In Silico Screening and In Vitro Assessment of Natural Products with Anti-Virulence Activity against Helicobacter pylori

Helicobacter pylori is one of the most frequent human pathogens and a leading etiological agent of various gastric diseases. As stringent response, coordinated by a SpoT protein, seems to be crucial for the survivability of H. pylori, the main goal of this article was to use in silico computational studies to find phytochemical compounds capable of binding to the active site of SpoT from H. pylori and confirm the ability of the most active candidates to interfere with the virulence of this bacterium through in vitro experiments. From 791 natural substances submitted for the virtual screening procedure, 10 were chosen and followed for further in vitro examinations. Among these, dioscin showed the most interesting parameters (the lowest MIC, the highest anti-biofilm activity in static conditions, and a relatively low stimulation of morphological transition into coccoids). Therefore, in the last part, we extended the research with a number of further experiments and observed the ability of dioscin to significantly reduce the formation of H. pylori biofilm under Bioflux-generated flow conditions and its capacity for additive enhancement of the antibacterial activity of all three commonly used antibiotics (clarithromycin, metronidazole, and levofloxacin). Based on these results, we suggest that dioscin may be an interesting candidate for new therapies targeting H. pylori survivability and virulence.

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Vitamin B6 Is Required for Full Motility and Virulence in Helicobacter pylori

mBio ◽

10.1128/mbio.00112-10 ◽

2010 ◽

Vol 1 (3) ◽

Cited By ~ 25

Author(s):

Alexandra Grubman ◽

Alexandra Phillips ◽

Marie Thibonnier ◽

Maria Kaparakis-Liaskos ◽

Chad Johnson ◽

...

Keyword(s):

Gene Expression ◽

Helicobacter Pylori ◽

Virulence Factors ◽

Bacterial Pathogens ◽

Therapeutic Targets ◽

Vitamin B ◽

Human Pathogens ◽

Chronic Colonization ◽

H Pylori

ABSTRACTDespite recent advances in our understanding of howHelicobacter pyloricauses disease, the factors that allow this pathogen to persist in the stomach have not yet been fully characterized. To identify new virulence factors inH. pylori, we generated low-infectivity variants of a mouse-colonizingH. pyloristrain using the classical technique ofin vitroattenuation. The resulting variants and their highly infectious progenitor bacteria were then analyzed by global gene expression profiling. The gene expression levels of five open reading frames (ORFs) were significantly reduced in low-infectivity variants, with the most significant changes observed for ORFs HP1583 and HP1582. These ORFs were annotated as encoding homologs of theEscherichia colivitamin B6biosynthesis enzymes PdxA and PdxJ. Functional complementation studies withE. coliconfirmedH. pyloriPdxA and PdxJ to bebona fidehomologs of vitamin B6biosynthesis enzymes. Importantly,H. pyloriPdxA was required for optimal growthin vitroand was shown to be essential for chronic colonization in mice. In addition to having a well-known metabolic role, vitamin B6is necessary for the synthesis of glycosylated flagella and for flagellum-based motility inH. pylori. Thus, for the first time, we identify vitamin B6biosynthesis enzymes as novel virulence factors in bacteria. Interestingly,pdxAandpdxJorthologs are present in a number of human pathogens, but not in mammalian cells. We therefore propose that PdxA/J enzymes may represent ideal candidates for therapeutic targets against bacterial pathogens.IMPORTANCEApproximately half of the world’s population is infected withH. pylori, yet howH. pyloribacteria establish chronic infections in human hosts remains elusive. From gene array studies, we identified two genes as representing potentially novel colonization factors forH. pylori. These genes encoded enzymes involved in the synthesis of vitamin B6, an important molecule for many metabolic reactions in living organisms. Little is currently known regarding vitamin B6biosynthesis in human pathogens. We showed that mutantH. pyloribacteria lacking an enzyme involved inde novovitamin B6biosynthesis, PdxA, were unable to synthesize motility appendages (flagella) and were unable to establish chronic colonization in mice. Thus, this work identifies vitamin B6biosynthesis enzymes as novel virulence factors for bacterial pathogens. Interestingly, a number of human pathogens, but not their mammalian hosts, possess these genes, which suggests that Pdx enzymes may represent ideal candidates for new therapeutic targets.

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In Silico and In Vitro Anti-Helicobacter Pylori Effects of Combinations of Phytochemicals and Antibiotics

Molecules ◽

10.3390/molecules24193608 ◽

2019 ◽

Vol 24 (19) ◽

pp. 3608 ◽

Cited By ~ 2

Author(s):

Pedro Fong ◽

Chon-Hou Hao ◽

Chi-Cheng Io ◽

Pou-Io Sin ◽

Li-Rong Meng

Keyword(s):

Helicobacter Pylori ◽

In Silico ◽

Inhibitory Concentration ◽

Positive Control ◽

Shikimate Kinase ◽

Semialdehyde Dehydrogenase ◽

Class 1 ◽

H Pylori ◽

Potential Inhibitors

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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Synthesis, In Silico Studies, and Evaluation of Syn and Anti Isomers of N-Substituted Indole-3-carbaldehyde Oxime Derivatives as Urease Inhibitors against Helicobacter pylori

Molecules ◽

10.3390/molecules26216658 ◽

2021 ◽

Vol 26 (21) ◽

pp. 6658

Author(s):

Ishani P. Kalatuwawege ◽

Medha J. Gunaratna ◽

Dinusha N. Udukala

Keyword(s):

Helicobacter Pylori ◽

Gastrointestinal Tract ◽

In Silico ◽

Hydroxylamine Hydrochloride ◽

In Silico Studies ◽

Urease Enzyme ◽

Oxime Derivatives ◽

Urease Inhibitory ◽

H Pylori

Gastrointestinal tract infection caused by Helicobacter pylori is a common virulent disease found worldwide, and the infection rate is much higher in developing countries than in developed ones. In the pathogenesis of H. pylori in the gastrointestinal tract, the secretion of the urease enzyme plays a major role. Therefore, inhibition of urease is a better approach against H. pylori infection. In the present study, a series of syn and anti isomers of N-substituted indole-3-carbaldehyde oxime derivatives was synthesized via Schiff base reaction of appropriate carbaldehyde derivatives with hydroxylamine hydrochloride. The in vitro urease inhibitory activities of those derivatives were evaluated against that of Macrotyloma uniflorum urease using the modified Berthelot reaction. Out of the tested compounds, compound 8 (IC50 = 0.0516 ± 0.0035 mM) and compound 9 (IC50 = 0.0345 ± 0.0008 mM) were identified as the derivatives with potent urease inhibitory activity with compared to thiourea (IC50 = 0.2387 ± 0.0048 mM). Additionally, in silico studies for all oxime compounds were performed to investigate the binding interactions with the active site of the urease enzyme compared to thiourea. Furthermore, the drug-likeness of the synthesized oxime compounds was also predicted.

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Functional Analysis of the cag Pathogenicity Island in Helicobacter pylori Isolates from Patients with Gastritis, Peptic Ulcer, and Gastric Cancer

Infection and Immunity ◽

10.1128/iai.72.2.1043-1056.2004 ◽

2004 ◽

Vol 72 (2) ◽

pp. 1043-1056 ◽

Cited By ~ 87

Author(s):

Steffen Backert ◽

Tobias Schwarz ◽

Stephan Miehlke ◽

Christian Kirsch ◽

Christian Sommer ◽

...

Keyword(s):

Gastric Cancer ◽

Helicobacter Pylori ◽

Peptic Ulcer ◽

Pathogenicity Island ◽

Epidemiological Studies ◽

Disease Development ◽

Gastric Diseases ◽

H Pylori ◽

Gastric Cancer Patients

ABSTRACT Helicobacter pylori is the causative agent of a variety of gastric diseases, but the clinical relevance of bacterial virulence factors is still controversial. Virulent strains carrying the cag pathogenicity island (cagPAI) are thought to be key players in disease development. Here, we have compared cagPAI-dependent in vitro responses in H. pylori isolates obtained from 75 patients with gastritis, peptic ulcer, and gastric cancer (n = 25 in each group). AGS gastric epithelial cells were infected with each strain and assayed for (i) CagA expression, (ii) translocation and tyrosine phosphorylation of CagA, (iii) c-Src inactivation, (iv) cortactin dephosphorylation, (v) induction of actin cytoskeletal rearrangements associated with cell elongation, (vi) induction of cellular motility, and (vii) secretion of interleukin-8. Interestingly, we found high but similar prevalences of all of these cagPAI-dependent host cell responses (ranging from 56 to 80%) among the various groups of patients. This study revealed CagA proteins with unique features, CagA subspecies of various sizes, and new functional properties for the phenotypic outcomes. We further showed that induction of AGS cell motility and elongation are two independent processes. Our data corroborate epidemiological studies, which indicate a significant association of cagPAI presence and functionality with histopathological findings in gastritis, peptic ulcer, and gastric cancer patients, thus emphasizing the importance of the cagPAI for the pathogenicity of H. pylori. Nevertheless, we found no significant association of the specific H. pylori-induced responses with any particular patient group. This may indicate that the determination of disease development is highly complex and involves multiple bacterial and/or host factors.

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The Role of E-cadherin in Helicobacter pylori-Related Gastric Diseases

Current Drug Metabolism ◽

10.2174/1389200219666180625113010 ◽

2019 ◽

Vol 20 (1) ◽

pp. 23-28

Author(s):

Yunzhan Zhang ◽

Danyan Li ◽

Yunkai Dai ◽

Ruliu Li ◽

Yong Gao ◽

...

Keyword(s):

Helicobacter Pylori ◽

Gastric Mucosa ◽

Precancerous Lesions ◽

Research Literature ◽

Gastric Diseases ◽

H Pylori ◽

The Relationship ◽

E Cadherin

Background: Helicobacter pylori (H. pylori)-related gastric diseases are a series of gastric mucosal disorders associated with H. pylori infection. Gastric cancer (GC) is widely believed to evolve from gastritis and gastric ulcer. As an important adhesion molecule of epithelial cells, E-cadherin plays a key role in the development of gastric diseases. In this review, we aim to seek the characteristic of E-cadherin expression at different stages of gastric diseases. Methods: We searched plenty of databases for research literature about E-cadherin expression in H. pylori-related gastric diseases, and reviewed the relationship of E-cadherin and H. pylori, and the role of E-cadherin at different stages of gastric diseases. Results: H. pylori was shown to decrease E-cadherin expression by various ways in vitro, while most of clinical studies have not found the relationship between H. pylori and E-cadherin expression. It is defined that poor outcome of GC is related to loss expression of E-cadherin, but it is still unclear when qualitative change of E-cadherin expression in gastric mucosa emerges. Conclusion: Expression level of E-cadherin in gastric cells may be a consequence of injury factors and body’s selfrepairing ability. More studies on E-cadherin expression in gastric mucosa with precancerous lesions need to be performed, which may be potential and useful for early detection, prevention and treatment of GC.

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The Effects of Sulglycotide on the Adhesion and the Inflammation of Helicobacter Pylori

International Journal of Environmental Research and Public Health ◽

10.3390/ijerph17082918 ◽

2020 ◽

Vol 17 (8) ◽

pp. 2918

Author(s):

Ji Yeong Yang ◽

Pumsoo Kim ◽

Seok-Hoo Jeong ◽

Seong Woong Lee ◽

Yu Sik Myung ◽

...

Keyword(s):

Helicobacter Pylori ◽

Protein A ◽

Enzyme Linked Immunosorbent Assay ◽

Etiologic Factor ◽

Dilution Method ◽

Gastric Diseases ◽

Ags Cells ◽

Human Gastric Adenocarcinoma ◽

H Pylori

Helicobacter pylori (H. pylori) is a primary etiologic factor in gastric diseases. Sulglycotide is a glycopeptide derived from pig duodenal mucin. Esterification of its carbohydrate chains with sulfate groups creates a potent gastroprotective agent used to treat various gastric diseases. We investigated the inhibitory effects of sulglycotide on adhesion and inflammation after H. pylori infection in human gastric adenocarcinoma cells (AGS cells). H. pylori reference strain 60190 (ATCC 49503) was cultured on Brucella agar supplemented with 10% bovine serum. Sulgylcotide-mediated growth inhibition of H. pylori was evaluated using the broth dilution method. Inhibition of H. pylori adhesion to AGS cells by sulglycotide was assessed using a urease assay. Effects of sulglycotide on the translocation of virulence factors was measured using western blot to detect cytotoxin-associated protein A (CagA) and vacuolating cytotoxin A (VacA) proteins. Inhibition of IL-8 secretion was measured using enzyme-linked immunosorbent assay (ELISA) to determine the effects of sulglycotide on inflammation. Sulglycotide did not inhibit the growth of H. pylori, however, after six and 12 hours of infection on AGS cells, H. pylori adhesion was significantly inhibited by approximately 60% by various concentrations of sulglycotide. Sulglycotide decreased H. pylori virulence factor (CagA and VacA) translocation to AGS cells and inhibited IL-8 secretion. Sulglycotide inhibited H. pylori adhesion and inflammation after infection of AGS cells in vitro. These results support the use of sulglycotide to treat H. pylori infections.

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In Vitro Activity of Sertraline, an Antidepressant, Against Antibiotic-Susceptible and Antibiotic-Resistant Helicobacter pylori Strains

Pathogens ◽

10.3390/pathogens8040228 ◽

2019 ◽

Vol 8 (4) ◽

pp. 228 ◽

Cited By ~ 3

Author(s):

Paweł Krzyżek ◽

Roman Franiczek ◽

Barbara Krzyżanowska ◽

Łukasz Łaczmański ◽

Paweł Migdał ◽

...

Keyword(s):

Helicobacter Pylori ◽

Morphological Transformation ◽

Gastric Diseases ◽

Additive Interaction ◽

Antibiotic Resistant ◽

Minimal Inhibitory Concentrations ◽

Killing Assay ◽

H Pylori ◽

Light Fluorescence

Antibiotic resistance of Helicobacter pylori, a spiral bacterium associated with gastric diseases, is a topic that has been intensively discussed in last decades. Recent discoveries indicate promising antimicrobial and antibiotic-potentiating properties of sertraline (SER), an antidepressant substance. The aim of the study, therefore, was to determine the antibacterial activity of SER in relation to antibiotic-sensitive and antibiotic-resistant H. pylori strains. The antimicrobial tests were performed using a diffusion-disk method, microdilution method, and time-killing assay. The interaction between SER and antibiotics (amoxicillin, clarithromycin, tetracycline, and metronidazole) was determined by using a checkerboard method. In addition, the study was expanded to include observations by light, fluorescence, and scanning electron microscopy. The growth inhibition zones were in the range of 19–37 mm for discs impregnated with 2 mg of SER. The minimal inhibitory concentrations (MICs) and minimal bactericidal concentrations (MBCs) counted for 2–8 µg/mL and 4–8 µg/mL, respectively. The time-killing assay showed the time-dependent and concentration-dependent bactericidal activity of SER. Bacteria exposed to MBCs (but not sub-MICs and MICs ≠ MBCs) underwent morphological transformation into coccoid forms. This mechanism, however, was not protective because these cells after a 24-h incubation had a several-fold reduced green/red fluorescence ratio compared to the control. Using the checkerboard assay, a synergistic/additive interaction of SER with all four antibiotics tested was demonstrated. These results indicate that SER may be a promising anti-H. pylori compound.

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Identification and In Silico Characterization of Novel Helicobacter pylori Glucose-6-Phosphate Dehydrogenase Inhibitors

Molecules ◽

10.3390/molecules26164955 ◽

2021 ◽

Vol 26 (16) ◽

pp. 4955

Author(s):

Beatriz Hernández-Ochoa ◽

Gabriel Navarrete-Vázquez ◽

Rodrigo Aguayo-Ortiz ◽

Paulina Ortiz-Ramírez ◽

Laura Morales-Luna ◽

...

Keyword(s):

Helicobacter Pylori ◽

In Silico ◽

Inhibitory Effect ◽

New Drugs ◽

Antibiotics Resistance ◽

Gastric Diseases ◽

Drug Candidates ◽

Current Therapies ◽

H Pylori ◽

Glucose 6 Phosphate Dehydrogenase

Helicobacter pylori (H. pylori) is a pathogen that can remain in the stomach of an infected person for their entire life. As a result, this leads to the development of severe gastric diseases such as gastric cancer. In addition, current therapies have several problems including antibiotics resistance. Therefore, new practical options to eliminate this bacterium, and its induced affections, are required to avoid morbidity and mortality worldwide. One strategy in the search for new drugs is to detect compounds that inhibit a limiting step in a central metabolic pathway of the pathogen of interest. In this work, we tested 55 compounds to gain insights into their possible use as new inhibitory drugs of H. pylori glucose-6-phosphate dehydrogenase (HpG6PD) activity. The compounds YGC-1; MGD-1, MGD-2; TDA-1; and JMM-3 with their respective scaffold 1,3-thiazolidine-2,4-dione; 1H-benzimidazole; 1,3-benzoxazole, morpholine, and biphenylcarbonitrile showed the best inhibitory activity (IC50 = 310, 465, 340, 204 and 304 μM, respectively). We then modeled the HpG6PD protein by homology modeling to conduct an in silico study of the chemical compounds and discovers its possible interactions with the HpG6PD enzyme. We found that compounds can be internalized at the NADP+ catalytic binding site. Hence, they probably exert a competitive inhibitory effect with NADP+ and a non-competitive or uncompetitive effect with G6P, that of the compounds binding far from the enzyme’s active site. Based on these findings, the tested compounds inhibiting HpG6PD represent promising novel drug candidates against H. pylori.

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The Effects of HP0044 and HP1275 Knockout Mutations on the Structure and Function of Lipopolysaccharide in Helicobacter pylori Strain 26695

Biomedicines ◽

10.3390/biomedicines10010145 ◽

2022 ◽

Vol 10 (1) ◽

pp. 145

Author(s):

Ai-Ning Liu ◽

Kai-Wen Teng ◽

Yongyu Chew ◽

Po-Chuan Wang ◽

Tram Thi Hong Nguyen ◽

...

Keyword(s):

Helicobacter Pylori ◽

Gene Knockout ◽

Membrane Vesicles ◽

Surface Hydrophobicity ◽

Outer Membrane Vesicles ◽

Gastric Diseases ◽

Knockout Mutants ◽

H Pylori ◽

And Function

Helicobacter pylori infection is associated with several gastric diseases, including gastritis, peptic ulcer, gastric adenocarcinoma and mucosa-associated lymphatic tissue (MALT) lymphoma. Due to the prevalence and severeness of H. pylori infection, a thorough understanding of this pathogen is necessary. Lipopolysaccharide, one of the major virulence factors of H. pylori, can exert immunomodulating and immunostimulating functions on the host. In this study, the HP0044 and HP1275 genes were under investigation. These two genes potentially encode GDP-D-mannose dehydratase (GMD) and phosphomannomutase (PMM)/phosphoglucomutase (PGM), respectively, and are involved in the biosynthesis of fucose. HP0044 and HP1275 knockout mutants were generated; both mutants displayed a truncated LPS, suggesting that the encoded enzymes are not only involved in fucose production but are also important for LPS construction. In addition, these two gene knockout mutants exhibited retarded growth, increased surface hydrophobicity and autoaggregation as well as being more sensitive to the detergent SDS and the antibiotic novobiocin. Furthermore, the LPS-defective mutants also had significantly reduced bacterial infection, adhesion and internalization in the in vitro cell line model. Moreover, disruptions of the HP0044 and HP1275 genes in H. pylori altered protein sorting into outer membrane vesicles. The critical roles of HP0044 and HP1275 in LPS biosynthesis, bacterial fitness and pathogenesis make them attractive candidates for drug inventions against H. pylori infection.

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In Vitro Screening Potential Antibacterial Properties of the Greek Oregano Honey against Clinical Isolates of Helicobacter pylori

Foods ◽

10.3390/foods10071568 ◽

2021 ◽

Vol 10 (7) ◽

pp. 1568

Author(s):

Chrysoula (Chrysa) Voidarou ◽

Georgios Rozos ◽

Athanasios Alexopoulos ◽

Stavros Plessas ◽

Ioanna Mantzourani ◽

...

Keyword(s):

Helicobacter Pylori ◽

Antibacterial Properties ◽

Gastric Ulcers ◽

Mountainous Area ◽

Bacterial Strains ◽

Gastric Diseases ◽

Popular Medicine ◽

Traditional Remedy ◽

H Pylori

Oregano honey is an exceedingly rare and distinct product, not commercially available, produced by bees bred in oregano fields of alpine altitudes at the mountainous area of Epirus, Greece. In ethnic popular medicine, this product is used as a therapeutic in various gastric diseases. To test this hypothesis, 14 strains of Helicobacter pylori (H. pylori), 6 isolated from gastric ulcers and 8 from cases of clinical gastritis, were employed in the present study. The above bacterial strains were exposed to various concentrations (75% v/v, 50% v/v, 25% v/v, 12.5% v/v, and 6% v/v) of 50 oregano honey samples by using the agar well method and the inhibition zones observed around each well were recorded. Although the inhibitory zones of the H. pylori isolated from the gastric ulcers were wide enough (0–34 mm), those strains, in general, appeared more resistant than the other eight (0–58 mm). The same result was observed when the same strains were tested against six antibiotics used in clinical practice. Extracts of oregano honey were prepared by extraction with four different organic solvents. N-hexane and chloroform extracts had the most potent antibacterial action. Finally, pure oregano honey and diethyl ether extracts of honey showed significant inhibitory activity against urease secreted by the pathogen. These results strongly indicate the susceptibility of H. pylori strains to the oregano honey by more than one mode of action. Consequently, this variety of honey seems to have potential therapeutic properties against gastric ulcers and gastritis, thus explaining the preference of the locals towards this traditional remedy.

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