In Vitro Framework to Assess the Anti-Helicobacter pylori Potential of Lactic Acid Bacteria Secretions as Alternatives to Antibiotics.
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ABSTRACT:
SUBMITTER: Whiteside SA
PROVIDER: S-EPMC8198849 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:We previously demonstrated (M. M. Exner, P. Doig, T. J. Trust, and R. E. W. Hancock, Infect. Immun. 63:1567-1572, 1995) that Helicobacter pylori has at least one nonspecific porin, HopE, which has a low abundance in the outer membrane but forms large channels. H. pylori is relatively susceptible to most antimicrobial agents but less susceptible to the polycationic antibiotic polymyxin B. We demonstrate here that H. pylori is able to take up higher basal levels of the hydrophobic fluorescent probe 1-N-phenylnaphthylamine (NPN) than Pseudomonas aeruginosa or Escherichia coli, consistent with its enhanced susceptibility to hydrophobic agents. Addition of polymyxin B led to a further increase in NPN uptake, indicative of a self-promoted uptake pathway, but it required a much higher amount of polymyxin B to yield a 50% increase in NPN uptake in H. pylori (6 to 8 microg/ml) than in P. aeruginosa or E. coli (0.3 to 0.5 microg/ml), suggesting that H. pylori has a less efficient self-promoted uptake pathway. Since intrinsic resistance involves the collaboration of restricted outer membrane permeability and secondary defense mechanisms, such as periplasmic beta-lactamase (which H. pylori lacks) or efflux, we examined the possible role of efflux in antibiotic susceptibility. We had previously identified in H. pylori 11637 the presence of portions of three genes with homology to potential restriction-nodulation-division (RND) efflux systems. It was confirmed that H. pylori contained only these three putative RND efflux systems, named here hefABC, hefDEF, and hefGHI, and that the hefGHI system was expressed only in vivo while the two other RND systems were expressed both in vivo and in vitro. In uptake studies, there was no observable energy-dependent tetracycline, chloramphenicol, or NPN efflux activity in H. pylori. Independent mutagenesis of the three putative RND efflux operons in the chromosome of H. pylori had no effect on the in vitro susceptibility of H. pylori to 19 antibiotics. These results, in contrast to what is observed in E. coli, P. aeruginosa, and other clinically important gram-negative bacteria, suggest that active efflux does not play a role in the intrinsic resistance of H. pylori to antibiotics.
| S-EPMC89666 | biostudies-literature
Project description:Helicobacter pylori infection is a WHO class 1 carcinogenic factor of gastric adenocarcinoma. In the past decades, many studies have demonstrated the increasing trend of antibiotic resistance and pointed out the necessity of new effective treatment. This study was aimed at identifying phytochemicals that can inhibit H. pylori and possibly serve as adjuvant treatments. Here, in silico molecular docking and drug-like properties analyses were performed to identify potential inhibitors of urease, shikimate kinase and aspartate-semialdehyde dehydrogenase. These three enzymes are targets of the treatment of H. pylori. Susceptibility and synergistic testing were performed on the selected phytochemicals and the positive control antibiotic, amoxicillin. The in-silico study revealed that oroxindin, rosmarinic acid and verbascoside are inhibitors of urease, shikimate kinase and aspartate-semialdehyde dehydrogenase, respectively, in which, oroxindin has the highest potency against H. pylori, indicated by a minimum inhibitory concentration (MIC) value of 50 μg/mL. A combination of oroxindin and amoxicillin demonstrated additive effects against H. pylori, as indicated by a fractional inhibitory concentration (FIC) value of 0.75. This study identified phytochemicals that deserve further investigation for the development of adjuvant therapeutic agents to current antibiotics against H. pylori.
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Project description:Secretion of proteins by Helicobacter pylori may contribute to gastric inflammation and epithelial damage. An in vitro analysis was designed to identify proteins released by mechanisms other than nonspecific lysis. The radioactivity of proteins in the supernatant was compared with that of the intact organism by two-dimensional gel phosphorimaging following a 4-h pulse-chase. The ratio of the amount of UreB, a known cytoplasmic protein, in the supernatant to that in the pellet was found to be 0.25, and this was taken as an index of lysis during the experiments (n = 6). Ratios greater than that of UreB were used to distinguish proteins that were selectively released into the medium. Thus, proteins enriched more than 10-fold in the supernatant compared to UreB were identified by mass spectrometry. Sixteen such proteins were present in the supernatant: VacA; a conserved secreted protein (HP1286); putative peptidyl cis-trans isomerase (HP0175); six proteins encoded by HP0305, HP0231, HP0973, HP0721, HP0129, and HP0902; thioredoxin (HP1458); single-stranded-DNA-binding 12RNP2 precursor (HP0827); histone-like DNA-binding protein HU (HP0835); ribosomal protein L11 (HP1202); a putative outer membrane protein (HP1564); and outer membrane proteins Omp21 (HP0913) and Omp20 (HP0912). All except HP0902, thioredoxin, HP0827, HP0835, and HP1202 had a signal peptide. When nalidixic acid, a DNA synthesis inhibitor, was added to inhibit cell division but not protein synthesis, to decrease possible contamination due to outer membrane shedding, two outer membrane proteins (Omp21 and Omp20) disappeared from the supernatant, and the amount of VacA also decreased. Thus, 13 proteins were still enriched greater than 10-fold in the medium after nalidixic acid treatment, suggesting these were released specifically, possibly by secretion. These proteins may be implicated in H. pylori-induced effects on the gastric epithelium.
| S-EPMC151949 | biostudies-literature
Project description:BackgroundResistance of Helicobacter pylori (H. pylori) to antibiotics is increasing worldwide. To determine the status of H. pylori resistance and its patterns in clinical patients, an investigation utilizing susceptibility testing for commonly used antibiotics was needed.MethodsTotal of 2283 H. pylori strains were collected from 2013 to 2016. The resistance and its patterns of these strains were tested by agar dilution method. The resistance rate and minimal inhibition concentration (MIC) in different gender groups were also analyzed.ResultsThe overall resistance rates were as following: amoxicillin (1.58%), clarithromycin (22.73%), levofloxacin (24.75%), furazolidone (1.49%), doxycycline (9.20%), cefetamet (97.20%), ceftriaxone (49.60%), cefuroxime (25.20%), gentamicin (3.73%), azithromycin (85.60%), rifampicin (2.80%), metronidazole (92.53%), ornidazole (94.27%), tinidazole (87.20%), ciprofloxacin (43.20%), and moxifloxacin (38.53%). There were only 64.08% strains pan-susceptible to amoxicillin, clarithromycin, levofloxacin, and furazolidone, followed by mono resistance (23.17%), double resistance (11.13%), triple resistance (1.36%), and quadruple resistance (0.26%). Significant differences in the resistance rate and MIC were also observed in different gender groups.ConclusionAntibiotic resistance trends of H. pylori is increasing in clinical patients. With the increasing resistance, it is imperative to individualized therapy based on the results of drug susceptibility testing.
| S-EPMC6817019 | biostudies-literature
Project description:Helicobacter pylori is a gastrointestinal pathogen with high prevalence that harms human health. Studies have shown that H. pylori can form antibiotic-tolerant biofilms, which may interfere with the efficacy of clinical antibiotic therapy. Probiotics can antagonize planktonic and biofilm pathogen cells and thus may play an auxiliary role in H. pylori antibiotic therapy. However, the effects of different probiotic strains and antibiotic combinations on H. pylori biofilms need to be further investigated. We determined the cell viability of H. pylori mature biofilms after treatment with Lactobacillus plantarum LN66 cell-free supernatant (CFS), clarithromycin (CLR), and levofloxacin (LVX) alone or in combination by the XTT method. Biofilm cells were observed by scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Subsequently, protein and polysaccharide concentrations in biofilm extracellular polymeric substances (EPSs) were quantitatively detected by the Bradford method and the phenol-sulfate method. The results showed that LN66 CFS had an eradication effect on mature H. pylori biofilm. When used in combination with CLR, LN66 CFS significantly attenuated the eradication effect of CLR on biofilms; in contrast, when used in combination with LVX, LN66 CFS enhanced the disrupting effect of LVX. We speculate that the different effects of CFS and antibiotic combinations on biofilms may be related to changes in the content of proteins and polysaccharides in EPS and that the combination of CFS and CLR might promote the secretion of EPS, while the combination of CFS and LVX might have the opposite effect. Accordingly, we suggest that supplementation with L. plantarum LN66 may provide additional help when therapy involving LVX is used for clinical H. pylori biofilm eradication, whereas it may impair CLR efficacy when therapy involving CLR is used.
| S-EPMC7923053 | biostudies-literature
Project description:BackgroundHelicobacter pylori infection is exceptionally prevalent and is considered to be acquired primarily early in life through person-to-person transmission within the family. H. pylori is a genetically diverse bacterial species, which may facilitate adaptation to new hosts and persistence for decades. The present study aimed to explore the genetic diversity of clonal isolates from a mother and her three children in order to shed light on H. pylori transmission and host adaptation.ResultsTwo different H. pylori strains and strain variants were identified in the family members by PCR-based molecular typing and sequencing of five loci. Genome diversity was further assessed for 15 isolates by comparative microarray hybridizations. The microarray consisted of 1,745 oligonucleotides representing the genes of two previously sequenced H. pylori strains. The microarray analysis detected a limited mean number (+/- standard error) of divergent genes between clonal isolates from the same and different individuals (1 +/- 0.4, 0.1%, and 3 +/- 0.3, 0.2%, respectively). There was considerable variability between the two different strains in the family members (147 +/- 4, 8%) and for all isolates relative to the two sequenced reference strains (314 +/- 16, 18%). The diversity between different strains was associated with gene functional classes related to DNA metabolism and the cell envelope.ConclusionThe present data from clonal H. pylori isolates of family members do not support that transmission and host adaptation are associated with substantial sequence diversity in the bacterial genome. However, important phenotypic modifications may be determined by additional genetic mechanisms, such as phase-variation. Our findings can aid further exploration of H. pylori genetic diversity and adaptation.
| S-EPMC1899507 | biostudies-literature
Project description:Transient gut microbiota alterations have been reported after antibiotic therapy for Helicobacter pylori. However, alteration in the gut virome after H. pylori eradication remains uncertain. Here, we apply metagenomic sequencing to fecal samples of 44 H. pylori-infected patients at baseline, 6-week (N = 44), and 6-month (N = 33) after treatment. Following H. pylori eradication, we discover contraction of the gut virome diversity, separation of virome community with increased community difference, and shifting towards a higher proportion of core virus. While the gut microbiota is altered at 6-week and restored at 6-month, the virome community shows contraction till 6-month after the treatment with enhanced phage-bacteria interactions at 6-week. Multiple courses of antibiotic treatments further lead to lower virus community diversity when compared with treatment naive patients. Our results demonstrate that H. pylori eradication therapies not only result in transient alteration in gut microbiota but also significantly alter the previously less known gut virome community.
| S-EPMC10110541 | biostudies-literature
Project description:Helicobacter pylori is the primary pathogen responsible for causing gastroduodenal ulcers and stomach cancer. The standard treatment for H. pylori typically involves a combination of antibiotics and acid-reducing medications. However, the recurrence of ulcers is closely linked to the emergence of antibiotic resistance in H. pylori, necessitating the development of alternative drugs. This report focuses on the investigation of artesunate as a potential alternative to reduce antibiotic use and enhance effectiveness against H. pylori. Unfortunately, commercial artesunate is available in an acid form, which has poor solubility, especially in gastric acid fluid. The aim of this study is to utilize a water-soluble formulation of artesunate called dry emulsion formulation (ADEF) and combine it with amoxicillin to eradicate H. pylori. In vitro studies were conducted to evaluate the activity of ADEF against H. pylori and determine its inhibitory concentrations. In addition, pharmacokinetic parameters of orally administered ADEF and native artesunate were investigated in rats for in vivo studies. The results showed that when combined with amoxicillin and pantoprazole, ADEF exhibited effectiveness against H. pylori. It is worth noting that the solubility of ADEF in gastric acid appears to be a critical factor for achieving successful treatment. Consequently, ADEF could be considered a promising candidate for H. pylori therapy.
| S-EPMC10342150 | biostudies-literature
Project description:BACKGROUND:Smectite can serve as a drug delivery system and gentamicin-intercalated smectite hybrids are expected to supersede the standard therapy for Helicobacter pylori eradication. The aim of this study was to confirm whether the minimum inhibitory concentration (MIC) of aminoglycosides applied as smectite hybrids remained low against recently isolated H. pylori strains. MATERIALS AND METHODS:A total of 140 strains were collected for a minimum period of 3 years. Antimicrobial susceptibility tests were performed, and the MICs of eight antibiotics (amoxicillin, clarithromycin, metronidazole, tetracycline, levofloxacin, gentamicin, netilmicin, and tobramycin) were determined by using the Epsilometer test and following the European Committee on Antimicrobial Susceptibility Testing recommendations. RESULTS:The resistance rate of clarithromycin was high, up to 30.7%, although it is a major antimicrobial agent used in standard therapy. The MIC₅₀ and MIC₉₀ of gentamicin (0.25 mg/L and 0.75 mg/L) and netilmicin (0.19 mg/L and 0.75 mg/L) were lower than other alternative therapies for H. pylori eradication. In clarithromycin-resistant strains, the MIC₅₀ was 0.25 mg/L and the MIC₉₀ was 1 mg/L for gentamicin; for netilmicin, the values were 0.25 mg/L and 0.75 mg/L, respectively. CONCLUSION:Through the use of gentamicin and netilmicin, which have low MICs for H. pylori, aminoglycoside-intercalated smectite hybrids are expected to emerge as a new standard therapy for H. pylori eradication.
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Project description:High-resolution imaging of biomolecular condensates in living cells is essential for correlating their properties to those observed through in vitro assays. However, such experiments are limited in bacteria due to resolution limitations. Here we present an experimental framework that probes the formation, reversibility, and dynamics of condensate-forming proteins in Escherichia coli as a means to determine the nature of biomolecular condensates in bacteria. We demonstrate that condensates form after passing a threshold concentration, maintain a soluble fraction, dissolve upon shifts in temperature and concentration, and exhibit dynamics consistent with internal rearrangement and exchange between condensed and soluble fractions. We also discover that an established marker for insoluble protein aggregates, IbpA, has different colocalization patterns with bacterial condensates and aggregates, demonstrating its potential applicability as a reporter to differentiate the two in vivo. Overall, this framework provides a generalizable, accessible, and rigorous set of experiments to probe the nature of biomolecular condensates on the sub-micron scale in bacterial cells.
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