In a recent study published in Gastroenterology, researchers presented intestinal microbiota modulation as a probable therapeutic option for diseases such as Crohn’s disease (CD).
Studies have reported that dysbiosis or imbalance in the intestinal microbiome could promote degenerative disorders, including diabetes, obesity, cardiovascular illness, cancer, inflammatory bowel disease, and hepatic illness. Thus, probiotic microbes, that benefit the host, could provide an option for improving disease outcomes by restoring the intestinal microbial balance.
About the study
In the present study, researchers investigated the therapeutic effects of modulating the intestinal microbiome using probiotics in diseases such as CD.
The team characterized the intestinal microbiota, comprising fungi, bacteria, and their NCDR (non-diseased first-degree relatives) among CD patients. Stool specimens were obtained from the study participants, following which, bacterial deoxyribonucleic acid (DNA) and fungal DNA were extracted and subjected to next-generation sequencing. Subsequently, PCA (principal component analysis), abundance analysis, and diversity analysis were performed.
In vitro analysis of the biofilm-forming ability of the microbes, by self or by interacting with one another, was performed. Confocal microscopy and electron microscopy (transmission electron microscopy and scanning electron microscopy) were performed. To identify beneficial microbes that could disrupt biofilms that were formed by pathogenic fungi and bacteria, correlation analysis was performed, as a result of which, four therapeutic strains, i.e., Lactobacillus acidophilus 16axg, Bifidobacterium breve 19bx, Saccharomyces boulardii 16mxg, and Lactobacillus rhamnosus 18fx were analyzed further.
The strains interfered with epithelial injury caused by S. marcescens, C. tropicalis, and E. coli. Additionally, the amylase enzyme was added to the candidate formulation for improved biofilm disruption. Further, validation analyses were performed (phase 3) by evaluating the effects of the probiotic combination in preventing/disrupting pathogenic biofilms (phase 3a).
Subsequently, in vivo preclinical analysis was performed to assess the effects of the probiotic candidate among spontaneous chronic Crohn’s disease-like-Ileitis (SAMP) mice. The mice were divided into three groups, one group received the probiotic candidate (with amylase) daily over 56.0 days, the second group received the candidate formulation lacking amylase, and the third group received only phosphate-buffered saline. Subsequently, mice ileac tissues were subjected to histological analysis.
In addition, fecal specimens were analyzed by GC/MS (16S ribosomal ribonucleic acid and gas chromatography/mass spectrometry) analyses. Lastly, the formulation was administered to 46 human volunteers (phase 4) once daily for four weeks, and assessments were performed four weeks post-administration. The microbiota of the volunteers was compared to those documented by HMP (human microbiome project).
Compared to healthy individuals, CD patients showed lowered Bacteroidetes and Faecalibacterium prausnitzii counts, and greater counts of Ruminococcus gnavus, E. coli, S. marcescens, and C.tropicals resulting in inflammation and disruption of the mucus layer. S. marcescens, C. tropicalis, and E. coli interacted with one another to form Candida-specific biofilms of elevated thickness and mass. E. coli appeared to be fused with the cellular wall of C.tropicalis, whereas S. marcescens interacted with C. tropicalis and E. coli with the help of fimbria.
The biofilm-forming abilities of the three microbes (E. coli, S. marcescens, and C.tropicals) were confirmed in the in vivo experiments. The candidate probiotic formulation could prevent biofilm formation and effectively disrupt the pathogenic biofilms, with significantly reduced germ tube formation by C. albicans, indicating lowered virulence of the fungal organism.
Mice receiving the candidate probiotic (with amylase) showed significantly less severe ileitis than the other groups. The GC/MS and sequencing analysis findings indicated an enhanced abundance of Lachnoclostridium species and Mucispirillum schaedleri species among probiotic (and amylase)-treated mice, related to short-chain fatty acid production. Further, elevated expression of 27.0 genes that contribute to the development of B memory lymphocytes and T lymphocyte infiltration and reduced expression of 17.0 genes were observed among the probiotic (with amylase enzyme)-treated murine animals.
The microbial strains in the candidate formulation were GRAS (generally regarded as safe) microorganisms. Among human volunteers, the probiotic significantly lowered Candida and Bacteroidetes species counts and increased Firmicutes counts. The lowered Candida counts could aid in preventing biofilm formation and associated inflammation in Crohn’s disease.
Overall, the study findings showed that the candidate probiotic formulation caused functional alterations resulting in the amelioration of Crohn’s disease-like ileitis severity. Additionally, amylase was critical in reducing intestinal inflammation. The probiotic-amylase combined therapy could reverse the Firmicutes/Bacteroidetes ratio and restore intestinal microbial balance. The findings indicated that altering the intestinal microbiome composition using probiotic microbes could regulate dysbiosis, providing a viable treatment approach for diseases with underlying microbial imbalances.