Quorum-quenching enzyme Est816 assisted antibiotics against periodontitis induced by Aggregatibacter actinomycetemcomitans in rats.

Introduction: Quorum-quenching enzyme Est816 hydrolyzes the lactone rings of N-acyl homoserine lactones, effectively blocking the biofilm formation and development of Gram-negative bacteria. However, its applications in the oral field is limited. This study aimed to evaluate the efficacy of enzyme Est816 in combination with antibiotics against periodontitis induced by Aggregatibacter actinomycetemcomitans in vitro and in vivo. Methods: The antimicrobial efficacy of enzyme Est816 in combination with minocycline, metronidazole, and amoxicillin was determined using the minimum inhibitory concentration test. The anti-biofilm effect of enzyme Est816 was assessed using scanning electron microscopy, live/dead bacterial staining, crystal violet staining, and real-time quantitative PCR. Biocompatibility of enzyme Est816 was assessed in human gingival fibroblasts (HGF) by staining. A rat model of periodontitis was established to evaluate the effect of enzyme Est816 combined with minocycline using micro-computed tomography and histological staining. Results: Compared to minocycline, metronidazole, and amoxicillin treatment alone, simultaneous treatment with enzyme Est816 increased the sensitivity of biofilm bacteria to antibiotics. Enzyme Est816 with minocycline exhibited the highest rate of biofilm clearance and high biocompatibility. Moreover, the combination of enzyme Est816 with antibiotics improved the antibiofilm effects of the antibiotics synergistically, reducing the expression of the virulence factor leukotoxin gene (ltxA) and fimbria-associated gene (rcpA). Likewise, the combination of enzyme Est816 with minocycline exhibited a remarkable inhibitory effect on bone resorption and inflammation damage in a rat model of periodontitis. Discussion: The combination of enzyme Est816 with antibiotics represents a prospective anti-biofilm strategy with the potential to treat periodontitis.

15N Hyperpolarization of Metronidazole Antibiotic in Aqueous Media Using Phase-Separated Signal Amplification by Reversible Exchange with Parahydrogen.

Metronidazole is a prospective hyperpolarized MRI contrast agent with potential hypoxia sensing utility for applications in cancer, stroke, neurodegenerative diseases, etc. We demonstrate a pilot procedure for production of ∼30 mM hyperpolarized [15N3]metronidazole in aqueous media by using a phase-separated SABRE-SHEATH hyperpolarization method, with nitrogen-15 polarization exceeding 2.2% on all three 15N sites achieved in less than 2 min. The 15N polarization T1 of ∼12 min is reported for the 15NO2 group at the clinically relevant field of 1.4 T in the aqueous phase, demonstrating a remarkably long lifetime of the hyperpolarized state. The produced aqueous solution of [15N3]metronidazole that contained only ∼100 µM of residual Ir was deemed biocompatible via validation through the MTT colorimetric test for assessing cell metabolic activity using human embryotic kidney HEK293T cells. This low-cost and ultrafast hyperpolarization procedure represents a major advance for the production of a biocompatible HP [15N3]metronidazole (and potentially other hyperpolarized drugs) formulation for MRI sensing applications.

Toxin genotypes, antibiotic resistance and their correlations in Clostridioides difficile isolated from hospitals in Xi'an, China.

BACKGROUND: Clostridioides difficile is the main pathogen of antimicrobial-associated diarrhoea and health care facility-associated infectious diarrhoea. This study aimed to investigate the prevalence, toxin genotypes, and antibiotic resistance of C. difficile among hospitalized patients in Xi'an, China. RESULTS: We isolated and cultured 156 strains of C. difficile, representing 12.67% of the 1231 inpatient stool samples collected. Among the isolates, tcdA + B + strains were predominant, accounting for 78.2% (122/156), followed by 27 tcdA-B + strains (27/156, 17.3%) and 6 binary toxin gene-positive strains. The positive rates of three regulatory genes, tcdC, tcdR, and tcdE, were 89.1% (139/156), 96.8% (151/156), and 100%, respectively. All isolates were sensitive to metronidazole, and the resistance rates to clindamycin and cephalosporins were also high. Six strains were found to be resistant to vancomycin. CONCLUSION: Currently, the prevalence rate of C. difficile infection (CDI) in Xi'an is 12.67% (156/1231), with the major toxin genotype of the isolates being tcdA + tcdB + cdtA-/B-. Metronidazole and vancomycin were still effective drugs for the treatment of CDI, but we should pay attention to antibiotic management and epidemiological surveillance of CDI.

Silver Complexes of Miconazole and Metronidazole: Potential Candidates for Melanoma Treatment.

Melanoma, arguably the deadliest form of skin cancer, is responsible for the majority of skin-cancer-related fatalities. Innovative strategies concentrate on new therapies that avoid the undesirable effects of pharmacological or medical treatment. This article discusses the chemical structures of [(MTZ)2AgNO3], [(MTZ)2Ag]2SO4, [Ag(MCZ)2NO3], [Ag(MCZ)2BF4], [Ag(MCZ)2SbF6] and [Ag(MCZ)2ClO4] (MTZ-metronidazole; MCZ-miconazole) silver(I) compounds and the possible relationship between the molecules and their cytostatic activity against melanoma cells. Molecular Hirshfeld surface analysis and computational methods were used to examine the possible association between the structure and anticancer activity of the silver(I) complexes and compare the cytotoxicity of the silver(I) complexes of metronidazole and miconazole with that of silver(I) nitrate, cisplatin, metronidazole and miconazole complexes against A375 and BJ cells. Additionally, these preliminary biological studies found the greatest IC50 values against the A375 line were demonstrated by [Ag(MCZ)2NO3] and [(MTZ)2AgNO3]. The compound [(MTZ)2AgNO3] was three-fold more toxic to the A375 cells than the reference (cisplatin) and 15 times more cytotoxic against the A375 cells than the normal BJ cells. Complexes of metronidazole with Ag(I) are considered biocompatible at a concentration below 50 µmol/L.

Evidence of Helicobacter pylori heterogeneity in human stomachs by susceptibility testing and characterization of mutations in drug-resistant isolates.

Heterogeneity of Helicobacter pylori communities contributes to its pathogenicity and diverse clinical outcomes. We conducted drug-susceptibility tests using four antibiotics, clarithromycin (CLR), amoxicillin (AMX), metronidazole and sitafloxacin, to examine H. pylori population diversity. We also analyzed genes associated with resistance to CLR and AMX. We examined multiple isolates from 42 Japanese patients, including 28 patients in whom primary eradication with CLR and AMX had failed, and 14 treatment-naïve patients. We identified some patients with coexistence of drug resistant- and sensitive-isolates (drug-heteroR/S-patients). More than 60% of patients were drug-heteroR/S to all four drugs, indicating extensive heterogeneity. For the four drugs except AMX, the rates of drug-heteroR/S-patients were higher in treatment-naïve patients than in primary eradication-failure patients. In primary eradication-failure patients, isolates multi-resistant to all four drugs existed among other isolates. In primary eradication-failure drug-heteroR/S-patients, CLR- and AMX-resistant isolates were preferentially distributed to the corpus and antrum with different minimum inhibitory concentrations, respectively. We found two mutations in PBP1A, G591K and A480V, and analyzed these in recombinants to directly demonstrate their association with AMX resistance. Assessment of multiple isolates from different stomach regions will improve accurate assessment of H. pylori colonization status in the stomach.

Promising effects of 1,8 Cineole to control Giardia lamblia infection: Targeting the inflammation, oxidative stress, and infectivity.

Reportedly, synthetic drugs such as metronidazole, furazolidone, tinidazole, and quinacrine are used for the treatment of giardiasis but are associated with adverse effects. In this study, we aimed to investigate the in vitro and in vivo effects of eucalyptol (ECT, 1,8 cineole) alone and in combination with metronidazole (MNZ) on Giardia lamblia. The effects of ECT on cell viability, plasma membrane permeability, and gene expression levels of adenylate cyclase (AK) and extracellular signal kinases 1 and 2 (ERK1 and ERK2) in trophozoites of G. lamblia were assessed. In vivo, the effects of ECT alone and in combination with MNZ were assessed on mice infected with G. lamblia. In addition, the gene expression of inflammatory genes (e.g., TNF-α, IL-1ß, and IL-10) and antioxidant genes (catalase (CAT), superoxide dismutase 1 (SOD1), glutathione peroxidase 2 (GPX2)) was determined by real-time PCR. The IC50 values of ECT, MNZ, and ECT+MNZ on trophozoites were 30.2 µg/mL, 21.6 µg/mL, and 8.5 µg/mL, respectively. The estimated Fractional inhibitory concentration index (FICI) values for ECT and MNZ were 0.28 and 0.39, respectively. The application of ECT on G. lamblia trophozoites resulted in a dose-dependent increase in plasma membrane permeability, particularly at concentrations of ½ IC50 and IC50 (P < 0.05). The treatment of infected mice with various doses of ECT, mainly in combination with MNZ for 7 days, resulted in a significant decrease (P < 0.001) in the average number and viability of cysts. ECT, especially when combined with MNZ, caused a significant (P < 0.001) reduction in the expression of TNF-α and IL-6 genes, and an increase (P < 0.05) in the expression of IL-10 genes. ECT alone and mainly in combination with MNZ leads to a significant (P < 0.001) increase in the gene expression of CAT, SOD, and GPX genes. These findings demonstrate that the use of ECT in these doses, even for 14 days, does not have any toxic effects on the function of vital liver and kidney tissues. The study findings confirmed the promising effects of ECT against G. lamblia infection both in vitro and in vivo. Considering the possible mechanisms, ECT increases plasma membrane permeability and reduces the expression levels of infectivity-related genes. In addition, ECT suppresses inflammation and oxidative stress, controlling giardiasis in mice. More studies are needed to clarify these findings.

Refractoriness to anti-Helicobacter pylori treatment attributed to phenotypic resistance patterns in patients with gastroduodenopathy in Guayaquil-Ecuador.

BACKGROUND: Treatment of Helicobacter pylori gastric infection is complex and associated with increased rates of therapeutic failure. This research aimed to characterize the H. pylori infection status, strain resistance to antimicrobial agents, and the predominant lesion pattern in the gastroduodenal mucosa of patients with clinical suspicion of refractoriness to first- and second-line treatment who were diagnosed and treated in a health center in Guayaquil, Ecuador. METHODS: A total of 374 patients with upper gastrointestinal symptoms and H. pylori infection were preselected and prescribed one of three triple therapy regimens for primary infection, as judged by the treating physician. Subsequently, 121 patients who returned to the follow-up visit with persistent symptoms after treatment were studied. RESULTS: All patients had H. pylori infection. Histopathological examination diagnosed chronic active gastritis in 91.7% of cases; premalignant lesions were observed in 15.8%. The three triple therapy schemes applied showed suboptimal efficacy (between 47.6% and 77.2%), with the best performance corresponding to the scheme consisting of a proton pump inhibitor + amoxicillin + levofloxacin. Bacterial strains showed very high phenotypic resistance to all five antimicrobials tested: clarithromycin, 82.9%; metronidazole, 69.7%; amoxicillin and levofloxacin, almost 50%; tetracycline, 38.2%. Concurrent resistance to clarithromycin-amoxicillin was 43.4%, to tetracycline-metronidazole 30.3%, to amoxicillin-levofloxacin 27.6%, and to clarithromycin-metronidazole 59.2%. CONCLUSIONS: In vitro testing revealed resistance to all five antibiotics, indicating that H. pylori exhibited resistance phenotypes to these antibiotics. Consequently, the effectiveness of triple treatments may be compromised, and further studies are needed to assess refractoriness in quadruple and concomitant therapies.

Conserved signatures of the canine faecal microbiome are associated with metronidazole treatment and recovery.

Antibiotic resistance is recognised as one of the biggest global threats to human and animal health. Understanding the influence of antibiotics on the canine microbiome is important to know the potential mid-to-long term effects on dysbiosis and mitigate side-effects such as antibiotic-associated diarrhoea. In this study, metronidazole was prescribed to 22 dogs for suspected giardiasis after exhibiting gastrointestinal symptoms such as diarrhoea and/or vomiting. Faecal samples were collected before, during seven days of treatment, and six months post-cessation. Faecal microbiota was assessed with 16S rRNA sequencing. Shannon diversity was reduced for up to three days after the treatment ended, and an altered community persisted for four to six weeks. All dogs recovered to a similar microbiome composition as pre-treatment. Immediately after receiving metronidazole, an increase in the relative abundance of the genera Lactobacillus, Bifidobacterium, and Enterococcus was observed. This may be due to antibiotic resistance commonly exhibited by these organisms. One-to-two weeks post-cessation, several other genera that were sensitive to the antibiotic recovered in abundances, with taxa belonging to the Erysipelotrichaceae family particularly driving composition change. Many of the bacteria initially reduced were associated with carbohydrate fermentation. This suggests scope exists to explore interventions to augment gastrointestinal health and support the re-establishment of the microbiome.

In vitro anti-Trichomonas gallinae effects of Ziziphus vulgaris L. and Camellia sinensis (L.) Kuntze extracts.

BACKGROUND: Trichomonas gallinae is a parasite that causes canker and severe loss and death, especially in young pigeons. Metronidazole (MTZ) is the recommended drug for treating avian trichomoniasis. Due to drug resistance, non-chemical alternatives, such as medicinal plant extracts, are also considered possible therapies for this disease. OBJECTIVES: This study compares the antitrichomonal effects of MTZ with extracts of Camellia sinensis and Ziziphus vulgaris on T. gallinae in vitro. METHODS: Samples of T. gallinae were taken from infected pigeons. Multi-well plates with different concentrations (5, 10, 25, 50 and 100 µg/mL) of plant extracts were used for the in vitro study. RESULTS: The minimum inhibitory concentration (MIC) of C. sinensis extract was 25 µg/mL over 24 h, compared to 50 µg/mL for MTZ. The MIC value of the Z. vulgaris extracts was 50 µg/mL. CONCLUSIONS: The results suggest that the extracts of Z. vulgaris and C. sinensis, as potential natural agents, could have anti-avian trichomoniasis properties. This study also shows that MTZ, C. sinensis and Z. vulgaris are equally effective in preventing the growth of T. gallinae trophozoites in the culture.

Development of New PCR Protocols to Detect Genetic Diversity in the Metronidazole Metabolism Genes in Susceptible and Refractory Clinical Samples of Giardia duodenalis.

PURPOSE: Investigating the genetic variation in thioredoxin reductase (TrxR) and nitroreductase (NR) genes in both treatment-resistant and -sensitive Giardia duodenalis isolates can provide valuable information in identifying potential markers of resistance to metronidazole. The rapid increase in metronidazole treatment failures suggests the presence of genetic resistance mechanisms. By analyzing these genes, researchers can gain insights into the efficacy of metronidazole against G. duodenalis and potentially develop alternative treatment strategies. In this regard, four G. duodenalis isolates (two clinically sensitive and two clinically resistant to metronidazole) were collected from various hospitals of Shiraz, southwestern Iran. METHODS: Parasitological methods including sucrose flotation and microscopy were employed for the primary confirmation of G. duodenalis cysts in stool samples. Microscopy-positive samples were approved by SSU-PCR amplification of the parasite DNA. All four positive G. duodenalis specimens at SSU-PCR were afterward analyzed utilizing designed primers based on important metronidazole metabolism genes including TrxR, NR1, and NR2. RESULTS: Unlike TrxR gene, the results of NR1 and NR2 genes showed that there are non-synonymous variations between sequences of treatment-sensitive and -resistant samples compared to reference sequences. Furthermore, the outcomes of molecular docking revealed that there is an interaction between the protein sequence and spatial shape of treatment-resistant samples and metronidazole in the position of serine amino acid based on the NR1 gene. CONCLUSION: This issue can be one of the possible factors involved in the resistance of Giardia parasites to metronidazole. To reach more accurate results, a large sample size along with simulation and advanced molecular dynamics investigations are needed.

Synthesis of new trypanocidal agents from the hybridisation of metronidazole and eugenol analogues.

Nitroimidazole compounds are well-known bioactive substances, and the structural activity relationship has been reported whereby the position of the nitro group within the imidazole ring has a large influence on the activity. This study focuses on synthesising new trypanocidal agents from the hybridisation of metronidazole with different natural phenols (eugenol, dihydroeugenol and guaiacol). Two different coupling methodologies have been explored in order to analyse the influence of the connector on bioactivity: i) classic direct esterification (AD compounds) and ii) "click" chemistry using a triazole connector (AC compounds). The in vitro trypanocidal tests show good results for both AC and AD hybrid compounds against both epimastigote and trypomastigote forms of T. cruzi. In silico studies showed positive data for most of the synthesised compounds and, in general present low toxicological risks. The AC compounds present lower ClogP (lipophilicity) values than those found for the AD series and higher TPSA (topological polar surface area) values, suggesting lower lipophilicity may be related to the presence of the triazole connector. The AD series compounds have higher Drug Score values than the AC series derivatives, suggesting better general properties for a pharmacological action.

Comparison of Antibacterial Efficacy of Triple Antibiotic-Loaded Hydrogel Versus Modified Triple Antibiotic-Loaded Hydrogel as Intracanal Medicament Against Enterococcus faecalis: An In vitro Study.

OBJECTIVE: Triple antibiotic paste (TAP) is known to have an essential role in the success of endodontic treatment by eliminating pathogens from the root canal system. Unfortunately, it causes discolouration and cytotoxicity at high concentrations. The objective of this research was to assess and compare the antimicrobial effectiveness of various concentrations (1 mg, 5 mg, 10 mg) of TAP, TAP hydrogel (TAPH), M-TAP, and M-TAP hydrogel (MTAPH) against Enterococcus faecalis. METHODS: The agar well diffusion method was used to assess the antibiotic sensitivity of the following intracanal medicaments: TAP (ciprofloxacin, metronidazole, and minocycline) mixed in a ratio of 1: 1: 1; TAPH, M-TAP (ciprofloxacin, metronidazole, and amoxicillin), M-TAPH and plain hydrogel. Each tested medicament was individually evaluated for its antimicrobial activity against Enterococcus faecalis. Structural and topographical characterisation were analysed using a Scanning Electron Microscope (SEM) and interpreted using ImageJ software. A microdilution broth test was performed to examine the minimum inhibitory concentration and minimum bactericidal concentration (MBC) of M-TAP and TAP. RESULTS: Except for the plain hydrogel, M-TAP and hydrogel and TAP and hydrogel showed significantly varied inhibitory zones at different concentrations. M-TAPH showed the highest mean zone of inhibition of 21.6, 33.33 and 38.0 mm at a concentration of 1, 5, and 10 mg/mL when compared to TAPH, which showed a mean zone of inhibition of 3.3 mm,12.3 mm, 21.3 mm at the respective concentrations. The MIC study shows that more than 75% of Enterococcus faecalis growth was inhibited by M-TAP at a concentration of 5 µg/mL, whereas TAP showed inhibition at a concentration of 35 µg/mL. MBC results indicate that almost 99.9% of the bacterial population was killed at a concentration of 100 µg/mL (10-1) for TAP and 10 µg/mL (10-2) for M-TAP. CONCLUSION: The antibacterial efficacy of M-TAP was significantly higher than TAP. Application of M-TAP at lower doses is advised to overcome the disadvantages seen with TAP.

Exploring novel pyrazole-nitroimidazole hybrids: Synthesis and antiprotozoal activity against the human pathogen trichomonas vaginalis.

Trichomoniasis, a prevalent sexually transmitted infection (STI) caused by the protozoan Trichomonas vaginalis, has gained increased significance globally. Its relevance has grown in recent years due to its association with a heightened risk of acquiring and transmitting the human immunodeficiency virus (HIV) and other STIs. In addition, many publications have revealed a potential link between trichomoniasis and certain cancers. Metronidazole (MTZ), a nitroimidazole compound developed over 50 years ago, remains the first-choice drug for treatment. However, reports of genotoxicity and side effects underscore the necessity for new compounds to address this pressing global health concern. In this study, we synthesized ten pyrazole-nitroimidazoles 1(a-j) and 4-nitro-1-(hydroxyethyl)-1H-imidazole 2, an analog of metronidazole (MTZ), and assessed their trichomonacidal and cytotoxic effects. All compounds 1(a-j) and 2 exhibited IC50 values ≤ 20 µM and ≤ 41 µM, after 24 h and 48 h, respectively. Compounds 1d (IC50 5.3 µM), 1e (IC50 4.8 µM), and 1i (IC50 5.2 µM) exhibited potencies equivalent to MTZ (IC50 4.9 µM), the reference drug, after 24 h. Notably, compound 1i showed high anti-trichomonas activity after 24 h (IC50 5.2 µM) and 48 h (IC50 2.1 µM). Additionally, all compounds demonstrated either non-cytotoxic to HeLa cells (CC50 > 100 µM) or low cytotoxicity (CC50 between 69 and 100 µM). These findings suggest that pyrazole-nitroimidazole derivatives represent a promising heterocyclic system, serving as a potential lead for further optimization in trichomoniasis chemotherapy.

Analysis of Helicobacter pylori resistance in patients with different gastric diseases.

Helicobacter pylori (H. pylori) resistance is the most important risk factor for eradication failure. However, in most regions, antibiotic resistance rates of H. pylori in patients with different types of gastric mucosal lesions are still unclear. An 8-year clinical retrospective cohort study involving 2847 patients was performed. In this study, we first summarized and compared the resistance status of H. pylori in different years, ages, sexes, and gastric diseases. The resistance profiles of amoxicillin (AMX), clarithromycin (CLR), levofloxacin (LVX) and furazolidone (FR) and their changing trends in the clinic were described. Then, multiple antibiotic resistance in different gastric diseases and years were described and compared. The relationship between proton pump inhibitor (PPI) medication history and antibiotic resistance in H. pylori was also explored. Finally, an antibiotic resistance risk model was constructed for clinical resistance risk prediction. The overall resistance rates of AMX, CLR, LVX and FR in gastric diseases were 8.18%, 38.11%, 43.98%, and 13.73%, respectively. The mono resistance, double resistance, triple resistance, and quadruple resistance rates were 30.17%, 25.96%, 6.46%, and 0.63%, respectively. Compared with the period from 2014 to 2016, the rates of mono-resistance and multiple resistance all showed relatively downward trends in the past 5 years. Factors including age, sex, type of gastric lesions and recent PPI treatment history are associated with the antibiotic resistance rate of H. pylori. Atrophic gastritis is an important clinical feature of high-risk antibiotic resistance in H. pylori-infected patients. Patients with atrophic gastritis have higher risk of resistant strains infection. In this study, our data provide the association between antibiotic resistance of H. pylori and gastritis pattern, which indicate the higher risk of resistant strain infection if the patients with atrophic gastritis, PPI history and older age.

Primary antibiotic resistance of Helicobacter pylori in India over the past two decades: A systematic review.

BACKGROUND: Helicobacter pylori antibiotic resistance has undergone vast changes in the last two decades. No systematic review has been done on the prevalence of antibiotic resistant H. pylori in India in the last two decades. We evaluated the pattern of resistance rates across various regions of India. MATERIALS AND METHODS: A systematic review of the geographical variations in antibiotic resistance pattern of H. pylori was conducted using PubMed, Google Scholar, Web of Science, Science Direct, etc. for articles published between January 1, 2000 and May 30, 2023. Random effects-model-based Cochran's Q test, I2 statistics, and chi-squared tests were used to measure heterogeneity. RESULTS: The overall resistance was highest against metronidazole (77.65%) followed by amoxicillin (37.78%), levofloxacin (32.8%), clarithromycin (35.64%), furazolidone (12.03%), and tetracycline (11.63%). 14.7% of the H. pylori isolates were multi-drug resistant. Under meta-analysis of each antibiotic, high heterogeneity levels were observed having I2 ranges from 86.53% to 97.70% at p < 0.0001. In sub-group analysis, Metronidazole has a stable rate of resistance as compared to other antibiotics. Other antibiotics have had a downtrend in the last 5 years except for levofloxacin, which has had an uptrend in the resistance rate for the past 5 years. Hence, one should avoid using metronidazole for any kind of first-line treatment. CONCLUSIONS: Metronidazole resistance is high in most regions of India except Assam and Mumbai while clarithromycin is found to be ineffective in South India, Gujarat, and Kashmir. As compared to other antibiotics, resistance to amoxicillin is generally low except in certain regions (Hyderabad, Chennai, and the Gangetic belt of North India). Tetracycline and Furazolidone have the least resistance rates and should be part of anti- H. pylori regimens. The resurgence of high single and multidrug resistance to the commonly used drugs suggests the need for newer antibiotics and regular resistance surveillance studies.

Antibacterial activity of ibezapolstat against antimicrobial-resistant clinical strains of Clostridioides difficile.

Antimicrobial resistance is emerging in clinical strains of Clostridioides difficile. Ibezapolstat (IBZ) is a DNA polymerase IIIC inhibitor that has completed phase II clinical trials. IBZ has potent in vitro activity against wild-type, susceptible strains but its effect on C. difficile strains with reduced susceptibility to metronidazole (MTZ), vancomycin (VAN), or fidaxomicin (FDX) has not been tested. The primary objective of this study was to test the antibacterial properties of IBZ against multidrug-resistant C. difficile strains. The in vitro activity, bactericidal, and time-kill activity of IBZ versus comparators were evaluated against 100 clinical strains of which 59 had reduced susceptibility to other C. difficile antibiotics. Morphologic changes against a multidrug resistance strain were visualized by light and scanning electron microscopy. The overall IBZ MIC50/90 values (µg/mL) for evaluated C. difficile strains were 4/8, compared with 2/4 for VAN, 0.5/1 for FDX, and 0.25/4 for MTZ. IBZ MIC50/90 values did not differ based on non-susceptibility to antibiotic class or number of classes to which strains were non-susceptible. IBZ bactericidal activity was similar to the minimum inhibitory concentration (MIC) and maintained in wild-type and non-susceptible strains. Time-kill assays against two laboratory wild-type and two clinical non-susceptible strains demonstrated sustained IBZ activity despite reduced killing by comparator antibiotics for IBZ and VAN non-susceptible strains. Microscopy visualized increased cell lengthening and cellular damage in multidrug-resistant strains exposed to IBZ sub-MIC concentrations. This study demonstrated the potent antibacterial activity of IBZ against a large collection of C. difficile strains including multidrug-resistant strains. This study highlights the therapeutic potential of IBZ against multidrug-resistant strains of C. difficile.

Electro-antibacterial therapy (EAT) to enhance intracellular bacteria clearance in pancreatic cancer cells.

Intratumoral bacteria have been implicated in driving tumor progression, yet effective treatments to modulate the tumor microbiome remain limited. In this study, we investigate the use of electroporation in combination with metronidazole to enhance the clearance of intracellular Fusobacterium nucleatum within pancreatic cancer cells. We explore various parameters, including electric field strength, pulse width, and pulse number to assess the permeability of pancreatic cancer cells infected with F. nucleatum, compared to non-infected cells of the same type. We subsequently quantify the clearance of intracellular bacteria when these pulsing schemes are applied to a suspension of infected pancreatic cancer cells in the presence of metronidazole. Our results reveal distinct differences in cell permeability between infected and non-infected cells, identifying a unique biophysical marker for host cells infected with F. nucleatum. We demonstrate that the combinatorial use of electroporation and metronidazole significantly enhances the delivery of metronidazole into host cells, leading to more effective clearance of intracellular F. nucleatum compared to independent treatments; we term this novel approach Electro-Antibacterial Therapy (EAT). EAT holds promise as an innovative strategy for addressing intratumoral bacteria in pancreatic cancer, other malignancies, and potentially treatment-resistant infections, offering new avenues for therapeutic intervention.

Development of a new antibiotic-induced dysbiosis model of the canine colonic microbiota.

As in humans, antibiotics are widely used in dogs to treat gastrointestinal infections, contributing to the global burden of antimicrobial resistance on both human and animal health. Close contact between pets and their owners can lead to horizontal transfer of gut microbes, including transmission of antibiotic resistance. Nevertheless, until now, the impact of antibiotics on the canine gut microbiota has been poorly described. The aim of this study was to adapt the canine mucosal artificial colon (CANIM-ARCOL) model, reproducing the main nutritional, physicochemical and microbial parameters found in the large intestine of the dog to simulate an antibiotic-induced perturbation. Following initial investigation of five antibiotic cocktails at in-field doses, a 5-day regimen of metronidazole/enrofloxacin (ME) was selected for further model development. Two CANIM-ARCOL bioreactors were inoculated with a faecal sample (n=2 donors) and run in parallel for 26 days under control or antibiotic conditions. ME reduced microbial diversity and induced major shifts in bacterial populations, leading to a state of dysbiosis characterized by an increase in the relative abundance of Streptococcaceae, Lactobacillaceae and Enterobacteriaceae, and a decrease in the relative abundance of Bacteroidaceae, Fusobacteriota and Clostridiaceae. Overall, mucus-associated microbiota were less impacted by antibiotics than luminal microbes. Microbial alterations were associated with drastic decreases in gas production and short-chain fatty acid concentrations. Finally, the model was well validated through in-vitro-in-vivo comparisons in a study in dogs. The CANIM-ARCOL model provides a relevant platform as an alternative to in-vivo assays for an in-depth understanding of antibiotic-microbiota interactions and further testing of restoration strategies at individual level.

Effects of Preoperative Quadruple Therapy for Helicobacter pylori on Bariatric Surgery Metabolic Outcomes.

PURPOSE: To assess the effects of Helicobacter pylori (HP) eradication with an omeprazole, clarithromycin, amoxicillin, and metronidazole (OCAM) regimen on the metabolic profile and weight loss 12 months after bariatric surgery (BS). METHODS: Retrospective analysis of a prospective cohort of patients with morbid obesity undergoing BS. HP presence was tested preoperatively by gastric biopsy and treated with OCAM when positive. Short-term metabolic outcomes and weight loss were evaluated. RESULTS: HP infection was detected in 75 (45.7%) of the 164 patients included. OCAM effectiveness was 90.1%. HP-negative patients had a greater reduction in glucose levels at 3 (-14.6 ± 27.5 mg/dL HP-treated vs -22.0 ± 37.1 mg/dL HP-negative, p=0.045) and 6 months (-13.7 ± 29.4 mg/dL HP-treated vs -26.4 ± 42.6 mg/dL HP-negative, p= 0.021) and greater total weight loss (%TWL) at 6 (28.7 ± 6.7% HP-treated vs 30.45 ± 6.48% HP-negative, p= 0.04) and 12 months (32.21 ± 8.11% HP-treated vs 35.14 ± 8.63% HP-negative, p= 0.023). CONCLUSIONS: Preoperative treatment with OCAM has been associated to poorer glycemic and weight loss outcomes after BS. More research is needed on the influence of OCAM on gut microbiota, and in turn, the effect of the latter on metabolic and weight loss outcomes after BS.

Combination of Doxycycline with Metronidazole Protects against Pyroptosis in Rats with Endometritis through the Modulation of TLR4/NF-κB Pathway.

BACKGROUND: Endometritis is a condition usually resulted from the bacterial infection of uterus, causing pelvic disease, sepsis, shock, uterine necrosis and even death if it is inappropriately treated. The aim of this study is to explore the pathogenesis of endometritis, and investigate whether the combination of doxycycline and metronidazole offers stronger protection against lipopolysaccharide (LPS)-induced endometritis, and decipher more about the mechanisms underlying endometritis-related pyroptosis. METHODS: Sprague-Dawley (SD) rats were divided into five groups (n = 8 per group): control, model, metronidazole, doxycycline, and combination groups. In control group, the rats were injected with saline, while in other groups, lipopolysaccharide was injected into uterus of the rats to establish endometritis. Hematoxylin-eosin (H&E) staining was performed as part of the histopathological examination of endometrium. The integrity of chromatin and pyroptosis were evaluated by terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labeling (TUNEL) assay. Western blot and quantitative real-time reverse-transcription polymerase chain reaction (qRT-PCR) were performed to ascertain the activation of toll-like receptors (TLR4)/nuclear factor-kappa B (NF-κB) pathway by detecting protein levels of phosphorylated p50 (p-p50)/p50, phosphorylated nuclear factor-kappa B (p-NF-κB)/NF-κB, phosphorylated IkappaB (p-IκB), and TLR4 protein and mRNA. Development of pyroptosis was also detected by determining the levels of caspase-1 and caspase-5 through Western blot and qRT-PCR. Enzyme-linked immunosorbent assay (ELISA) was used to detect levels of interleukin (IL)-1ß, IL-18, IL-2, IL-4, IL-6 and tumor necrosis factor alpha (TNF-α), and flow cytometry was adopted to determine T-helper (Th)1 and Th2 cell percentage to assess the extent of pyroptosis and Th1/Th2 imbalance. RESULTS: The uterine of the model group exhibited pathological alterations and higher degree of cell apoptosis. Compared with the control rats, model group showed lower protein levels of p-p50/p50 (p < 0.001), p-NF-κB/NF-κB (p < 0.001), p-IκB (p < 0.001), and TLR4 protein (p < 0.001) and mRNA (p < 0.001). Elevated levels of caspase-1 (p < 0.001), caspase-5 (p < 0.001), IL-1ß (p < 0.001), IL-18 (p < 0.001), IL-2 (p < 0.01), TNF-α (p < 0.05) and Th1/Th2 (p < 0.001) as well as reduced levels of IL-4 (p < 0.05) and IL-6 (p < 0.01) were observed in the model group, which could however be reversed by metronidazole (p < 0.01) or doxycycline (p < 0.01), with a more significant effect detected if a combination of the two drugs was administered (p < 0.01). CONCLUSIONS: The combination of doxycycline and metronidazole protects against rat endometritis by inhibiting TLR4/NF-κB pathway-mediated inflammation and suppressing pyroptosis.