Hollow versatile Ag@Pt alloy nanoparticles with nanozyme activity for detection and photothermal sterilization of Helicobacter pylori.

In view of a large number of people infected with Helicobacter pylori (H. pylori) with great harm followed, there is an urgent need to develop a non-invasive, easy-to-operate, and rapid detection method, and to identify effective sterilization strategies. In this study, highly specific nanoprobes with nanozyme activity, Ag@Pt nanoparticles (NPs) with the antibody, were utilized as a novel lateral flow immunoassay (LFIA). The optical label (Ag@Pt NPs) was enhanced by the introduction of the chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) and compared with a gold nanoparticles (Au NPs) optical label. Under the optimal condition, Ag@Pt-LFIA and TMB-enhanced Ag@Pt-LFIA for H. pylori were successfully established, two of which were over twofold and 100-fold more sensitive than conventional visual Au NP-based LFIA, respectively. Furthermore, Ag@Pt NPs with the antibody irradiated with NIR laser (808 nm) at a power intensity of 550 mW/cm2 for 5 min exhibited a remarkable antibacterial effect. The nanoprobes could close to bacteria through effective interactions between antibodies and bacteria, thereby benefiting photothermal sterilization. Overall, Ag@Pt NPs provide promising applications in pathogen detection and therapeutic applications.

Selective photodynamic inactivation of Helicobacter pylori by a cationic benzylidene cyclopentanone photosensitizer - an in vitro and ex vivo study.

The rise in the antibiotic resistance rate of Helicobacter pylori has led to an increasing eradication failure of this carcinogenic bacterial pathogen worldwide. This underlines the need for alternative antibacterial strategies against H. pylori infection. Antimicrobial photodynamic therapy (aPDT) is a promising non-pharmacological antibacterial technology. In this study, the selective killing activities of three benzylidene cyclopentanone (BCP) photosensitizers (Y1, P1 and P3) towards H. pylori over normal human gastric epithelial GES-1 cells were evaluated and the ex vivo photodynamic inactivation effect was preliminarily assessed on twelve H. Pylor-infected mice. Results showed that under the irradiation of 24 J/cm2 532 nm laser, Y1, P1 and P3 at 2.5 µM induced a 3-log10 reduction of H. pylori CFU (99.9% killing). Confocal images showed that P3, unlike Y1 and P1, could not be uptaken by GES-1 cells. P3 at 2.5 to 20 µM showed not significant (p > 0.05) phototoxicity to GES-1 cells, nevertheless, Y1 and P1 under the same concentrations exhibited remarkable phototoxicity to GES-1 cells. In the co-culture of H. pylori and GES-1 cells, P3 at 2.5 µM led to a complete eradication of H. pylori under the irradiation of 24 J/cm2 532 nm laser. While for the GES-1 cells, no significant (p > 0.05) phototoxicity was observed under the same aPDT dosage. The ex vivo experiments showed that P3 mediated aPDT resulted in 82.4% to 100% reduction of H. pylori CFU without damaging the gastric mucosa. To sum up, P3 is a promising anti-H. pylori photosensitizer with the ability to selectively photo-inactivate H. pylori while sparing normal gastric tissues.

Blue light emitting diodes enhance the antivirulence effects of Curcumin against Helicobacter pylori.

Introduction. Growing concern about the increasing frequency of resistance of Helicobacter pylori to the available antimicrobial agents worldwide has encouraged the search for new strategies in treating and eradicating H. pylori infections. Endoscopic blue-light therapy has been used in patients with H. pylori gastritis with limited success due to subsequent repopulation with H. pylori. Clinical trials using Curcumin could not eradicate infection either.Aim. We studied the effect of blue light emitting diodes (LEDs) in conjunction with Curcumin on H. pylori, since this has not been previously reported.Methodology. We examined the effect of Curcumin with and without irradiation with blue LEDs on the viability of H. pylori and four key factors important for colonization and establishment of H. pylori infection, namely urease production, motility, adhesion and biofilm formation.Results. We found that a combination of Curcumin and blue LEDs caused significant reductions in viability, urease production, motility, haemagglutination activity, as well as increased disruption of mature preformed biofilms of H. pylori, in comparison to Curcumin alone (P<0.0001), at sublethal concentrations of Curcumin.Conclusion. Targeting the virulence factors of H. pylori with blue LED photoactivated Curcumin would theoretically cripple this pathogen from colonizing and causing tissue damage and perhaps overcome the problem of repopulation with H. pylori that often occurs following endoscopic blue-light therapy.

Blue light emitting diodes cripple Helicobacter pylori by targeting its virulence factors.

BACKGROUND: The endogenous photosensitizing porphyrins in Helicobacter pylori (H. pylori), make blue light therapy an attractive addition to the armamentarium in the war against this very prevalent and difficult to treat infectious agent. METHODS: In the current study we examined in vitro the effect of blue LED (Light Emitting Diode) irradiation for 1-6 minutes on the viability and virulence factors of H. pylori, which allow this microorganism to colonize and establish infection. Specifically, we examined the effects of blue LED on urease production, motility, adhesion and biofilm formation. RESULTS: We found that exposure to blue LED for 1-6 minutes significantly decreased the viability of H. pylori and caused decreased urease activity, as well as, swarming motility. Furthermore, blue LED irradiation for 6 minutes caused greater than 50% disruption of preformed mature biofilms of H. pylori, relative to controls. CONCLUSIONS: Collectively, the results of our in-vitro study indicate that therapy with blue LED may be an added weapon in the eradication of H. pylori by targeting the virulence factors of this very common pathogen. We envisage that phototherapy will have an adjuvant effect on conventional anti-H. pylori therapy, especially considering its efficacy in biofilm disruption and the fact that microorganisms are unlikely to develop resistance as a result of the multi-target effects.

Side effects of intra-gastric photodynamic therapy: an in vitro study.

Since many years it has been acknowledged that some bacterial species, among which H. pylori, P. aeruginosa, P. acnes accumulate endogenous photosensitizers (PS) in the form of porphyrins. This makes antibacterial photodynamic therapy (PDT) easier to perform due to the possible avoidance of external PS. In this study, we focus on gastric infections associated with the presence of Helicobacter pylori (H. pylori), known to accumulate and release both protoporphyrin IX (PPIX) and coproporphyrins. PDT versus H. pylori can be carried out by modified endoscopes or by new ingestible luminous devices under development. In both cases of in vitro and in vivo applications, either for therapy (PDT) or diagnosis, scientific literature lacks studies on the possible side-effects of light treatments on the surrounding tissues. To this aim we evaluated in vitro side-effects due to a possible intrinsic photosensitivity of gastric mucosa or to a photosensitization by the PS released from the bacterium itself. Photo-toxicity studies were conducted on the AGS cell line (ATCC® CRL-1739™), commonly used as a model for the stomach mucosa tissue, considering PPIX as the photosensitizing agent. After first evaluations of PPIX dark toxicity, its uptake and accumulation sites, photo-toxicity tests were conducted using a LED light source peaked at 400 nm, by varying both PPIX concentration (50 nM - 2 µM) and light dose in the range 0.6-13 J/cm2, representing different treatment procedures found in literature. The oxidative stress consequent to irradiation was investigated both in terms of ROS production and assessment of the activity of enzymes involved in ROS-related biological mechanisms. A significant phototoxic effect was found only for PPIX concentration > 100 nM for all tested light doses. This indicates that the evaluated photo-treatments do not cause side effects even with the sensitization due to PPIX released by the bacteria.

Treatment of Helicobacter pylori with dielectric barrier discharge plasma causes UV induced damage to genomic DNA leading to cell death.

Gastrointestinal endoscopy is an important tool for the indentification and treatment of disorders of the gastrointestinal tract. However, nosocomial infections of Helicobacter pylori have been linked to the use of contaminated endoscopes. Disinfectants such as glutaraldehyde, ortho-phthalaldehyde and peracetic acid are generally used in the reprocesssing of endoscopes, but these chemicals are hazardous to human health. Thus, safer reprocessing and disinfecion methods are needed. In this study, we applied a dielectric barrier discharge (DBD) plasma torch for inactivation of H. pylori to investigate a potential new methodology to disinfect endoscopes. Suspensions of H. pylori in 10% glycerol were subjected to the DBD plasma torch, which reduced the viable cell count to undetectable levels after 2 min of treatment. Furthermore, urease activity of H. pylori was eliminated after 2 min-plasma treatment, while plasma-treatment reduced the intact DNA of H. pylori in a time-dependent manner. Next, we examined several potential bactericidal factors produced by the DBD plasma torch. Two min-plasma treatment resulted in a small temperature rise (4 °C), ultraviolet radiation (UV) generation, and the production of hydrogen peroxide. H. pylori samples were then exposed to equivalent levels of each of these factors in turn. Our results showed that treatment with heat and hydrogen peroxide at the levels produced after 2-min of plasma treatment did not efficiently inactivate H. pylori, whereas exposure to UV had a significant bactericidal effect. Taken together, UV generated by the plasma torch may be crucial for efficient inactivation of H. pylori by damaging the bacterial DNA.

Anti-proliferation effect of blue light-emitting diodes against antibiotic-resistant Helicobacter pylori.

BACKGROUND AND AIM: Infection by Helicobacter pylori is implicated in a wide range of upper gastrointestinal diseases. Owing to the rapid emergence of antibiotic-resistant strains of H. pylori, the development of novel treatment modalities for antibiotic-resistant H. pylori infection is a key priority. Blue light-emitting diodes (LED) may represent a unique option owing to their antimicrobial effect. In this study, we aimed to evaluate the anti-proliferative effect of blue LED against antibiotic-resistant H. pylori. METHODS: Ten antibiotic-resistant strains and one sensitive H. pylori strain were used in this study. After irradiation by blue LED along time course, the viability of H. pylori was evaluated by enumerating colony forming units. Morphological changes in H. pylori were observed using a scanning electron microscope. Reductase activity was measured as an indicator of bacterial cellular activity. Total reactive oxygen species was monitored using fluorescence intensity and fluorescence microscope imaging. RESULTS: After irradiation by blue LED, the numbers of H. pylori in all the strains were significantly reduced compared with control group. The H. pylori exhibited a short rod-shaped morphology after irradiation; no such change was observed in H. pylori not exposed to blue LED. Re-irradiation of surviving strain after the initial irradiation also exhibited the same anti-proliferation effect. After blue LED irradiation, bacterial cellular activity was lower, and total reactive oxygen species production was significantly higher in blue LED group, compared with that in control. CONCLUSIONS: Blue LED could be a new treatment to eradicate infection with antibiotic-resistant H. pylori.

Iodine-131 in Helicobacter pylori-positive patients: preliminary accidental finding and in differentiated thyroid cancer.

OBJECTIVE: To investigate the eradicate effect of Helicobacter pylori in differentiated thyroid cancer patients who underwent I therapy. MATERIALS AND METHODS: A total of 130 patients with differentiated thyroid cancer underwent I treatment. None of the patients had a history of stomach-related diseases. The C-urea breath test (UBT) was carried out for H. pylori examination before I treatment. The cut-off value for C-UBT was 4%. For H. pylori test-positive patients, a second C-UBT was performed 4-6 weeks after I treatment. T-tests were used to compare the difference before and 4-6 weeks after I treatment. RESULTS: A total of 42 of 130 (32.31%) patients were H. pylori positive. The average value of H. pylori was 28.36% before I, whereas the value was 18.18% after I therapy. A significant decrease in C-UBT was found after I treatment compared with before therapy (P<0.01). CONCLUSION: Our preliminary data showed that I exerts certain eradication effects on H. pylori, which could provide a new approach to multidrug resistance pathogens. As the exact molecular mechanism of this phenomenon is still unclear, future clinical applications of this anecdotal finding require further assessments.

Capsule Design for Blue Light Therapy against Helicobacter pylori.

A photo-medical capsule that emits blue light for Helicobacter pylori treatment was described in this paper. The system consists of modules for pH sensing and measuring, light-emitting diode driver circuit, radio communication and microcontroller, and power management. The system can differentiate locations by monitoring the pH values of the gastrointestinal tract, and turn on and off the blue light according to the preset range of pH values. Our experimental tests show that the capsule can operate in the effective light therapy mode for more than 32 minutes and the wireless communication module can reliably transmit the measured pH value to a receiver located outside the body.

In vitro studies of different irradiation conditions for Photodynamic inactivation of Helicobacter pylori.

Helicobacter pylori (HP) infections are considered to be the main cause for chronic gastritis and gastric ulcers, whereby more than half of the world's population is nowadays infected. The increased use of antibiotics is leading to an enhanced resistance. Photodynamic inactivation of bacteria seems to be a potential alternative for antibiotic therapies. In our study we used the photosensitizer Chlorin e6 (Ce6) in combination with red light-emitting diodes to inactivate HP in vitro. Ce6 uptake is determined by spectroscopy. Furthermore diverse experiments of different concentrations in the range of 0-100 µM of the photosensitizer and exposure times up to 300 s are carried out in order to find optimal irradiation parameters (wavelength: 660 nm, power density: 9 mW/cm(2), absorbed dose: up to 2.7 J/cm(2)). The data show a significant reduction after already a few seconds of illumination, even with a low Ce6 concentration in the sub-µM-region. At a concentration of 100 µM a nearly total inactivation (6-log10-reduction) of HP was achieved within 60s of irradiation.

Synergistic in vitro photodynamic antimicrobial activity of methylene blue and chitosan against Helicobacter pylori 26695.

BACKGROUND: Photodynamic therapy (PDT) is a method for killing cells (bacterial, fungal and cancer cells) or virus using photosensitizers (PS) and light of various wavelengths. In vitro PDT using endoscopic light against H. pylori was effective at a concentration of 0.2mg/mL of MB. The purpose of this study was to increase the effect of photodynamic modality against H. pylori by addition of chitosan to MB. METHODS: The bactericidal effect was measured by counting viable cells after PDT. The degree of damage to DNA was confirmed using alkaline gel electrophoresis. Cellular DNA damage was demonstrated by ethidium bromide monoazide-quantitative polymerase chain reaction (EMA-qPCR). RESULTS: In the groups treated with either 0.04 mg/mL MB alone or 0.02 mg/mL MB with endoscopic light for 15 min, viable cells were decreased approximately tenfold. The group treated with 0.04 mg/mL of MB with light, showed more effective bactericidal activity than 0.02 mg/mL of MB treatment. By 0.05% chitosan pre-treatment followed with 0.04 mg/mL of MB and light irradiation, viable cells were decreased 10(7)-fold. The DNA damage caused by PDT as demonstrated by alkaline gel electrophoresis was greater in the MB plus chitosan-treated group than in control and MB-treated groups. Cellular DNA damage demonstrated by EMA-qPCR was also greater in the group treated with MB plus chitosan than in the MB-treated group. CONCLUSION: The bactericidal effects with PDT using MB were increased with the concentration of photosensitizer and chitosan treatment, peculiarly before endoscopic light irradiation.

The effect of radioactive iodine treatment on 14C urea breath test results in patients with hyperthyroidism.

PURPOSE: Radioactive Iodine therapy (RAIT) plays a major role in the treatment of hyperthyroidism. In addition to the thyroid gland, significant amounts of radioactive iodine are maintained in the stomach. The aim of this study was to determine if RAIT has any effect on Helicobacter pylori infection, based on the C urea breath test (UBT). MATERIALS AND METHODS: The study included 85 patients with hyperthyroidism scheduled to undergo RAIT and 69 hyperthyroid subjects in whom methimazole treatment was planned. All subjects had pretreatment-positive UBT results, and the test was repeated on the first and third months after RAIT and methimazole treatment. RESULTS: After a mean RAIT dose of 15 mCi (range, 10-20 mCi), UBT became negative in 13 (15.3%) of 85 patients on the first month and 18 (21.2%) of 85 patients on the third month. All subjects treated with methimazole remained UBT positive on the first and third months of methimazole treatment (100%). Reduction in the number of UBT-positive patients on both the first and the third months after RAIT was statistically significant (P < 0.001). Distribution of hyperthyroidism etiologies and thyroid autoantibody levels in subjects with UBT that became negative and in subjects with UBT that remained positive were similar in the RAIT group (P > 0.05). Urea breath test negativity rates did not differ according to the radioiodine dose. CONCLUSIONS: Our findings indirectly showed that RAIT might have an antimicrobial effect on H. pylori. Clinical applications of this beneficial effect of RAIT on H. pylori should be further evaluated.

In vitro bactericidal effects of near-ultraviolet light from light-emitting diodes on Helicobacter pylori.

We investigated whether near-ultraviolet light emitted from light-emitting diodes (LEDs) effects Helicobacter pylori viability and whether this new method can potentially apply to eradication therapy. Three H. pylori strains were used for near-ultraviolet (UV) LED irradiation experiments. Viability of isolates exposed to near-UV light was compared with controls by counting colony forming units. A time-dependent bactericidal effect of near-UV light was definitely observed. LED irradiation with near-UV light showed effective bactericidal activity against H. pylori strains. Eradication therapy with LED might provide a new avenue of treatment in patients refractory to eradication due to antibiotic resistance and/or adverse effects of antibiotics.

Helicobacter pylori inactivation and virulence gene damage using a supported sensitiser for photodynamic therapy.

About half of the world's population is currently infected with Helicobacter pylori, which is involved in the development of several gastro-duodenal pathologies. The increasing number of antibiotic resistance reduces the effectiveness of the first-line therapy, so new strategies to improve the H. pylori eradication rates are needed. Antimicrobial Photodynamic Therapy (APDT) benefits from photogenerated reactive oxygen species, such as singlet oxygen, which inactivate microorganisms by means of photosensitising dyes and visible light. Therefore, it could be a suitable alternative for H. pylori eradication in the gastro-duodenal tract, particularly in patients infected with antibiotic resistant strains. We evaluated APDT against H. pylori, in vitro, using a new photosensitising material (PSM) based on a ruthenium(II) complex covalently bound to micrometric glass beads. Five H. pylori isolates (classified according to cagA genotype, and metronidazole-clarithromycin resistance) were used. Bacteria were mixed with the PSM and incubated in the dark or illuminated by blue light. Aliquots (min 1', 2', 5', 15' and 30') were cultured and colonies were counted after 2-3 days. A 99.99999% decrease was detected in the number of colonies in the irradiated wells where the bacterium was mixed with the PSM, compared to non-illuminated wells or with irradiated wells without PSM. It was also confirmed that DNA is a molecular target for oxidant species released during APDT (evaluated by alkaline gel electrophoresis after endonuclease III incubation, ureC and cagA RT-PCR, and bacterial fingerprint). Results were independent of cagA gene and antibiotic resistances.

Comparison of in vitro photodynamic antimicrobial activity of protoporphyrin IX between endoscopic white light and newly developed narrowband endoscopic light against Helicobacter pylori 26695.

Helicobacter pylori might be readily affected with photodynamic therapy (PDT) by weak wavelengths, because it has few repair genes. Recently, gastrointestinal endoscopy emitting specific wavelengths (narrowband imaging, NBI) has been developed for the early detection of tumors. Coincidentally, its wavelength (415 nm) is very similar to the wavelength (410 nm) that activates protoporphyrin IX (PpIX) as a photosensitizer (PS). Therefore, we studied in vitro PDT against H. pylori using NBI and conventional white light (WL) according to low or high concentration of PpIX along with exposure time. The bactericidal effects, the degree of oxidative DNA damage and membrane integrity of H. pylori after PDT were evaluated. In the control, the numbers of viable cells remained constant during the experiment. Viable cells after PDT using both endoscopic light irradiation, were decreased approximately 10(3) - 10(5) fold at low concentration of PpIX and below 0.80 × 10 at high concentration of PpIX. Only membrane damage after PDT was observed microscopically in H. pylori without DNA injury. Conclusively, either the bactericidal effect in high concentration or the decrease of bacterial loading in low concentration of PpIX, would be expected with PDT using endoscopic light (NBI or WL).

Blue light for infectious diseases: Propionibacterium acnes, Helicobacter pylori, and beyond?

Blue light, particularly in the wavelength range of 405-470 nm, has attracted increasing attention due to its intrinsic antimicrobial effect without the addition of exogenous photosensitizers. In addition, it is commonly accepted that blue light is much less detrimental to mammalian cells than ultraviolet irradiation, which is another light-based antimicrobial approach being investigated. In this review, we discussed the blue light sensing systems in microbial cells, antimicrobial efficacy of blue light, the mechanism of antimicrobial effect of blue light, the effects of blue light on mammalian cells, and the effects of blue light on wound healing. It has been reported that blue light can regulate multi-cellular behavior involving cell-to-cell communication via blue light receptors in bacteria, and inhibit biofilm formation and subsequently potentiate light inactivation. At higher radiant exposures, blue light exhibits a broad-spectrum antimicrobial effect against both Gram-positive and Gram-negative bacteria. Blue light therapy is a clinically accepted approach for Propionibacterium acnes infections. Clinical trials have also been conducted to investigate the use of blue light for Helicobacter pylori stomach infections and have shown promising results. Studies on blue light inactivation of important wound pathogenic bacteria, including Staphylococcus aureus and Pseudomonas aeruginosa have also been reported. The mechanism of blue light inactivation of P. acnes, H. pylori, and some oral bacteria is proved to be the photo-excitation of intracellular porphyrins and the subsequent production of cytotoxic reactive oxygen species. Although it may be the case that the mechanism of blue light inactivation of wound pathogens (e.g., S. aureus, P. aeruginosa) is the same as that of P. acnes, this hypothesis has not been rigorously tested. Limited and discordant results have been reported regarding the effects of blue light on mammalian cells and wound healing. Under certain wavelengths and radiant exposures, blue light may cause cell dysfunction by the photo-excitation of blue light sensitizing chromophores, including flavins and cytochromes, within mitochondria or/and peroxisomes. Further studies should be performed to optimize the optical parameters (e.g., wavelength, radiant exposure) to ensure effective and safe blue light therapies for infectious disease. In addition, studies are also needed to verify the lack of development of microbial resistance to blue light.

DprB facilitates inter- and intragenomic recombination in Helicobacter pylori.

For naturally competent microorganisms, such as Helicobacter pylori, the steps that permit recombination of exogenous DNA are not fully understood. Immediately downstream of an H. pylori gene (dprA) that facilitates high-frequency natural transformation is HP0334 (dprB), annotated to be a putative Holliday junction resolvase (HJR). We showed that the HP0334 (dprB) gene product facilitates high-frequency natural transformation. We determined the physiologic roles of DprB by genetic analyses. DprB controls in vitro growth, survival after exposure to UV or fluoroquinolones, and intragenomic recombination. dprB ruvC double deletion dramatically decreases both homologous and homeologous transformation and survival after exposure to DNA-damaging agents. Moreover, the DprB protein binds to synthetic Holliday junction structures rather than double-stranded or single-stranded DNA. These results demonstrate that the dprB product plays important roles affecting inter- and intragenomic recombination. We provide evidence that the two putative H. pylori HJRs (DprB and RuvC) have overlapping but distinct functions involving intergenomic (primarily DprB) and intragenomic (primarily RuvC) recombination.

Genome sequencing reveals a phage in Helicobacter pylori.

UNLABELLED: Helicobacter pylori chronically infects the gastric mucosa in more than half of the human population; in a subset of this population, its presence is associated with development of severe disease, such as gastric cancer. Genomic analysis of several strains has revealed an extensive H. pylori pan-genome, likely to grow as more genomes are sampled. Here we describe the draft genome sequence (63 contigs; 26× mean coverage) of H. pylori strain B45, isolated from a patient with gastric mucosa-associated lymphoid tissue (MALT) lymphoma. The major finding was a 24.6-kb prophage integrated in the bacterial genome. The prophage shares most of its genes (22/27) with prophage region II of Helicobacter acinonychis strain Sheeba. After UV treatment of liquid cultures, circular DNA carrying the prophage integrase gene could be detected, and intracellular tailed phage-like particles were observed in H. pylori cells by transmission electron microscopy, indicating that phage production can be induced from the prophage. PCR amplification and sequencing of the integrase gene from 341 H. pylori strains from different geographic regions revealed a high prevalence of the prophage (21.4%). Phylogenetic reconstruction showed four distinct clusters in the integrase gene, three of which tended to be specific for geographic regions. Our study implies that phages may play important roles in the ecology and evolution of H. pylori. IMPORTANCE: Helicobacter pylori chronically infects the gastric mucosa in more than half of the human population, and while most of the infected individuals do not develop disease, H. pylori infection doubles the risk of developing gastric cancer. An abundance and diversity of viruses (phages) infect microbial populations in most environments and are important mediators of microbial diversity. Our finding of a 24.6-kb prophage integrated inside an H. pylori genome and the observation of circular integrase gene-containing DNA and phage-like particles inside cells upon UV treatment demonstrate that we have discovered a viable H. pylori phage. The additional finding of integrase genes in a large proportion of screened isolates of diverse geographic origins indicates that the prevalence of prophages may have been underestimated in H. pylori. Since phages are important drivers of microbial evolution, the discovery should be important for understanding and predicting genetic diversity in H. pylori.

Biochemical and cellular characterization of Helicobacter pylori RecA, a protein with high-level constitutive expression.

Helicobacter pylori is a bacterial pathogen colonizing half of the world's human population. It has been implicated in a number of gastric diseases, from asymptomatic gastritis to cancer. It is characterized by an amazing genetic variability that results from high mutation rates and efficient DNA homologous recombination and transformation systems. Here, we report the characterization of H. pylori RecA (HpRecA), a protein shown to be involved in DNA repair, transformation, and mouse colonization. The biochemical characterization of the purified recombinase reveals activities similar to those of Escherichia coli RecA (EcRecA). We show that in H. pylori, HpRecA is present in about 80,000 copies per cell during exponential growth and decreases to about 50,000 copies in stationary phase. The amount of HpRecA remains unchanged after induction of DNA lesions, suggesting that HpRecA is always expressed at a high level in order to repair DNA damage or facilitate recombination. We performed HpRecA localization analysis by adding a Flag tag to the protein, revealing two different patterns of localization. During exponential growth, RecA-Flag presents a diffuse pattern, overlapping with the DAPI (4',6-diamidino-2-phenylindole) staining of DNA, whereas during stationary phase, the protein is present in more defined areas devoid of DAPI staining. These localizations are not affected by inactivation of competence or DNA recombination genes. Neither UV irradiation nor gamma irradiation modified HpRecA localization, suggesting the existence of a constitutive DNA damage adaptation system.