Exploring the effects of transport duration on the fecal microbial communities of surplus dairy calves.

Transportation significantly affects the health and welfare of surplus dairy calves, largely due to the various stressors and pathogen exposures encountered during the process. Concurrently, an animal's microbiome is known to correlate with its health status, with stress-induced alterations in the microbiota potentially precipitating various diseases. This study aimed to compare the effects of transportation durations of 6, 12, or 16 h on the fecal microbiota in young surplus dairy calves. We used a randomized controlled design in which surplus dairy calves aged 1 to 19 d from 5 commercial dairy farms in Ontario were allocated into 1 of 3 transportation groups (6, 12, and 16 h of continuous transportation). Health assessments were conducted before, immediately after, and for 2 wk following transportation. Fecal samples were collected before, immediately after, and at 24 and 72 h after transportation and subjected to 16S rRNA sequencing. Alpha diversity metrics showed no significant differences between the 3 transportation groups at any of the sampling time points. Although ß diversity metrics revealed no clustering by transportation groups, they indicated significant differences across sampling time points within each group. The overall analysis revealed a total of 22 phyla and 353 genera, with Firmicutes, Bacteroidetes, Proteobacteria, Actinobacteria, and Fusobacteria being the most abundant phyla. Bacteroides, Escherichia/Shigella, Lactobacillus, Collinsella, and Bifidobacterium were the most abundant genera. The reduction in Fusobacteria abundance before and after transport was significantly larger in the 16-h transportation group when compared with the 6-h transportation group. We also identified several genus-level and amplicon sequence variation-level taxa that displayed significant differences in their abundances across various transportation groups, observed at all sampling time points investigated. This research identifies microbiota changes due to varying transportation durations in surplus dairy calves, providing a broad understanding of the microbial shifts in surplus dairy calves after transportation across varying durations. Although these variations may not directly correlate with overall calf health or indicate dysbiosis, these results emphasize the importance of further investigating transportation practices to enhance calf health and well-being. Further studies are warranted to elucidate the relationship between microbiota and calf health.

Protective Effect of Baicalin against Clostridioides difficile Infection in Mice.

This study investigated the prophylactic and therapeutic efficacies of baicalin (BC), a plant-derived flavone glycoside, in reducing the severity of Clostridioides difficile infection (CDI) in a mouse model. In the prophylactic trial, C57BL/6 mice were provided with BC (0, 11, and 22 mg/L in drinking water) from 12 days before C. difficile challenge through the end of the experiment, whereas BC administration started day 1 post challenge in the therapeutic trial. Both challenge and control groups were infected with 106 CFU/mL of hypervirulent C. difficile BAA 1803 spores or sterile PBS, and the clinical and diarrheal scores were recorded for 10 days post challenge. On day 2 post challenge, fecal and tissue samples were collected from mice prophylactically administered with BC for microbiome and histopathologic analysis. Both prophylactic and therapeutic supplementation of BC significantly reduced the severity of colonic lesions and improved CDI clinical progression and outcome compared with control (p < 0.05). Microbiome analysis revealed a significant increase in Gammaproteobacteria and reduction in the abundance of protective microbiota (Firmicutes) in antibiotic-treated and C. difficile-infected mice compared with controls (p < 0.05). However, baicalin supplementation favorably altered the microbiome composition, as revealed by an increased abundance in beneficial bacteria, especially Lachnospiraceae and Akkermansia. Our results warrant follow-up investigations on the use of BC as an adjunct to antibiotic therapy to control gut dysbiosis and reduce C. difficile infection in humans.

In vitro antivirulence activity of baicalin against Clostridioides difficile.

Introduction. Clostridioides difficile is an enteric pathogen that causes a serious toxin-mediated colitis in humans. Bacterial exotoxins and sporulation are critical virulence components that contribute to pathogenesis, and disease transmission and relapse, respectively. Therefore, reducing toxin production and sporulation could significantly minimize C. difficile pathogenicity and disease outcome in affected individuals.Aim. This study investigated the efficacy of a natural flavone glycoside, baicalin, in reducing toxin synthesis, sporulation and spore germination in C. difficile in vitro.Methodology. Hypervirulent C. difficile isolates BAA 1870 or 1803 were cultured in brain heart infusion broth with or without the subinhibitory concentration (SIC) of baicalin, and incubated at 37 °C for 24 h under strictly anaerobic conditions. The supernatant was harvested after 24 h for determining C. difficile toxin production by ELISA. In addition, a similar experiment was performed wherein samples were harvested for assessing total viable counts, and heat-resistant spore counts at 72 h of incubation. Furthermore, C. difficile spore germination and spore outgrowth kinetics, with or without baicalin treatment, was measured in a plate reader by recording optical density at 600 nm. Finally, the effect of baicalin on C. difficile toxin, sporulation and virulence-associated genes was investigated using real-time quantitative PCR.Results. The SIC of baicalin significantly reduced toxin synthesis, sporulation and spore outgrowth when compared to control. In addition, C. difficile genes critical for pathogenesis were significantly down-regulated in the presence of baicalin.Conclusion. Our results suggest that baicalin could potentially be used to control C. difficile, and warrant future studies in vivo.

In vitro efficacy of sodium selenite in reducing toxin production, spore outgrowth and antibiotic resistance in hypervirulent Clostridium difficile.

PURPOSE: This study investigated the efficacy of the essential mineral, selenium (sodium selenite), in reducing the toxin production, spore outgrowth and antibiotic resistance of Clostridium difficile in vitro. METHODOLOGY: Two hypervirulent C. difficile isolates were cultured in brain heart infusion broth with and without a sub-minimum inhibitory concentration (sub-MIC) of sodium selenite, and the supernatant and bacterial pellet were harvested for total toxin quantitation and RT-qPCR analysis of toxin-encoding genes, respectively. Additionally, C. difficile isolates were cultured in brain heart infusion broth containing 0.5 or 1× the minimum inhibitory concentration (MIC) of either ciprofloxacin or vancomycin with or without sub-MICs of sodium selenite. Further, the effect of sodium selenite on C. difficile germination and spore outgrowth was also determined by exposing C. difficile spores to a sub-MIC of sodium selenite in a germination medium and measuring the germination and outgrowth by measuring the optical density at 600 nm. RESULTS: Sodium selenite significantly reduced C. difficile toxin synthesis, cytotoxicity and spore outgrowth. Further, the expression of the toxin production genes, tcdA and tcdB, was downregulated in the presence of sodium selenite, while sodium selenite significantly increased the sensitivity of C. difficile to ciprofloxacin , but not vancomycin, as revealed by decreased bacterial growth in samples containing ciprofloxacin+selenium compared to the antibiotic control. Although the sub-MIC of sodium selenite did not inhibit spore germination, it was capable of completely inhibiting spore outgrowth. CONCLUSION: Our results suggest that sodium selenite could potentially be used to control C. difficile and indicate that future in vivo studies are warranted.