Search Results (175)

Clostridioides difficile is a leading pathogen involved in healthcare-associated diarrhea. With its increasing incidence, mortality, and antibiotic resistance, there is an urgent need for novel therapeutic strategies to address the infection and prevent its recurrence. Gegen Qinlian Decoction (GQD) is a traditional Chinese medicine for the treatment of diarrhea, but its main active ingredient is not known. Therefore, in this study, we evaluated the biological activity of berberine (BER) and baicalein (BAI), key components of GQD, against C. difficile. Time–kill curves and scanning electron microscopy were employed to assess their effects on C. difficile growth, while Enzyme-Linked Immunosorbnent Assay (ELISA) and cytotoxicity assays were used to examine their impact on toxin production. We also employed Quantitative Reverse Transcription PCR (qRT-PCR) to examine how BER and BAI influenced the expression of toxin-associated genes. At sub-inhibitory concentrations, these compounds exerted antibacterial activity against C. difficile by disrupting the integrity of the cell membrane and cell wall. Furthermore, BER and BAI also suppressed toxin production, demonstrating effects comparable to those of vancomycin. This suppression likely resulted from their bactericidal activity and the inhibition of toxin gene expression. This study not only highlights the potential application of GQD in treating C. difficile infections but also offers promising options for developing drugs targeting the growth and virulence of this pathogen. C. difficile infection (CDI) is a leading cause of severe diarrhea, and its treatment remains challenging due to limited drug options and its high recurrence rate. BAI and BER, the main active components of the traditional Chinese medicinal formula GQD, inhibited the growth of C. difficile by disrupting its cellular structure and significantly reduced the production of toxins associated with disease severity. Furthermore, the effects of BAI and BER on C. difficile were comparable to those of conventional antibiotics, suggesting that these compounds could be potential alternative therapies for CDI. This study not only highlights the therapeutic potential of GQD in treating CDI but also provides a replicable research strategy for the development of novel anti-CDI agents. Full article

Background/Objectives: Clostridioides difficile is a “One Health” pathogen and a cause of antibiotics-associated diarrhea and pseudomembranous colitis. Mobile genetic elements (MGEs) have been documented in the genomes of clinical C. difficile strains; however, the presence of MGEs in environmental strains remains poorly characterized. Thus, the present study was conducted with the objective of identifying the prevalence of MGEs, including mobilizable transposons (MTns), conjugative transposons (CTns), plasmids, and insertion sequences, in whole genome sequences (WGSs) of environmental C. difficile isolates. Methods: The analysis of MGEs was conducted using 166 WGSs obtained from C. difficile strains isolated from various environmental sources contaminated with feces. The MGEs were identified using bioinformatic tools. Results: A total of 48.2% (80/166) of the studied genomes were identified to harbor nine transposons, including Tn916, Tn6194-like, Tn5397, Tn6215, Tn4001, Tn6073, Tn6110, Tn6107, or Tn5801-like. The majority of MTns and CTns could be found within C. difficile sequence types ST11, ST3, and ST35. The results demonstrated close genetic relatedness among the studied genomes, the array of antimicrobial resistance (AMR) genes, such as tetM, ermB, and aac(6′)-aph(2″), and the presence of CTns. Furthermore, the analysis revealed that 24.7% (41/166) of the genome sequences of isolates were associated with various predominant plasmid groups, including pCD6, pCD-ECE4-6, pCD-WTSI1-4, pCDBI1, and pCd1_3, which belonged to 16 different sequence types. Furthermore, several plasmids were identified as harboring the prophage phiCDHM19. Conclusions: The results of the current study suggest that the identified plasmids are abundant and may encode functions that are relevant to C. difficile physiology. The genomes of C. difficile strains examined contain closely related CTns, suggesting that horizontal transfer of AMR is important in this species or other bacterial species. Further research is required to ascertain the effect of these genetic elements and their transferability on the biology of C. difficile. Full article

Defensins, one of the members of the antimicrobial peptide family, play a vital role in resisting microbial invasion and immune regulation. Porcine β-defensin 2 possesses excellent stability, making it an ideal antibiotic alternative for feed additives. In this study, a total of 15 piglets were used to investigate the effects of supplementing diets with 2.5 mg/kg (LP group) and 5 mg/kg (HP group) of pBD2 to weaned piglets. The results revealed that pBD2 significantly increased the total weight gain and average daily weight gain (p < 0.05), the contents of T-AOC, SOD, IgM, and IL-10 in serum (p < 0.05), the villus-to-crypt ratios, and the expression of tight-junction proteins ZO-1 and claudin-1 (p < 0.05) in the small intestine. Furthermore, pBD2 increased the abundance of beneficial bacteria related to nutrient and energy metabolism while decreasing the abundance of harmful bacteria associated with intestinal inflammation and diarrhea. Alterations in the gut microbiota were closely associated with the levels of T-AOC, SOD, IgM, and IL-10 in serum. pBD2 primarily enhanced the health of weaned piglets by influencing antioxidant capacity, intestinal barrier function, and the intestinal microbiota. Our research provides a novel perspective for addressing the issue of antibiotic residues in feed. Full article

Clostridioides difficile is a Gram-positive bacterium recognized for its ability to produce toxins and form spores. It is mainly accountable for the majority of instances of antibiotic-related diarrhea. Background. Bacterial persister represent a minor fraction of the population that shows temporary tolerance to bactericidal agents, and they pose considerable medical issues because of their link to the rise of antibiotic resistance and challenging chronic or recurrent infections. Our previous research has shown a persister-like phenotype associated with treatments that include pefloxacin. Nonetheless, the mechanism is still mostly unclear, mainly because of the difficulty in isolating this small group of cells. Objectives. To enhance the understanding of C. difficile persister cells, we made an enrichment and characterization of these cells from bacterial cultures during the exponential phase under pefloxacin treatment and lysis treatment. Results. We demonstrate the appearance of cells with lower metabolism and DNA damage. Furthermore, we noted the participation of toxin–antitoxin systems and Clp proteases in the generation of persister cells. Conclusions. This work demonstrates the formation of C. difficile persister cells triggered by a lethal concentration of pefloxacin. Full article

Piglet diarrhea caused by weaning stress will increase the mortality rate and seriously affect swine industry production efficiency. Probiotic supplementation has been reported to effectively alleviate weaning diarrhea by inhibiting the colonization of pathogenic microorganisms; however, the underlying mechanisms remain unclear. In this study, we isolated a strain of Bacillus velezensis and conducted a series of in vivo and in vitro experiments to explore its effects on weaned piglets. The piglets were fed for a 28-day period, and the results showed that dietary supplementation of B. velezensis 411 significantly alleviated weaning diarrhea (p = 0.019) and improved the average daily gain (ADG) of piglets throughout the experimental period (p = 0.004). The intestinal antioxidant capacity of piglets was also significantly enhanced. Whole-genome sequencing revealed that B. velezensis 411 contains a protein-encoding circular chromosome, which is involved in biological processes such as sporulation and antibiotic secretion. Supplementation with B. velezensis 411 significantly increased the abundance of Akkermansia in intestine samples and significantly decreased the abundance of pathogenic bacteria, including Escherichia coli and Staphylococcus aureus, in piglets (p < 0.05). The transcriptomic results suggest that B. velezensis 411 supplementation may alter the composition of intestinal microorganisms through regulating the expression of MPEG1. Collectively, dietary B. velezensis can relieve diarrhea in piglets and improve their production performance by influencing the antioxidant capacity of the intestines and the balance of the intestinal flora. This study provides valuable insights into the potential application of Bacillus velezensis in mitigating weaning-associated issues in piglets. Full article

Background: Antibiotic-associated diarrhea (AAD) is a common adverse effect of pediatric antibiotic therapy, often linked to gut microbiota disruption. Probiotics may help prevent AAD when appropriately selected and dosed. Methods: We conducted a cross-sectional survey to assess the attitudes and prescribing habits of Italian Primary Care Pediatricians (PCPs) regarding the use of probiotics during antibiotic treatment. A digital questionnaire comprising 23 mandatory multiple-choice items was distributed to 980 PCPs across Italy between July and October 2024. The survey explored probiotic prescribing frequency, indications, strains used, dosage, duration, and sources of information. Descriptive statistics and subgroup analyses by years of clinical experience were performed. Results: A total of 279 PCPs (response rate: 28%) completed the survey; 66.7% were female, and 77.1% had over 20 years of clinical experience. Probiotics were prescribed primarily to restore microbiota balance (81.1%) and prevent AAD (47.3%). The most common barriers included additional cost (35.1%) and perceived lack of evidence (26.5%). Lactobacillus rhamnosus GG (91.8%) and Saccharomyces boulardii (41.9%) were the most frequently recommended strains. Daily doses of 5–10 billion CFU were preferred by 44.4% of respondents, with typical durations of 1–2 weeks (40.1%) or one week (31.2%). Conclusions: Probiotics are widely used by Italian PCPs during antibiotic therapy, especially for microbiota support and AAD prevention. However, variability in practice underscores the need for clearer, evidence-based guidelines regarding probiotic strain selection, dosing, and treatment duration. Full article

Objective: This review aims to comprehensively evaluate the current evidence on the role of prebiotics, probiotics, synbiotics, and postbiotics—collectively referred to as “biotics”—in modulating the human gut microbiota and enhancing intestinal epithelial integrity. Findings: Biotics exert their beneficial effects through several mechanisms, including by promoting the growth of beneficial microbes, producing short-chain fatty acids (SCFAs), strengthening the gut barrier, and regulating immune responses. Prebiotics selectively stimulate beneficial bacteria, probiotics introduce live microorganisms with therapeutic functions, synbiotics combine the strengths of both, and postbiotics offer non-viable microbial components and metabolites that mimic probiotic benefits with enhanced safety profiles. Each type of biotic demonstrates unique and complementary effects across a range of conditions, such as inflammatory bowel disease, irritable bowel syndrome, obesity, constipation, and antibiotic-associated diarrhea. Implications: As disruptions in the gut microbiota and intestinal barrier are increasingly linked to chronic and immune-mediated diseases, leveraging biotics offers promising avenues for personalized nutrition, preventive healthcare, and adjunct therapies. The integration of biotics into clinical and dietary strategies may significantly contribute to improving gastrointestinal and systemic health. Full article

Background: Enterohemorrhagic Escherichia coli (EHEC) is a considerable public health concern associated with several foodborne outbreaks of bloody diarrhea (BD) and the potentially lethal hemolytic uremic syndrome (HUS), the pathophysiology of which is attributable to the Shiga toxin (Stx) produced by this bacterium. In most patients, supportive treatment will be sufficient; however, in some cases, antibiotic treatment may be necessary. Most antibiotics are not recommended for EHEC infection treatment, particularly those that kill the bacteria, since this triggers the release of Stx in the body, inducing or worsening HUS. Azithromycin, which prevents the release of Stx and is a weaker inducer of the SOS system, has been successfully used to reduce EHEC shedding. It is necessary to identify compounds that eliminate EHEC without inducing Stx release. The use of natural compounds such as curcumin (CUR), a polyphenol derived from turmeric, has been highlighted as an alternative bactericidal treatment approach. Objective: The objective of this study was to establish the effect of CUR and its interactions with selected antibiotics on resistant EHEC O157/H7/EDL933. Methods: Bacterial cultures were exposed to CUR at three different concentrations (110, 220, and 330 µg/mL) and 1.2% DMSO, and the antimicrobial activity of CUR was assessed by measuring the optical density at 600 nm (OD600). The synergy of CUR and the antibiotics was determined with the FIC method. RT-PCR was performed to determine the expression levels of the bla_CTX-M-15, catA1, acrAB-tolC stx2A, and stx2B genes. Results: Our data indicate that CUR did not affect the growth of EHEC, but when combined with the antibiotics, it acted as a bacterial resistance breaker. Synergistic combinations of CUR and cefotaxime or chloramphenicol significantly reduced colony counts. Conclusions: Our findings support the potential of CUR as a sensitizer or in combination therapy against EHEC. Full article

Clostridioides difficile infection (CDI) appears mainly as nosocomial antibiotic-associated diarrhea, and community-acquired infection is increasingly being recognized. The threshold of asymptomatic colonization and the clinical manifestation of CDI need further elucidation. Community-acquired CDI (CA-CDI) should be considered when the disease commences within 48 h of admission to hospital or more than 12 weeks after discharge. Although CDI is not established as a food-borne or zoonotic disease, some data support that direction. The spores’ ability to survive standard cooking procedures and on abiotic surfaces, the formation of biofilms, and their survival within biofilms of other bacteria render even a low number of spores capable of food contamination and spread. Adequate enumeration methods for detecting a low number of spores in food have not been developed. Primary care physicians should take CA-CDI into consideration in the differential diagnosis of diarrhea, as there is a thin line between colonization and infection. In patients diagnosed with inflammatory bowel disease and other comorbidities, C. difficile can be the cause of recurrent disease and should be included in the estimation of diarrhea and worsening colitis symptoms. In the community setting, it is difficult to distinguish asymptomatic carriage from true infection. For asymptomatic carriage, antibiotic therapy is not suggested but contact isolation and hand-washing practices are required. Primary healthcare providers should be vigilant and implement infection control policies for the prevention of C. difficile spread. Full article

Campylobacter spp. is one of the most prevalent causes of zoonotic foodborne infections associated with diarrhea in humans. The growing threat of antibiotic resistance calls for innovative approaches. The antimicrobial lipopeptide brevibacillin produced by Brevibacillus laterosporus and its synthetic analog brevibacillin Thr1 showed promising activity against Salmonella and E. coli. The latter is a 1602.13 Da positively charged (+3) synthetic peptide of 13 residues that showed reduced cytotoxicity (IC₅₀ of 32.2 µg/mL against Caco-2 cells) and hemolytic activity (1.2% hemolysis at 128 µg/mL) compared to the native peptide. It contains an N-terminal L-isoleucic fatty acid chain and four non-proteinogenic amino acids and ends with valinol at its C-terminus. One key structural modification is the substitution of α,β-dehydrobutyric acid with threonine. We investigated the antimicrobial potential of the synthetic brevibacillin Thr1 analog against a collection of 44 clinical Campylobacter spp. that were obtained from two reference laboratories. Susceptibility testing revealed marked resistance to ciprofloxacin, tetracycline, and ampicillin among the strains, with more than half expressing a multidrug-resistant phenotype. The genomes of the 44 strains were sequenced to study the genes responsible for their antimicrobial resistance. Tetracycline resistance was associated with tet(O), ciprofloxacin resistance with mutations in gyrA and regulatory sequences modulating the expression of an efflux system, and aminoglycoside resistance with genes of the aph family. The brevibacillin Thr1 analog was produced by chemical synthesis, and evaluation of its activity against a subset of clinical strains by microdilution revealed minimum inhibitory concentration and minimum bactericidal concentration ranging from 8 µg/mL to 64 µg/mL. The peptide was active against multidrug-resistant isolates with a bactericidal effect. Of note, despite numerous attempts, it proved impossible to select Campylobacter spp. for resistance to the brevibacillin Thr1 analog. These results underline the potential of lipopeptides, notably brevibacillin, as antimicrobial alternatives against antibiotic-resistant Campylobacter bacterial infections. Full article

Probiotics have gained increasing recognition for their potential to mitigate antibiotic-associated diarrhea (AAD). However, the precise mechanisms underlying their effects remain unclear. This study developed a mouse model of AAD using ceftriaxone to investigate the alleviating effects and mechanisms of Bifidobacterium animalis A6 (A6). The findings indicated that A6 supplementation effectively attenuated ceftriaxone-associated diarrhea in mice. The morphological damage to the villi and crypts was partially restored and more neatly reorganized following the A6 intervention. Additionally, intestinal morphology observations revealed a significant increase in the thickness of the mucus layer in the A6-treated group. Further examination of key regulatory genes associated with mucus secretion demonstrated that the A6 intervention effectively upregulated the expression of mucin1, thereby reinforcing the mucus layer. Concurrently, the A6 intervention upregulated the expression of the AQP4 and SLC26A3 genes in the intestine, which is responsible for restoring water absorption capacity in AAD mice. Additionally, the A6 treatment reduced ceftriaxone-induced harm to the intestinal microbiota of the mice, boosting beneficial bacteria like Bacteroidales, Akkermansia, Bifidobacterium, and Lactobacillus. Overall, this study offers valuable insights into the potential therapeutic role of A6 in restoring intestinal homeostasis and alleviating symptoms associated with AAD. Full article

Background/Objective. Toxin-producing strains of Clostridioides difficile (C. diff) are the most commonly identified cause of healthcare-associated infection in the elderly. Risk factors include advanced age, hospitalization, prior or concomitant systemic antibacterial therapy, chemotherapy, and gastrointestinal surgery. Patients with unspecified and new-onset diarrhea with ≥3 unformed stools in 24 h are the target population for C. diff infection (CDI) testing. To present data on the risks of laboratory misdiagnosis in managing CDI. Materials. In two general hospitals, we examined 116 clinical stool specimens from hospitalized patients with acute diarrhea suspected of nosocomial or antibiotic-associated diarrhea (AAD) due to C. diff. Enzyme immunoassay (EIA) tests for the detection of C. diff toxins A (cdtA) and B (cdtB) in stool, automated CLIA assay for the detection of C. diff GDH antigen and qualitative determination of cdtA and B in human feces and anaerobic stool culture were applied for CDI laboratory diagnosis. MALDI-TOF (Bruker) was used to identify the presumptive anaerobic bacterial colonies. The following methods were used as confirmatory diagnostics: the LAMP method for the detection of Salmonella spp. and simultaneous detection of C. jejuni and C. coli, an E. coli Typing RT-PCR detection kit (ETEC, EHEC, STEC, EPEC, and EIEC), API 20E and aerobic stool culture methods. Results. A total of 40 toxigenic strains of C. diff were isolated from all 116 tested diarrheal stool samples, of which 38/40 produced toxin B and 2/40 strains were positive for both cdtA and cdtB. Of the stool samples positive for cdtA (6/50) and/or cdtB (44/50) by EIA, 33 were negative for C. diff culture but positive for the following diarrheal agents: Salmonella enterica subsp. arizonae (1/33, LAMP, culture, API 20E); C. jejuni (2/33, LAMP, culture, MALDI TOF); ETEC O142 (1/33), STEC O145 and O138 (2/33, E. coli RT-PCR detection kit, culture); C. perfringens (2/33, anaerobic culture, MALDI TOF); hypermycotic enterotoxigenic K. pneumonia (2/33) and enterotoxigenic P. mirabilis (2/33, culture; PCR encoding LT-toxin). Two of the sixty-six cdtB-positive samples (2/66) showed a similar misdiagnosis when analyzed using the CLIA method. However, the PCR analysis showed that they were cdtB-negative. In contrast, the LAMP method identified a positive result for C. jejuni in one sample, and another was STEC positive (stx1+/stx2+) by RT-PCR. We found an additional discrepancy in the CDI test results: EPEC O86 (RT-PCR eae+) was isolated from a fecal sample positive for GHA enzyme (CLIA) and negative for cdtA and cdtB (CLIA and PCR). However, the culture of C. diff was negative. These findings support the hypothesis that certain human bacterial pathogens that produce enterotoxins other than C. diff, as well as intestinal commensal microorganisms, including Klebsiella sp. and Proteus sp., contribute to false-positive EIA card tests for C. diff toxins A and B, which are the most widely used laboratory tests for CDI. Conclusions. CDI presents a significant challenge to clinical practice in terms of laboratory diagnostic management. It is recommended that toxin-only EIA tests should not be used as the sole diagnostic tool for CDI but should be limited to detecting toxins A and B. Accurate diagnosis of CDI requires a combination of laboratory diagnostic methods on which proper infection management depends. Full article

Clostridioides difficile (C. difficile) is a major pathogen responsible for antibiotic-associated diarrhea, frequently observed in hospital settings. Due to the widespread use of antibiotics, the incidence and severity of C. difficile infection (CDI) are rising across the world. CDI is primarily driven by two homologous protein exotoxins, toxin A (TcdA) and toxin B (TcdB). Other putative virulence factors include binary toxin CDT, surface layer proteins, phosphorylated polysaccharides, and spore coat proteins. These C. difficile virulence factors are potential targets for vaccine development. Although several C. difficile vaccines have entered clinical trials, there is currently no approved vaccine on the market. This review outlines the intoxication mechanism during CDI, emphasizing the potential antigens that can be used for vaccine development. We aim to provide a comprehensive overview of the current status of research and development of C. difficile vaccines. Full article

The weaning period is a critical phase for nursery pigs that is characterized by rapid growth and alterations in the intestinal microbiome associated with nutrient utilization. The present study aimed to investigate the efficacy of halquinol, when used as an antibiotic (ABO), on the growth performance, diarrhea incidence, coefficient of apparent total tract digestibility (CATTD), fecal volatile fatty acids (VFAs), and microbiota in pigs. A total of 210 healthy weaned pigs with an average initial weight of 6.9 kg and aged 28 ± 2 days were assigned to five treatments (six pens/treatment) in a complete randomized design, including a control group (T1, CON; feed with no ABO), a colistin group (T2, CLT; feed containing 120 ppm colistin), and three halquinol groups (T3 to T5, HAL; feed containing 180, 240, and 360 ppm halquinol, respectively). The experiment period lasted for 10 days. Field recordings, observation, and feces collection were performed on D1, D5, and D10. CATTD and VFA assessments were conducted on D10. The composition of the fecal microbiota was analyzed via 16S rRNA gene sequencing using the Illumina Miseq platform. The results demonstrated that the in-feed ABO groups exhibited a significantly lower ADFI (p < 0.01). Pigs fed the T3 and T4 diets had the lowest FCR (p < 0.01) on D5 and D10 and, thus, had reduced ADFI (p < 0.01). A quadratic contrast was found in ADFI and FCR on D5 and D10, indicating a negative correlation with HAL concentration (p < 0.01). Pigs fed CLT and HAL had significantly reduced levels of coliform (p < 0.01) and E. coli (p < 0.01). Moreover, pigs receiving ABO also had a lower fecal score compared to those on the CON diet (p < 0.01). Dietary in-feed ABO had no effect on all the parameters of the CATTD on D10 (p > 0.05), except for fat digestibility in pigs that received T4 (p < 0.01). Pigs fed the T4 and T5 diets had higher propionate concentrations and lower A/P ratios than pigs fed T1, T2, and T3 (p < 0.01). The microbial diversity shifted quickly through the early weaning period. The relative abundance of beneficial Enterococcus microbes increased in pigs fed in-feed ABO, whereas the relative prevalence of pathogenic bacteria, such as Escherichia and Klebsiella, decreased. Escherichia and Bacteroides were negatively correlated with carbohydrate digestibility and butyric and valeric acid production (p < 0.05). Overall, the appropriate HAL dosage was 240 ppm (T4), and this antimicrobial can potentially be characterized as an in-feed colistin replacer that improves feed efficiency and fat digestion, enhancing VFA production, alleviating post-weaning diarrhea, and protecting ABO-resistant piglets. Full article

Background: Dysbiosis and antibiotic-associated diarrhea (AAD) are significant concerns in clinical settings. Probiotics, such as Bacillus clausii (O/C, N/R, SIN, T), a spore-forming bacterium resistant to gastrointestinal conditions and most commonly used antibiotics, emerge as a promising approach. We aim to assess the role of B. clausii in preventing AAD in children and adults during antibiotic therapy. Materials and methods: A systematic literature search was conducted across multiple databases, including MEDLINE, EMBASE, Cochrane Library, LILACS, and SciELO, up to May 2024. Studies were included if they involved B. clausii (O/C, N/R, SIN, T) administration during antibiotic treatment and reported AAD-related outcomes. Results: A total of four studies were included in the review. The studies comprised two randomized controlled trials (RCTs), one meta-analysis of RCTs, and one expert consensus. The primary outcome was the effectiveness of B. clausii (O/C, N/R, SIN, T) in reducing the incidence of diarrhea. Results showed a significant reduction in the risk of AAD and gastrointestinal symptoms in patients receiving it at a dosage of 4 × 10⁹ CFU/day for children and 6 × 10⁹ CFU/day for adolescents and adults for up to 14 days. Conclusions: B. clausii (O/C, N/R, SIN, T) appears to be an effective probiotic for preventing AAD in adults and children. It significantly improves gastrointestinal symptoms associated with antibiotic treatment, including diarrhea, nausea, and epigastric pain. Future studies are recommended to further elucidate its effectiveness in diverse populations, especially in low- and middle-income countries. Full article