.



  Biopsy Top


Earlier, when CD was recognized as a newly evolved disease, its diagnosis was exclusively based on the finding of villous atrophy during a small intestine biopsy. The European Society for Pediatric Gastroenterology, Hepatology and Nutrition (ESPGHN) published the initial biopsy-based criteria for the diagnosis of CD.[8],[11] These criteria required three biopsies over a period exceeding 1 year. In recent time, serologic tests are used only for screening purposes and not to confirm the diagnosis of CD owing to some limitations that has already been shown in the beginning of this article. The diagnosis should be taken seriously because it is not a trivial disorder. It is a lifelong condition, which requires a strict GFD along with all its challenges of cost, complexity, and social restrictions. The most relevant feature of the disease was histological change that made it a gold standard test for diagnosis.[51] The patient having CD needs to be followed regularly on a long-term basis for monitoring of associated complications and development of other autoimmune disorders.[46] Patients found positive after serological test results should be referred to a gastroenterologist for small intestinal biopsies to confirm the diagnosis of CD.[52],[53]

Despite substantial changes in the mode of presentation and the availability of new diagnostic tools, small bowel mucosal biopsy has remained the gold standard for CD diagnosis until now. Endoscopic small intestinal biopsies are safe and carry minimal morbidity.[52] However, timely availability of endoscopy remains a problem that requires attention. It is important to note that a GFD should not be started before the biopsy is performed, as such a diet will lead to healing of the small intestinal mucosa and complicate the interpretation of the biopsy results.[54] Some patient suffering from allergic reaction (chronic, severely itchy, blistering rash) known as dermatitis herpetiformis is “CD of the skin.”[55] In such patients, a skin biopsy can confirm the diagnosis and small intestinal biopsy is not required because most of these patients will have villous atrophy. The ESPGHN proposed a no biopsy approach to the diagnosis of CD in children.[56] These guidelines suggest that, for avoiding small intestinal biopsy, a symptomatic patient must fulfill the following criteria:

  • Patient have a positive tTG antibody test result more than 10 times the upper limit of normal,
  • Moreover have a positive endomysial antibody test result, and
  • Positive results for HLA-DQ2 or HLA-DQ8 testing.


If the patient meets all 3 criteria, the family might be offered the opportunity to decline a biopsy because CD is highly likely. It is important to note that just having a very high level of tTG antibodies is not enough to make a diagnosis of CD. Also, according to the diagnostic algorithm proposed in the guidelines, a patient with a positive tTG antibody test result should be referred by the primary care physician to a pediatric gastroenterologist for further assessment and workup.[56]

False-positive results in serologic tests are uncommon but can occur. Because CD is a chronic disorder with substantial lifelong implications, all attempts should be made to confirm the diagnosis. It is bad to miss a diagnosis of CD, but it is also harmful to diagnose it in someone who does not have it.[57],[58],[59]


  Limitations of Biopsy Top
[11],[46],[51],[52],[53],[54],[55],[56],[57],[58],[59]

  • Biopsy is different, difficult and invasive procedure
  • Facilities for biopsy are not available in medium class cities and need lot of expertise
  • For better results, at least four biopsy samples should be taken that results multifold puncturing of intestinal wall
  • Neglect in standardized operative procedures in many medical centers
  • Low degree of specificity.


With its low degree of specificity, histology fails to be the gold standard of CD diagnosis, and it is time to recognize that for the majority of gluten-sensitive cases, histology alone cannot provide the diagnosis.


  Operating Procedure for Biopsy Top


Biopsy is referred to as the confirmatory test in the diagnosis of CD. This test requires a lot of expertise and so is conducted by a gastroenterologist most often in an outpatient surgical department. The procedure takes around half an hour by inducing sedation through local anesthesia. Biopsy specimens are taken from the distal part of the duodenum upon endoscopy; in some small children they are taken from the proximal jejunum with the Watson capsule.[60] Specimens are then stained with h and e, studied under light microscopy, and classified according to Marsh. [Figure 3] represents the procedure followed during biopsy [61] and [Table 2] represents specificity and sensitivity ranges of various available diagnostic tools.
Figure 3: Operating procedure for Biopsy

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Table 2: Sensitivity and specificity range of various available diagnostic tools

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  Upcoming Diagnostic Tools for Celiac Disease Top


NonInvasive diagnostic methods

Biochemical estimation of celiac disease

In 2017, Oldenburger et al. performed a study to develop a biochemical method for the noninvasive diagnosis of CD. They on the basis of literature survey observed intestinal fatty acid–binding protein (I-FABP) as a potential marker in CD. It is a small (15 kDa), cytosolic, water-soluble protein which is released upon enterocyte damage. Previous studies showed that serum I-FABP is a marker of acute intestinal ischemia, Crohn's disease, sepsis, and necrotizing enterocolitis. I-FABP has also been examined as a marker in CD diagnostics by some researchers and is both found to be significantly elevated in CD patients and correlates with the percentage of mucosal damage. To establish this they selected a study population which included all children of (0–18 years) that were newly referred to the Wilhelmina Children's Hospital and were having tTG level < 10x of Upper Limit of Normal (ULN) and also were genetically predisposed to CD genes. A highly specific commercially available ELISA kit capable of selectively detecting human I-FABP provided by HBT (Uden, the Netherlands) was used by them to determine serum I-FABP levels. The results observed by them showed that I-FABP level was significantly increased in patients with histologically proven CD when compared with other healthy controls. However diagnostic performance of the results used alone was not sufficient. They further suggested that a combination of I-FABP and tTG may increase the proportion of patients that can be diagnosed by this noninvasive diagnostic technique.[66]

Adriaanse et al. in 2017 performed a study to find out the usefulness of plasma I-FABP in the noninvasive diagnosis of CD. There results demonstrated that a higher plasma I-FABP level, increased CD autoantibody titres and HLA-DQ2 and/or-DQ8 might be sufficient for the diagnosis of CD and could increase the number of patients that can be benefited from a noninvasive approach. Moreover, I-FABP levels recover rapidly after gluten elimination from the diet of CD patients, which shows that plasma I-FABP may also be used as a monthly monitoring tool for disease activity in CD patients. This study also confirms the study of Oldenburger as the results observed in both the tests are almost similar.[67]

Determination of Bacterial composition of fecal microbiota as a tool for the diagnosis of celiac disease

Di Cagno et al. in 2009 performed a study to determine the concentration of fecal microbiota and volatile organic compounds in treated and un-treated CD patients. For this they selected study groups of children b/w 6-12 years of age having symptoms of CD, healthy children and children on GFD. He compared the microbiota composition of each group by collecting stool samples of the study population, extraction of DNA from those samples and subjected the samples to polymerase chain reaction amplification and denaturing gradient gel electrophoresis. Later they performed gene sequencing to identify the specific microbial composition. He also performed gas chromatography-mass spectrometry-solid-phase micro extraction analysis of fecal volatile organic compounds. There results demonstrated that, fecal microbiota and volatile organic compound concentrations were different in healthy, treated and untreated celiac children. The microbiota and volatile organic compounds of treated and healthy children was almost similar while different in untreated CD patient. This study if further carried out can help in development of a microbial map for the noninvasive diagnosis of CD.[68]

Video capsule endoscopy as a diagnostic tool for celiac disease

Rokkas et al. in 2012 performed a meta-analysis in order to determine the usefulness of video capsule endoscopy as a diagnostic tool for CD. He performed an extensive literature survey of the already performed experiments and on the basis of that extracted the data to demonstrate the effectiveness of video capsule endoscopy in the diagnosis of CD. The results of his study demonstrated that video capsule endoscopy (VCE) is not as accurate as pathology, but instead it can be a reasonable alternative tool for diagnosing CD. This method is capable of expanding the portfolio of available diagnostic methods, especially for the patients who are unwilling to undergo gastroscopy due to its perceived inconvenience and discomfort. However, there is no evident larger, multicenter, and well-designed trials yet which could fully demonstrate its usefulness in diagnosis of CD. He and the team concluded that larger, multicenter, and well-designed trials are needed to further establish the role of VCE in the diagnosis of CD.[69],[70]

The role of human leukocyte antigen typing in diagnosis of celiac disease

More than 97% of those with CD have the DQ2 and / or DQ8 marker genes. As many as 25%–40% of Caucasians carry the genotypes that encode for the HLA molecules DQ2 or DQ8, so a positive test has very poor positive predictive value. However, there are patients in whom serology and biopsy are equivocal yet in whom there is a strong suspicion that CD is there, HLA typing can be utilized there for the diagnosis. After typing, if the patient lacks the risk haplotypes (DQ2/DQ8), then they can be excluded from the risk of having CD. DNA-based methods are now used for HLA typing and may report the genotypes that encode the old serotype equivalents DQA1 * 05 with DQB1 * 02 (encodes DQ2) and DQA1 * 03 with DQB1 * 0302 (encodes DQ8). It should be noted that HLA typing is often performed in a central laboratory and is not always readily available and this is the biggest demerit of this diagnostic technique.[71]


  Step-by-Step Diagnostic Approach for Actual Diagnosis of Celiac Disease Top


The major problem with CD is that people do not follow the proper diagnostic approach leading to false diagnosis as well as prevalence of CD. For proper diagnosis, first, the age group, gender, and area at risk should be identified followed by identification of associated symptoms.[72],[73],[74] The symptoms for CD vary from person to person as CD may either be symptomatic or asymptomatic. In some patients, there may be GI symptoms while in others there may be no associated symptoms at all. Hence, in order to diagnose CD accurately, step-by-step diagnostic approach should be followed.

IgA-TTG tissue tTG test is recommended for initial testing of CD.[52],[70] Endomysial antibody (EMA) test can also be used as an alternative of IgA-TTG if IgA-tTG is unavailable but the test is having greater specificity and lower sensitivity.[75] Individuals with CD who are also IgA deficient will not have abnormally elevated levels of TTG IgA or EMA IgA. The occurrence of both CD and IgA deficiency in the same individual appears to be rare in asymptomatic individuals (approximately 1:8500 of the general population) but is more likely in symptomatic children with CD (approximately 2%). Therefore, when testing for CD in children with symptoms suspicious for CD, measurement of quantitative serum IgA can facilitate interpretation when the TTG IgA is low. In individuals with known selective IgA deficiency and symptoms suggestive of CD, testing with TTG IgG is recommended.[75],[76] Even when serological tests for CD are negative, in children with chronic diarrhea or FTT and in those belonging to a group at risk (e.g., selective IgA deficiency or a positive family history of CD) who have symptoms compatible with CD, an intestinal biopsy can be helpful to identify the unusual case of seronegative CD or to detect other mucosal disorders accounting for the symptoms.[77] It has been recommended that confirmation of the diagnosis of CD requires an intestinal biopsy in all cases. Because the histological changes in CD may be patchy, it has been recommended that multiple biopsy specimens be obtained from the second or more distal part of the duodenum. There is good evidence that villous atrophy (Marsh type 3) is a characteristic histo-pathologic feature of CD. The presence of infiltrative changes with crypt hyperplasia (Marsh type 2) on intestinal biopsy is compatible with CD but with less clear evidence.[78],[79] Diagnosis in these cases is strengthened by the presence of positive serological tests (TTG or EMA) for CD. In the event the serological tests are negative, other conditions for the intestinal changes are to be considered and if excluded, the diagnosis of CD is reconsidered. The presence of infiltrative changes alone (Marsh type 1) on intestinal biopsy is not specific for CD in children. Concomitant positive serological tests for CD (TTG or EMA) increase the likelihood that an individual has CD. In circumstances where the diagnosis is uncertain additional strategies, including determination of the HLA type, repeat biopsy or a trial of treatment with a GFD, repeat serology and biopsy can be considered. The diagnosis of CD is considered definitive when there is complete symptom resolution after treatment with a strict GFD in a previously symptomatic individual with characteristic histological changes on small intestinal biopsy. A positive serological test that reverts to negative after treatment with a strict GFD in such cases is further supportive evidence for the diagnosis of CD.[80] [Figure 4] represents the diagnostic approach that needs to be followed for the proper diagnosis of CD.
Figure 4: Representation of systematic diagnostic approach

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The approach described here is the accurate/appropriate approach for confirmation of the disease but in regular practice people only go for serological testing and on the basis of obtained results they start following GFD. This gives rough estimation of prevalence of CD as well as improper diagnosis of the disease. CD can finally be confirmed only after undergoing biopsy testing. Once the obtained result of biopsy testing is positive, only then a strict GFD should be taken.


  Conclusion Top


Serology is widely available and predominantly performed in primary health care for diagnosis and prognosis of diseases. Diagnostic tools for CD evolved from diagnosis based on primitive symptomatic features to latest sequencing techniques in last 70 years. It has been observed in a recent study in Italy that when diagnosis of CD is based only on serology then there are chances of two third of the incorrect cases. At the same time some other studies showed that over reliance on serological markers may lead to underdiagnosis of CD. The data in the compilation reflects that although there is a great change and evolution in diagnostic techniques yet there is a need of developing noninvasive, specific and sensitive diagnostic tests for diagnosis of CD. There is a great need and potential in developing microbiome and micronome-based methods that could provide a sure shot picture of the disease and enable the physicians to take early informed decisions.

Financial support and sponsorship

DST-SYST (Project No- SP/YO/097/2017.

Conflicts of interest

There are no conflicts of interest.



 
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