A Potential Probiotic for Diarrhea: Clostridium tyrobutyricum Protects Against LPS-Induced Epithelial Dysfunction via IL-22 Produced By Th17 Cells in the Ileum

Probiotics are clinically used for diarrhea and inflammatory bowel diseases in both humans and animals. Previous studies have shown that Clostridium tyrobutyricum (Ct) protects against intestinal dysfunction, while its regulatory function in the gut needs further investigation and the related mechanisms are still not fully elucidated. This study aims to further verify the protective function of Ct and reveal its underlying mechanisms in alleviating diarrhea and intestinal inflammation. Ct inhibited LPS-induced diarrhea and intestinal inflammation in the ileum. IL-22 was identified and the protective role of Ct in the ileum presented an IL-22-dependent manner according to the transcriptomic analysis and in vivo interference mice experiments. The flow cytometric analysis of immune cells in the ileum showed that Ct enhanced the proportions of Th17 cells in response to LPS. The results of in situ hybridization further verified that Ct triggered Th17 cells to produce IL-22, which combined with IL-22RA1 expressed in the epithelial cells. Moreover, Ct was unable to enhance the levels of short-chain fatty acids (SCFAs) in the ileum, suggesting that the protective role of Ct in the ileum was independent of SCFAs. This study uncovered the role of Ct in alleviating diarrhea and inflammation with the mechanism of stimulating Th17 cells in the lamina propria to produce IL-22, highlighting its potential application as a probiotic for diarrhea and inflammation in the ileum.

Optimization of Clostridium tyrobutyricum encapsulation by extrusion method and characterization of the formulation

Purpose: To optimize the process parameters for the encapsulation of Clostridium tyrobutyricum (Ct) and to determine its in vitro characteristics.Methods: The process parameters, including the concentration of the wall and hardening material, Ct to gelatin ratio and hardening time, were studied by single factor analysis, while optimization was performed by orthogonal experimental design for the encapsulation rate of Ct.Results: Optimal conditions exhibited by orthogonal experimental design at a 92.17 % encapsulation rate with a viable count of 9.61 ± 0.06 lgCFU/g were: 6 % modified starch, 3 % sodium alginate, and 2 % CaCl2 at a Ct to gelatin ratio of 1:1 with a hardening time of 30 min. The survival rates of encapsulated Ct were higher than free Ct in simulated gastric (6.22 %) and intestinal juices (15.55 %). Reduction in viable counts of Ct at 90 °C were higher for free cells (44.76 %) than encapsulated cells (28.09 %) after 30 min of heat treatment. Correspondingly, encapsulation boosted the capacity of Ct to withstand the strong acidic conditions of the stomach and improved the storage properties of Ct.Conclusion: The results suggested that extrusion is a good technique for the encapsulation of Ct, as it enhances the viability of Ct during their transit through the gastrointestinal tract. Furthermore, encapsulation is favorable for Ct if planned for use in formulations where high temperature treatment is required.

Draft Genome Sequences of 12 Clostridium tyrobutyricum Strains Isolated from Raw Milk and Cheese

Clostridium tyrobutyricum is recognized as the main causative agent of late blowing defect—severe spoilage of hard and semihard cheeses. In this work, we present the draft genome sequences of 12 C. tyrobutyricum strains isolated from raw milk and cheese.

Colorimetric Point-of-Care Detection of Clostridium tyrobutyricum Spores in Milk Samples

Clostridium tyrobutyricum represents the main spoiling agent responsible for late blowing defects (LBD) in hard and semi-hard cheeses. Its spores are resistant to manufacturing procedures and can germinate during the long ripening process, causing the burst of the cheese paste with a consequent undesirable taste. The lower quality of blown cheeses leads to considerable financial losses for the producers. The early identification of spore contaminations in raw milk samples thus assumes a pivotal role in industrial quality control. Herein, we developed a point of care (POC) testing method for the sensitive detection of C. tyrobutyricum in milk samples, combining fast DNA extraction (with no purification steps) with a robust colorimetric loop-mediated isothermal amplification (LAMP) technique. Our approach allows for the sensitive and specific detection of C. tyrobutyricum spores (limit of detection, LoD: ~2 spores/mL), with the advantage of a clear naked-eye visualization of the results and a potential semi-quantitative discrimination of the contamination level. In addition, we demonstrated the feasibility of this strategy using a portable battery-operated device that allowed both DNA extraction and amplification steps, proving its potential for on-site quality control applications without the requirement of sophisticated instrumentation and trained personnel.