Investigating the correlation between gut microbiota levels and coeliac disease in the Maltese population

Abstract

Coeliac Disease (CD) is an immune-mediated enteropathy triggered by gluten consumption in genetically predisposed individuals. In recent years, gut microbiota has emerged as a contributing factor in the development and progression of CD, particularly through the potential role of probiotic microorganisms in maintaining gut health. This study sought to investigate the presence and abundance of two probiotic microorganisms, Lactobacillus acidophilus and Saccharomyces boulardii, in coeliac and healthy control individuals, that is, individuals without CD, though not necessarily free of other diseases, within the Maltese population. It explored whether there is any correlation between differences in gut microbiota levels and CD status. A cross-sectional study was conducted involving 65 participants (15 coeliac, 50 control participants). Participants submitted a one-time faecal sample and completed a health-related questionnaire. S. boulardii was assessed using culture-based methods on Candida Chromogenic Agar (CCA) followed by MALDI-TOF for species confirmation. L. acidophilus detection involved manual DNA extraction from faecal samples, with quantitative PCR (qPCR) performed using species-specific primers. A reference strain (ATCC4356) was used for validation. Statistical analyses were performed using SPSS version 29. None of the samples showed the presence of S. boulardii, despite its growth on the CCA media used as a control, possibly due to its transient presence in the gut or limitations in culture recovery. Candida albicans was the most commonly identified yeast in both groups. DNA extracted from faecal samples showed visible amplification products on agarose gel electrophoresis, indicating the presence of both human and bacterial DNA. However qPCR results for L. acidophilus showed irregularities, suggesting that further optimisation, particularly in primer specificity and reaction conditions, is required. While this study did not find statistically significant microbial differences between CD and non-CD groups, trends, such as the predominance of C. albicans and the absence of S. boulardii, raise important questions about yeast dynamics in CD. Methodological limitations such as the small sample size, use of a single reference strain, and control group heterogeneity may have impacted findings. Future research with larger, longitudinal cohorts and high-throughput sequencing such as 18 rRNA gene analysis is guaranteed to elucidate the gut microbiota’s role in CD pathogenesis.