Chronic diarrhea is a frequent clinical issue that can be seen in outpatient and inpatient clinic and is considered one of the major morbidity issues globally. It is typically considered to be a duration of loose or watery stools over four weeks long, which can result due to a long range of etiologies, including both functional disorders and severe organic pathologies [1]. The unremitting diarrhea does not only impact the nutritional status and quality of life but also leads to a heightened usage of healthcare and economic impact [2]. Chronic diarrhea is considered to be a complicated diagnosis due to the possibility of clinical manifestation overlap between infectious, inflammatory, malabsorptive, and neoplastic diseases. As a result, the correct diagnosis of the underlying cause is critical to the correct management and prevention of complications [3].

Gastrointestinal (GI) biopsies under histopathological analysis are a central part of diagnostic work up of chronic diarrhea. Endoscopic biopsy permits close evaluation of the mucosal architecture, inflammatory patterns, epithelial integrity, and cellular infiltration which are useful in addition to clinical and laboratory results [4]. Histological analyses can be used in the diagnosis of inflammatory bowel disease, microscopic colitis, celiac disease, intestinal tuberculosis, parasitic infestations, and neoplastic or pre-neoplastic lesions. It is also used to be able to distinguish between infectious and non-infectious causes in case gross endoscopic results are inconclusive [5]. Thus, histopathology could serve as one of the pillars of making conclusive diagnoses and informative therapeutic decision-making in patients presenting with a long-term history of diarrheal symptoms [6].

Histopathological observations were found to be diverse and varied in each geographical location as a result of changes in environmental exposure, sanitation, diets, and the presence of infectious agents. Infectious and parasitic etiologies are common in developing countries, such as Pakistan, whereas inflammatory and autoimmune diseases are gaining more and more popularity as a result of shifting lifestyles and the better access to diagnosis [7, 8]. Although the overall endoscopic services are increasing in number, few regional data has been documented to explain the distribution and pattern of histopathological abnormalities of GI biopsies of patients presenting with chronic diarrhea. This information plays a crucial role in helping the clinicians and pathologists to interpret the local trends of the disease and maximize the diagnostic approaches [9].

Despite the international literature on the heterogeneity of histological aspects in chronic diarrhea, there exist gaps in the literature to provide comprehensive, locally determined evidence to evaluate biopsy-based findings in chronic diarrhea and align with its clinical presentation. The inability to create context-specific diagnostic algorithms due to this gap in region-specific data can also result in the under-identification of some pathological conditions. Thus, the current research paper will fill this gap by assessing the histopathological characteristics of gastrointestinal biopsies collected in the participants of chronic diarrhea in Pakistan.

The aim of this study is to establish the range and frequency of histopathological observation among such patients and thus lead to better diagnostic understanding and clinical management among the population in the area.

Study Design and Setting

This paper was done on a descriptive cross-sectional research at the Department of Histopathology with the Gastroenterology Unit of an academic teaching hospital in Pakistan. The trial was an assessment of histopathological findings of gastropod intestinal biopsies of clients who presented with chronic diarrhea. The data was collected in the twelve months, i.e., January 2025 to December 2025.

Sample Size Determination

To compute the sample size, the formula of single population proportion was used in cases of descriptive research:

:n=Z2⋅p(1−p)/ d2

N was the required sample size, Z was the standard deviation of the normal distribution of the 95-percent confidence level (1.96), p was the anticipated prevalence of the significant histopathological abnormalities in patients with chronic diarrhea, and d was the margin of error. Because of the lack of local prevalence data, a prevalence of 50% was used to guarantee the maximum sample size and the level of precision was fixed at 8%. This computation gave a minimum of 150 participants. In order to consider all unfinished samples of biopsy and gaps in data, 10% was added which made the final sample size 165 patients who were included in the study.

Sampling Technique

Non-probability consecutive sampling method was used. All patients with chronic diarrhea, who were eligible and had gastrointestinal endoscopy study to evaluate their problems, were recruited until the necessary sample was obtained during the period of the study.

Study Population and Criteria

The population of the study was patients of both sexes aged 18 years and older who reported a chronic diarrhea of greater than 4 weeks and had undergone gastrointestinal biopsy as one of their diagnostic tests. The inclusion criteria were that patients had relentless diarrhea with or without other gastrointestinal tract symptoms like abdominal pain, bloating, or wasting, and gave an informed consent that their clinical data and the result of the biopsy can be used in the research. Patients were also excluded in case they had insufficient or in poor conditions of the biopsies or had a previous diagnosis of gastrointestinal malignancy that was still under treatment, previous gastrointestinal surgery that had changed the mucosal anatomy, and lack of full clinical records. Moreover, patients who had systemic illnesses that might cause diarrhea on their own, including uncontrolled hyperthyroidism or chronic kidney disease were also excluded to make the confounding factors as minimal as possible.

Data Collection Procedure

Medical records were used to retrieve clinical and demographic data such as age, gender, duration of symptoms and any clinical history using a structured proforma. Samples of endoscopic biopsy were taken on pertinent gastrointestinal sites based on clinical finding. The specimens were fixed in 10 percent buffered formalin and subjected to the routine paraffin embedding procedures and stained with hematoxylin and eosin. Special stains were used as necessary to identify particular diagnoses.

The histopathological study involved highly qualified consultant histopathologists who evaluated the mucosal architecture, presence of inflammatory infiltrates, epithelial changes and the presence of infectious or neoplastic changes. The results were grouped into diagnostic categories; nonspecific inflammation, inflammatory bowel disease, microscopic colitis, celiac disease, infectious pathology, and neoplastic lesions.

Variables and Outcome Measures

The spectrum and frequency of histopathological diagnoses in gastrointestinal biopsies was the main outcome measure. The independent variables were demographic and clinical presentation. The patterns of histology and diagnostic categories were put in records to be analyzed.

Statistical Analysis

Statistical Package of Social Sciences (SPSS) 25 was used to enter and analyze data. Mean and standard deviation were provided in quantitative variables like age whereas frequencies and percentages were used to describe qualitative variables like gender and histopathological categories. The Chi-square test or the Fisher exact test were used to determine the relationship between categorical variables where they were appropriate. Where needed, independent sample t-tests or one-way ANOVA were used to compare the mean values in different groups. A p-value of [?]0.05 was taken to be statistically 

The study involved 165 patients who had persistent diarrhea. The average age of the study population was 42.5 ± 15.3 years, and the age of the subjects was between 18 and 78 years. The number of the participants was 92 (55.8) boys and 73 (44.2) girls. The average time of diarrhea was 7.2 ±  3.8 weeks. Other symptomatology recorded were abdominal pain in 102 (61.8%) patients, bloating in 78 (47.3%) patients and accidental weight loss in 41 (24.8%) patients.

Table 1: Demographic and Clinical Characteristics of Study Participants

Gastrointestinal biopsies were examined by the use of histopathology and a wide range of results were observed. The most prevalent abnormality was nonspecific inflammation and was detected in 61 patients (37.0%). The 32 patients (19.4%), who were diagnosed with inflammatory bowel disease (IBD) included 20 cases of ulcerative colitis and 12 cases of Crohn. Microscopic colitis such as lymphocytic and collagenous colitis was detected in 14 patients (8.5%). The diagnosis of celiac disease was made in 18 participants (10.9) on the basis of the villous atrophy and intraepithelial lymphocytosis. There were 21 patients with infectious etiologies (12.7%), parasitic infestations, and chronic bacterial infection. 11 patients (6.7%), had neoplastic (or pre-neoplastic) lesions, such as adenomatous polyps and early gastrointestinal malignancies. Normality was seen in 8 patients (4.8 per cent).

The patients were divided into three age groups, i.e., 18-35 years (n = 56), 36-55 years (n = 62), and >55 years (n = 47). The patterns showing the distribution of histopathological findings were different among age groups (Chi-square test, χ²  = 18.7, p = 0.002). Nonspecific inflammation was more typical in younger patients (18-35 years: 25.0%), IBD and neoplastic lesions were mostly detected among patients aged 36-55 years and >55 years, respectively. The younger sample identified celiac disease higher.

Table 2: Age-wise Distribution of Histopathological Findings

The correlation between histopathological diagnoses and gender was studied. In males, nonspecific inflammation was identified in 38 patients (41.3%), whereas in females, it was seen in 23 patients (31.5%). IBD showed a male predominance with 21 cases (22.8%) in males compared to 11 cases (15.1%) in females. Microscopic colitis was more frequent in females (9 cases, 12.3%) than males (5 cases, 5.4%). Celiac disease showed a slight female predominance (10 cases, 13.7% vs 8 cases, 8.7%). The association between histopathological category and gender was not statistically significant (Chi-square test, χ² = 7.84, p = 0.098).

Table 3: Gender-wise Distribution of Histopathological Findings

The colon (n = 112, 67.9%), duodenum (n = 39, 23.6%) and stomach (n = 14, 8.5%) were the locations where most of the biopsies were taken. Nonspecific inflammation (42 patients, 37.5%) and IBD (30 patients, 26.8%) were mainly found in the colonic biopsies. Celiac disease was found in 16 cases (41.0%) in duodenal biopsies and infectious gastritis was found in 6 cases (42.9%) in gastric biopsies. The biopsy site and histopathological diagnosis were statistically related (Chi-square test, χ²  = 35.6, p < 0.001).

Table 4: Biopsy Site-wise Distribution of Histopathological Findings

Clinical symptom analysis combined with histopathological diagnosis showed that patients who presented with weight loss had more chances of having IBD or neoplastic lesions (p = 0.021, Chi-square test). IBD and microscopic colitis were linked to abdominal pain (p = 0.032), whereas bloating was observed more frequently in the case of celiac disease (p = 0.045). These observations are evidence that certain pattern of symptoms could give hints to the underlying histopathological pathology, but these phenomena overlapped with each other in most instances.

Table 5: Correlation of Clinical Symptoms with Histopathological Diagnosis

Gastrointestinal biopsies of patients with chronic diarrhea exposed a broad range of abnormal histopathological changes in this research. The most common finding was nonspecific inflammation, then inflammatory bowel disease, infectious pathology, celiac disease, microscopic colitis and neoplastic lesions. This distribution shows the predominance of inflammatory and infectious etiologies in patients with chronic diarrhea and functional or normal histology was relatively rare. This has been supported by the increased incidence of non-specific inflammation implying that a significant percentage of chronic diarrhea cases could be attributed to mild or subclinical mucosal alterations that have not yet been categorized as specific diseases.

According to the age-wise analysis, younger patients had greater chances of having celiac disease and nonspecific inflammation, whereas middle-aged and older patients displayed more rates of IBD and neoplastic lesions [10]. Not statistically significant gender distribution differences were observed, however, microscopic colitis and celiac disease were slightly more common in females. Biopsy site and histopathology correlation established that site-specific analysis is essential to the accurate diagnosis of celiac disease and IBD and nonspecific inflammation is mostly detected in colonic biopsies. Correlation of the symptoms indicated that the weight loss and abdominal pains were likely to predict more severe underlying pathology, such as IBD and neoplastic lesions, compared with bloating, which had the correlation with celiac disease [11].

In comparison with the available literature, the prevalence of nonspecific inflammation coincides with the results published on similar populations of patients in developing countries, where environmental and infectious pathogenesis play a major role in the chronic diarrhea [12]. The presented prevalence of IBD correlates with the emerging data about the rising number of autoimmune gastrointestinal disorders in the area, and the finding of celiac disease confirms the tendency in the world in terms of the improved recognition of the diseases as a result of the increased access to their diagnostics. The prevalence of infectious etiologies highlights the chronicity of parasitic and bacterial infections in persistent diarrhea in the endemic gastrointestinal pathogen areas [13]. The comparatively lower number of neoplastic lesions is due to the age distribution of the study population, and is consistent with the previous results that malignancy is a relatively less common but clinically relevant cause of chronic diarrhea [14, 15].

Although it was a valuable study, there were a number of limitations. As a single center cross-sectional research, the results might not be generalized to the full population of Pakistan. The sample is also sufficient to carry out descriptive analysis, but will not permit a large amount of subgroup analysis and follow-up data on clinical outcome was unavailable. Also, nonspecific inflammation (which can contain early or evolving disease) was found to be considered as a type of biopsy.

Further research can be based on multicenter designs with increased numbers and longitudinal follow-ups to check the progression of the histopathological abnormalities. The use of molecular diagnostics, immunohistochemistry, and microbiological testing can be used to clarify additional information about the nonspecific inflammatory alterations and facilitate an early identification of the autoimmune or neoplastic states. Moreover, the comparison of the biopsy results with the nutritional, environmental, and genetic factors may enhance learning about the patterns of diarrhea chronicity in the region and allow creating specific diagnostic algorithms.

This paper has shown that a broad range of histopathological abnormalities is linked to chronic diarrhea in Pakistani patients. The most prevalent was nonspecific inflammation, then inflammatory bowel disease, infectious pathology, celiac disease, microscopic colitis and neoplastic lesions. The pattern of histopathology was affected by age and site of biopsy, but not by gender. There were clinical symptoms like the loss of weight and pain in the abdomen that were associated with more severe pathology. The findings reiterate the role of gastrointestinal biopsy in making a final diagnosis and treatment in managing chronic diarrhea especially where the etiology varies.

1. Yasin MA, Farooq MA, Uddin N, Parveen B, Asghar A, Ahsan J. Spectrum of histopathological patterns of duodenal biopsies in patients with unexplained anaemia and chronic diarrhea and their correlation with endoscopic findings and serological markers. Pakistan Armed Forces Medical Journal. 2021 Oct 31;71(5):1893-6.
2. Manzoor M, Maqbool M, Sarwar A, Khan M, Mir H, Batool B, Bangash SA, Muthanna FM. Adults with celiac disease: histopathological and immunohistochemical analysis of small intestinal biopsies: histopathological and immunohistochemical analysis of small intestinal biopsies. Pakistan BioMedical Journal. 2022 Jan 31:249-52.
3. Ullah O, Rahman H, Ijaz S. Helicobacter pylori-Associated Infection: A Comprehensive Histopathological Analysis of Gastric Biopsies from Patients of Pakistan. Microbiology Research. 2025 Nov 2;16(11):232.
4. Muhammad N, Rahat A, Jafri AA. Frequency of microscopic colitis in patients presenting with chronic diarrhea. The Professional Medical Journal. 2024 Mar 4;31(03):422-6.
5. Ahmad Z, Siddique SU. Histopathology of the gastrointestinal tract. InAtlas of Diagnostic and Predictive Histopathology 2020 Jun 2 (pp. 205-237). Singapore: Springer Singapore.
6. Zubair M, Mubeen F, Farooq MA, Hanif A, Rana MZ, Khan S. Spectrum of Duodenal & Small Intestinal Diseases in Quetta: A Hospital-Based Study. Annals of PIMS-Shaheed Zulfiqar Ali Bhutto Medical University. 2025 Dec 20;21(4):716-21.
7. Khan B, Khan MS, Naeem A, Naim F, Utmani N. Frequency of TTG positivity in diarrhea predominant IBS patients at Peshawar, Pakistan: TTG positivity in diarrhea predominant IBS patients. Pakistan Journal of Health Sciences. 2024 Oct 31:74-8.
8. Shaikh SS, Khalid R, Bukhari U, Shaikh H, Rizvi SA, Shabbir A. Frequency and risk factors of microscopic colitis as a cause of chronic watery diarrhoea. Journal of Ayub Medical College Abbottabad. 2023 Jul 9;35(3):394-9.
9. Liu TC, VanBuskirk K, Ali SA, Kelly MP, Holtz LR, Yilmaz OH, Sadiq K, Iqbal N, Amadi B, Syed S, Ahmed T. A novel histological index for evaluation of environmental enteric dysfunction identifies geographic-specific features of enteropathy among children with suboptimal growth. PLoS neglected tropical diseases. 2020 Jan 13;14(1):e0007975.
10. Shaikh SS, Shaikh H, Ali SA, Khalid R, Shabbir A, Afnan BH. Clinical, endoscopic, and histopathological characteristics of gastrointestinal polyps. Journal of University Medical & Dental College. 2023 Apr 18;14(2):590-5.
11. Lail RA, Tahir QA, Sabir SH, Siraj SA, Khan TM, Parveen S. Histopathological Features of Helicobacter Pylori Positive and Helicobacter Pylori Negative Gastritis in Endoscopic Biopsies: First Comparative Study from Sahiwal, Pakistan. Pakistan J Medical & Health Sciences. 2022;6(1):303-5.
12. Anwar A, Cheema HA, Saeed A. SEROLOGICAL VERUS HISTOLOGICAL DIAGNOSIS IN PEDIATRIC CELIAC DISEASE: IS THERE A NEED FOR SMALL BOWEL BIOPSY?. Pakistan Armed Forces Medical Journal. 2021 Feb 1;71(1).
13. Khassaf MB, Qasim BJ. Histopathological assessment of colonoscopic biopsies in patients with bleeding per rectum. Medical Journal of Babylon. 2022 Apr 1;19(2):203-9.
14. Moussa F, Abd El Gawad W, Nosseir NS, Hassan M. Colonoscopic and histopathological findings in patients with various lower gastrointestinal symptoms: a single-center experience. Suez Canal University Medical Journal. 2020 Mar 1;23(1):62-70.
15. Gul R, Shah SA, Alam S, Khan AM, Khan S. PREVALENCE AND HISTOPATHOLOGICAL PATTERN OF HELICOBACTER PYLORI IN PATIENTS PRESENTING WITH GASTROINTESTINAL SYMPTOMS: A CLINICAL STUDY.: JPUMHS; 2024: 14: 04, 32-40. http://doi. org/10.46536/jpumhs/2024/14.04. 558. Journal of Peoples University of Medical & Health Sciences Nawabshah.(JPUMHS). 2024 Dec 31;14(4):32-40.