Update on the pathogenesis and clinical management of Helicobacter pylori gastric infection and associated diseases

Abstract

Helicobacter pylori (H. pylori) is a gram-negative, spiral-shaped, microaerophilic bacterium that infects over 43% of the global population, with higher prevalence in regions of low socioeconomic status and poor sanitation. It is transmitted mainly through oral-oral and fecal-oral routes and has evolved multiple mechanisms that allow colonization of the acidic gastric environment, including urease production, chemotaxis, and a variety of adhesins. The bacterium expresses several virulence factors that enhance its pathogenicity, such as cytotoxin-associated antigen A, vacuolating cytotoxin A, and the small regulatory RNA NikS, found to be essential for the fine-tuning of the bacterial virulence. Although many infected individuals remain asymptomatic, H. pylori infection is associated with multiple clinical outcomes, including chronic gastritis, peptic ulcers, gastric adenocarcinoma, and mucosa-associated lymphoid tissue lymphoma, all correlated to the host immune response and chronic inflammation. Diagnosis relies on both invasive and non-invasive methods, and growing antibiotic resistance poses a major challenge to treatment. New therapeutic strategies, such as tailored therapy, potassium-competitive acid blockers, and probiotics are under investigation. Vaccine development remains a key area of research, with several candidates currently in preclinical and clinical evaluation.

Key Words: Helicobacter pylori; Virulence factors; Immunological response; Tailored therapy; Treatment adjuvants

Core Tip: Helicobacter pylori (H. pylori) is a gastric pathogen that infects more than 40% of the world’s population. Pathophysiological mechanisms of infection and evolution to severe diseases range from host’s immunological responses and virulence factors, which are intricately regulated. Diagnosis and treatment strategies have been evolving in order to overcome the burden of antibiotic resistance and the need for invasive procedures. This article aims to provide an overview of H. pylori infection, highlighting important advances in the understanding of pathogenesis and new insights for effective clinical management.

INTRODUCTION

Helicobacter pylori (H. pylori) is a microaerophilic, spiraled, gram-negative bacterium that infects the stomach and leads to chronic inflammation[1]. It is transmitted mainly through the fecal-oral and oral-oral routes, and less frequently by the gastric-oral pathway, especially in patients prone to vomiting[2]. These routes correlate to its prevalence in populations with lower socioeconomic status, lack of sanitation and poor household conditions[3,4], due to the transmission between family members and the contamination of water and utensils.

Although present in more than 43% of individuals and often causing no symptoms[5], H. pylori is still considered a gastric pathogen, since it not only disrupts the gastric microbiota, but also potentially leads to severe diseases such as peptic ulcers and gastric cancer (GC)[6]. The genesis of these conditions and their differential presentation is directly linked to the host immunological response, as well as H. pylori virulence factors, such as cytotoxin-associated antigen A (CagA) and vacuolating cytotoxin A (VacA)[7]. In this regard, a recent discovery of a small non-coding RNA, NikS, has demonstrated the inner regulation of H. pylori virulence program is extremely intricate[8], allowing its persistence on the gastric environment, and also correlating to the severity of clinical outcomes.

Proper diagnosis and treatment of H. pylori-positive patients has helped prevent severe complications[9], however, the extensive use of antibiotics has compromised the efficacy of standard empirical therapies, renewing the interest in tailored therapy guided by susceptibility tests and novel adjuvants for treatment, such as potassium-competitive acid blockers (P-CABs) and probiotics, although further evidence is needed to actually ensure the implementation of these strategies[10,11], along with the remaining need for the development of prophylactic and therapeutic vaccines[12]. Overall, clinical research on H. pylori has focused on optimizing treatment strategies, while also minimizing the rise of antibiotic resistance.

This review aims to provide a comprehensive and up-to-date overview of H. pylori pathogenesis and clinical management, focusing on its mechanisms of persistence, immune evasion, and disease progression, as well as clinical management strategies, with a particular look to recent discoveries in these fields and new insights for effective clinical assessment based on current ongoing research.

PATHOGENESIS

Bacterial adherence and colonization

After encountering the gastric environment, H. pylori employs a variety of mechanisms to assure its survival and adhere to the gastric epithelium. In order to deal with the stomach's low pH, the bacterium resorts mainly to urease, a nickel-dependent enzyme capable of catalyzing the hydrolysis of urea into ammonia and carbon dioxide, thereby increasing the pH around the bacterial cells[13]. Ammonia production also serves as a wide source of nitrogen, and has shown correlation to enhanced cytokine-mediated and VacA-mediated apoptosis of gastric epithelial cells[14,15].

The reaching and adherence of H. pylori to the gastric mucosa is especially mediated by two different types of outer membrane proteins: Chemoreceptors and adhesins. Chemoreceptors, especially transducer-like proteins sense attractants and repellents in order to direct the flagellar movement towards regions with higher pH, closer to the epithelium and away from harmful chemicals[16], while adhesins mediate binding to antigens present on the gastric tissue. The intricate adaptation of H. pylori has enabled the bacterium to adhere to the healthy tissue on a first encounter, but also to survive within the inflamed environment induced by the infection itself, since it differentially expresses adhesins such as BabA[17] and HopZ[18], important for adherence with the healthy epithelium, and others like SabA[17], OipA[19] and HopQ[20], that bind to antigens overexpressed in inflamed environments. Figure 1 summarizes the most important adhesin-receptor interactions.

Figure 1 Simplified representation of Helicobacter pylori adhesin-receptor interactions. Le^b: Lewis b antigen; sLe^a: Sialyl-Lewis an antigen; sLe^x: Sialyl-Lewis x antigen; CEACAMs: Carcinoembryonic Antigen-Related Cell Adhesion Molecules; T4SS: Type IV secretion system.

Adhesion of H. pylori to the gastric epithelium is also critical for the effective functioning of its type IV secretion system (T4SS). The proper assembly and positioning of the T4SS machinery is specially mediated by HopQ and BabA, allowing the bacterium to inject effector molecules into host cells, the main one being the oncoprotein CagA[21].

Virulence factors of H. pylori

CagA: Cytotoxin-associated gene A (cagA), contained within the cag pathogenicity island, encodes the CagA oncoprotein, extremely important virulence factor of H. pylori. It is translocated into gastric epithelial cells via type IV secretion system and undergoes tyrosine phosphorylation at specific sequences known as EPIYA (Glu-Pro-Ile-Tyr-Ala) motifs located at the C-terminal of the protein[22]. These motifs are variable, consisting of different segments (A, B, C, D). While EPIYA-A and EPIYA-B are common in most H. pylori strains, EPIYA-C and EPIYA-D are characteristic of Western and East Asian strains, respectively, and the presence of EPIYA-D or multiple EPIYA-C segments is associated with an increased risk of GC development[23-26].

Older data has reported that, in Western countries, approximately 60%-70% of isolates are CagA-positive. However, in East Asian populations, notably Japan, China, and South Korea, nearly 100% of H. pylori strains contain the cagA gene[27]. Although more recent and broader studies regarding cagA prevalence in the west are scarce, a recent meta-analysis from 2024 further supports this data, reporting an 83% CagA positivity rate among H. pylori strains associated with gastroduodenal disease in the Indo-Pacific region[28]. These epidemiological differences correlate, especially, to the heightened risk of GC observed in East Asia.

Upon phosphorylation, CagA binds to SHP-2 phosphatase, leading to hyperactivation of Ras/ERK signaling and inducing cytoskeletal rearrangement, which promotes the “hummingbird phenotype”, as well as increased cell proliferation and motility through Rac1 and MAPK pathways[29,30]. Non-phosphorylated CagA can also disrupt tight junctions by interacting with E-cadherin, leading to activation of β-catenin signaling[31] and interfere with cell polarity by inhibiting PAR1/MARK family kinases[32]. Altogether, these mechanisms favor malignant transformation but are not crucial for cancer maintenance, as the pro-oncogenic actions of CagA are eventually replaced by genetic and epigenetic alterations in cancer-prone cells[33].

VacA: Another important virulence factor is the vacuolating cytotoxin A (VacA), encoded by the vacA gene, found in virtually all strains. It presents variations mainly on the signal (s), middle (m), and intermediate (i) regions[24]. The s1, i1, and m1 vacA genotypes are linked to an increased risk of GC or precancerous conditions when compared to strains carrying the type s2, i2, or m² of vacA[34], and the s1 and m1 strains have also been correlated with greater occurrence of severe gastric inflammation and ulceration[35].

VacA is able to insert into host epithelial cell membranes and form anion-selective pores that disrupt intracellular ionic gradients and induce the formation of large acidic vacuoles[36]. It also targets organelles like mitochondria, where it induces the release of cytochrome c and activates caspase-dependent apoptosis, and the endoplasmic reticulum, triggering autophagy[37]. Additionally, VacA is correlated to immune evasion: It disrupts antigen presentation by interfering with MHC class II trafficking[38], and is also able to inhibit interleukin (IL)-2 production by T cells through suppression of the NFAT signaling pathway[38].

NikS: A small nickel regulated non-coding RNA (sRNA) molecule that acts as a post-transcriptional regulator of genes related to H. pylori virulence factors, such as VacA and CagA[8]. Like other trans-encoded sRNAs, NikS acts by pairing complementary bases in regions of mRNA genes (usually closer to the 5' end in the translation initiation region), which prevents contact with ribosomes and downregulates gene expression[8,39]. The influence of nickel on NikS activity is directly related to the transcriptional regulator NikR, which responds to the availability of nickel in the environment[40]. An increased concentration of nickel induces NikR to bind to the promoter region of NikS, suppressing its expression[40,41]. Therefore, the higher the nickel concentration and the greater the suppressive action of NikR on NikS, which favors the expression of virulence factors.

Furthermore, Eisenbart et al[8] demonstrated that a variation in the number of thymine bases (Ts) in the hypermutable simple sequence repeats region can alter the transcription rate of NikS. This sequence of Ts is located immediately before the promoter, near the TATAAT box (box -10), so changes in the number of Ts alter the distance between box -10 and the binding sites of the NikR transcription factor, which can alter the affinity with RNA polymerase and, consequently, alter the transcription of NikS[8].

The modulation of CagA expression by NikS appears to be favorable for a better prognosis of H. pylori infection, since it down regulates its production[41]. Accordingly, it has been observed that GC patients have reduced NikS expression when compared to a non-cancer group[42].

Recently explored virulence aspects: In addition to the already described virulence factors, recent studies have highlighted other important components involved in H. pylori pathogenicity. For example, studies have demonstrated that bacterial strains containing Integrative and Conjugative Elements (ICEHptfs), mobile genetic regions that encode distinct type IV secretion systems apart from the cagPAI, could induce more IL-8 secretion and exacerbate inflammation and tissue damage[43,44], in accordance to epidemiological studies that have found a higher prevalence of ICEHptfs genes in strains isolated from patients with GC and duodenal ulcers[45]. However, the actual effector proteins delivered and the extent of their pathogenic mechanisms remain unknown.

Additionally, Sharafutdinov et al[46] found that a specific single-nucleotide polymorphism of the gene that encodes the High-temperature requirement A (HtrA), a bacterial serine protease that cleaves E-cadherin and occludin, is present in GC-associated strains. This gene variant promotes HtrA trimerization and enhances its proteolytic activity, leading to increased disruption of epithelial junctions, β-catenin signaling and DNA double-strand breaks, therefore favoring malignant transformations, and emerging as a potential biomarker for cancer risk.

Immunological response and chronic inflammation

The immunological response against H. pylori is initiated when toll-like receptors (TLRs) detect pathogen-associated molecular patterns[47-50], triggering the activation of nuclear factor-kappaB (NF-κB), activator protein-1, and interferon regulatory factors, resulting in the expression of pro-inflammatory cytokines[51], antimicrobial peptides, type I interferons, and the recruitment of neutrophils, macrophages, and dendritic cells[47]. In this regard, H. pylori evasion mechanisms take place: Bacterial ligands like lipopolysaccharide (LPS/Lipid A) for TLR4[52] and flagellins for TLR5[53], have evolved to exhibit low intrinsic activation potential[54], antigen presentation is also impaired by metabolites like ADP-heptose, which activate microRNAs that suppress the antigen-presenting HLA-II expression[55], and modulates phagocytic destruction by interfering with reactive oxygen species mediated and nitric oxide mediated processes[56]. Together, these mechanisms help H. pylori evade the initial immune response and persist in the gastric environment.

As mentioned, NF-κB activation is a hallmark of H. pylori infection. It is closely linked to gastric inflammation and also plays important roles in carcinogenesis[57], either by impairing apoptosis, allowing gastric epithelial cells to survive and accumulate mutations, eventually contributing to malignant transformations[58-61] and also by modulating the infiltration of leukocytes and cytokine expression in the tumor microenvironment[62]. Moreover, Frauenlob et al[63], found that H. pylori triggers a form of innate immune memory in monocytes that leads to hypersensitivity to future challenges via NF-κB pathway amplification, which may worsen conditions like gastritis and GC.

The establishment of chronic inflammation is also linked to a complex interplay of different T helper cell profiles. While Th1 cells secrete interferon-gamma and IL-12, promoting macrophage activation and contributing to mucosal damage[64,65], Th17 cells produce IL-17A, IL-21, and IL-22, which recruit neutrophils and sustain chronic inflammation. Elevated levels of these cytokines were associated with increased gastric mucosal damage and progression to atrophic gastritis[64,65]. Moreover, Tumor necrosis factor-alpha (TNF-α) was also linked to an amplification of the inflammatory response and correlates with the severity of gastric lesions[66]. Nonetheless, regulatory T cells (Treg) are also pivotal for H. pylori establishment, as a balanced immunosuppression impairs bacterial clearance[67]. A predominance of Treg in detriment to the pro-inflammatory profiles in pediatric patients has been proposed as an explanation for the higher susceptibility to infection[68], along with the milder manifestations[69], and may even play a role in the down regulation of allergic reactions[70].

Variations in the balance between Th1 and Th17 responses have also been correlated to differences in disease progression. A prospective study showed that H. pylori positive patients with duodenal ulcers presented higher gastric levels of Th1 representative cytokines, while those who evolved to GC presented a more prominent Th17 response[71], in accordance to a meta-analysis by Yu et al[72], which showed GC is associated to increased TNF-α and IL-6. Interestingly, patients who evolved to duodenal ulcers rather than GC presented increased levels of IL-27[71], highlighting its dual role, as both a Th1 enhancer and a regulator of excessive inflammation, as well as its potential therapeutic use.

Lastly, regarding the humoral response against H. pylori, antibodies are incapable of clearing the infection, due to the bacterium’s evasion mechanisms and localization within the gastric mucus layer[73]. Anti-H. pylori IgG is also not the first choice as a diagnostic tool, as it shows limited sensitivity and cannot distinguish active from past infection[74]. As for biomarker use, Anti-CagA antibodies have been studied as possible indicators of cagA-positive strains and therefore, cancer risk, with good outcomes regarding sensibility and specificity[75,76], but further research is needed for its use to be included in guidelines.

CLINICAL MANAGEMENT

Gastric histopathology

H. pylori infection is the most important cause of chronic gastritis, characterized by persistent gastric inflammation[74]. The lesions typically begin in the antrum, causing non-atrophic gastritis, and may progress to atrophic gastritis, marked by the loss of gastric glands and the predisposition to intestinal metaplasia[77]. To better stratify cancer risk, the Operative Link on Gastritis Assessment staging system integrates the severity and distribution of atrophy into a stage-based model, from stage 0 to IV, with higher stages strongly associated with GC development[78]. H. pylori-induced gastritis is dynamic, and is influenced by bacterial virulence, host response, and environmental factors[79]. Its importance lies on its role as a precursor to peptic ulcer disease and gastric malignancies, making accurate assessing and classification essential for effective management.

The treatment for H. pylori related gastritis is multifaceted. Bacterial eradication by antibiotic regimen is an essential first step, however, antimicrobial therapy alone often cannot restore the gastric mucosa and resolve active inflammation[80], which is why the standard H. pylori eradication regimens also include proton pump inhibitors for managing gastric acidity[81]. While H₂ receptor blockers and mucosal protective agents such as rebamipide are not part of standard eradication therapy, they may be useful for symptom relief following eradication, seeing they do not impact H. pylori and thus, do not interfere with short-term follow-up for treatment success[74]. Even so, mainly on atrophic variations, the gastric acid secretion may be compromised, and the administration of proton pump inhibitors may worsen problems associated with low stomach acidity, such as alterations in the gastric microbiota or reduced absorption of nutrients[82].

Even with successful eradication of H. pylori, residual atrophic changes or intestinal metaplasia often persist, highlighting the importance of continued monitoring and treatment to promote mucosal recovery and control inflammation[83].

Diagnosis and assessment of infection

For optimal accuracy in diagnosing H. pylori infection, current guidelines recommend at least two positive results in distinct diagnostic tests. However, achieving this standard can be challenging in routine clinical practice, and it is therefore not universally implemented[84]. There are invasive and non-invasive methods of diagnosing H. pylori infection.

Invasive methods: H. pylori culture is a complex and time-consuming procedure that is generally considered unnecessary in routine clinical practice, seen that other diagnostic methods, including invasive and non-invasive tests, are capable of detecting the organism in most patients with greater efficiency[85,86].

Endoscopic imaging of H. pylori-infected patients shows mucosal edema, patchy redness, diffuse erythema, nodularity, and loss of regular arrangement of collecting venules pattern (RAC), an important parameter, as recent data suggests that the integrity of RAC in endoscopic imaging can accurately rule out H. pylori infection on the gastric corpus[87-90]. The Kyoto classification of gastritis is also implied, it is a score based on 5 endoscopic findings: Atrophy, intestinal metaplasia (IM), enlarged folds, nodularity, and diffuse redness, developed to evaluate risk of GC[91]. These scores provide high specificity and accuracy in identifying current H. pylori infections[92-96].

Histopathological examination is the most informative test in case of gastroscopy. The histological characteristics of chronic active gastritis, combined with the distinctive morphology of H. pylori and associated to diagnostic methods provide a definitive diagnosis of the infection[84]. A variety of special histochemical stains are available to enhance the detection of H. pylori in gastric tissue, including Warthin-Starry, Gimenez, Genta, toluidine blue, and Giemsa, the latter being considered the most popular due to its cost-effectiveness[97].

This approach also facilitates a detailed evaluation of the associated inflammatory changes, enabling predictions about treatment efficacy, infection prognosis, and individual risk of developing GC[84]. Biopsies should ideally be obtained from areas of the gastric mucosa that appear even in apparently endoscopic normal aspects, seen that mucosal alterations, such as ulceration, erosions, atrophy, or intestinal metaplasia, may be associated with a reduced H. pylori density, which may compromise diagnostic accuracy[98-102].

The rapid urease test (RUT) leverages the high urease production of H pylori as a surrogate marker for bacterial detection in gastric biopsies[85]. Biopsies obtained from both the antrum and body of the stomach for RUT are preferred, as this approach demonstrates superior sensitivity compared to using antral biopsies alone[103]. Several diagnostic techniques are available for detecting H. pylori urease activity, including the Campylobacter-like organism (CLO) test, which utilizes a solid supporting medium containing urea and a pH-sensitive indicator[104]. Alternative methods, such as strip-based tests, are better suited for use in endoscopy clinics due to their faster turnaround time, typically providing results within approximately one hour, compared to the 24-hour incubation period required for the CLO test. Among the commercially available kits, Pyloritek (Serim Research Corp.) and Helicocheck (Institute of Immunology Co. Ltd., Tochigi, Japan) are frequently used, both demonstrating sensitivity and specificity exceeding 90% when compared[104-106].

Polymerase chain reaction (PCR) testing is another good alternative for diagnosis, because infection can be reliably detected using PCR, even at very low bacterial densities, where other diagnostic methods may yield negative results[107-109]. There are several CE-certified kits available for routine use, and, among biopsy-based tests, is considered one of the most accurate methods, particularly in patients with active bleeding[108]. PCR is not only used for detecting infection, but also for identification and analysis of pathogenic genes and specific mutations linked to antimicrobial resistance, such as the 16S rRNA[110] and 23S rRNA[111] genes.

Non-invasive methods: The Urea Breath Test (UBT) is frequently regarded as the gold standard for diagnosing H. pylori infection[112]. To conduct the UBT, the patient ingests urea labeled with either 13C or 14C isotopes. If H. pylori is present, its urease enzyme will hydrolyze the urea in the stomach, releasing labeled carbon dioxide, which can be detected in the patient's exhaled breath using either isotope ratio mass or infrared-spectrometry[85]. A recent meta-analysis demonstrated that 13C-UBT outperforms the 14C-UBT, establishing it as the preferred diagnostic method for diagnosis, while also highlighting the importance of carefully optimizing urea dosage, timing of assessment, and measurement techniques for both tests in order to improve diagnostic accuracy[113]. In the majority of studies, the sensitivity and specificity of the UBT exceed 90%, and it is also an effective tool for assessing the success of eradication therapy[114], but proton pump inhibitors and antimicrobial agents should be discontinued for 2 weeks and 4 weeks, respectively, seeing their use could limit test sensibility[115]. Another important drawback for UBT is the need for specialized equipment, especially when it comes to obtaining the labeled isotope, which might limit accessibility in some areas[116].

Detection of H. pylori antigen in stool specimens is also a form of diagnosis with high sensitivity and specificity (94.1% and 91.8%)[117], as well as an effective tool to evaluate eradication therapy. However, results may be affected due to degradation of the antigen when passing through intestine, variation of antigen excretion, and can be affected by the use of N-acetylcystein, a mucolytic agent[118].

Serology can also be employed in specific cases, including patients with bleeding ulcers, gastric atrophy, mucosa-associated lymphoid tissue lymphoma, gastric carcinoma, or those who have recently used antibiotics or proton pump inhibitors (PPIs), as these conditions can be related to low bacterial load that might result in a false negative in other tests[117]. Conversely, a major limitation for serology is that it cannot distinguish between active or past infection, as antibodies may persist long after bacterial clearance[119]. This limits its utility in monitoring eradication or in regions with high background seroprevalence.

Standard eradication therapy

Eradication therapy of H. pylori by antibiotics has been increasingly influenced by changes in the resistance profile around the world. Current resistance rates and their demographic variability point to local susceptibility testing and knowledge of the epidemiological profile of resistance as fundamental tools for successful and effective prescribing[120].

The most widely used therapeutic regimen is PPI-clarithromycin triple therapy. However, the elevated rates of resistance to clarithromycin[121,122] and frequent allergies to penicillin[123], often require alternatives, such as replacing clarithromycin with levofloxacin (250 mg-500 mg once a day), or changing to bismuth quadruple therapy (BQT). Interestingly, studies have shown that BQT has superior efficacy to PPI-clarithromycin triple therapy in bacterial eradication[124,125]. Rifabutin triple therapy[126] is also an alternative, typically used in patients who have recurrent H. pylori infections, but there is little comparative evidence to clarithromycin-triple therapy and BQT. For the treatment of children and adolescents, specific recommendations are given by the North American Societies of Pediatric Gastroenterology Hepatology and Nutrition, such as prioritizing therapy guided by antimicrobial susceptibility testing, using a high dosage of PPIs, and avoiding empirical clarithromycin regimens when susceptibility test or local resistance profile data is unavailable[127]. The therapeutic strategies according to the Maastricht VI/Florence Consensus[74] and American College of Gastroenterology guidelines[126] are summarized in Table 1.

Table 1 Therapeutic regimens for Helicobacter pylori infection according to the Maastricht consensus and the American College of Gastroenterology.

Regimen	Drugs	Dosage	Recomendations
PPI-clarithromycin triple therapy	Clarithromycin; Amoxicillin; Omeprazole	500 mg 12-hourly; 1 g 12-hourly; 20 mg 12-hourly	First line of treatment in situations of low local resistance to clarithromycin (< 15%); people without penicillin allergy; favorable individual susceptibility test for this regimen
Bismuth quadruple therapy	Metronidazole; Tetracycline; PPI; Bismuth subcitrate	500 mg 8-hourly; 500 mg 6-hourly; 120-300 mg	First line of treatment in situations of high local resistance to clarithromycin (> 15%); people with penicillin allergy; when the individual susceptibility testing isn’t available
Rifabutin triple therapy	Rifabutin; Amoxicillin; Omeprazole	50 mg 8-hourly; 1 g 8-hourly; 40 mg 8-hourly	Failure of other lines of treatment; recurrent Helicobacter pylori infections

PPI: Proton pump inhibitor.

Few changes in therapeutic strategies can be seen in other regional treaties, as they adapt, for example, to the context of pharmacological availability and the resistance profile. Coelho et al[128] recommends PPI-clarithromycin triple therapy as the first line of treatment, despite increasing resistance rates, and rifabutin should only be used in the event of availability and lack of therapeutic response to other lines of treatment, following antimicrobial susceptibility testing. As for the Chinese consensus on H. pylori infection, modified dual therapy (Amoxicillin + Potent acid suppression) is considered the most recommended empirical therapeutic strategy and there is no division into therapeutic lines, as the therapy chosen will be based on local resistance, adverse effects, costs and efficacy[129]. Fallone et al[130] recommends BQT as the first line of treatment, except when resistance rates to clarithromycin are < 15% or the eradication rate is > 85%, and the use of rifabutin triple therapy is restricted to cases of failure of the other three previous therapeutic alternatives.

Novel and emerging therapy strategies

Although test-and-treat has been the standard method of prevention of H. pylori associated diseases, it has significant impact in terms of antibiotic consumption and resistance[74]. In this context, tailored therapy (therapy guided by antibiotic susceptibility tests) emerges as a way to minimize these issues, while also increasing effectiveness. However, there are still many obstacles to that practice. Culture based methods are extremely specific but require extensive and specialized labor[131], and molecular based methods (for identification of specific gene mutations that provide resistance), even if more practical, are still expensive and may not always reflect the actual phenotypic resistance[132].

Another limitation to tailored therapy is the dependency to an invasive procedure like an endoscopy, but recent studies have demonstrated promising results for the implying of molecular based tests using stool samples[133,134]. Assessing metronidazole susceptibility is also a difficulty for both test methods[134,135]. In this regard, a recent trial has proposed the use of single-cell Raman spectrometry as an alternative, with positive outcomes[136]. Despite all the prospective findings, studies still show that tailored therapy as it is today is not necessarily more effective than the empirical regimens[137,138], highlighting the importance of broader research in order to optimize these methods, maximize eradication and minimize the rise of antibiotic resistance.

P-CABs are drugs that have been on the rise in gastroenterology as an alternative to PPIs. Its mechanism of action consists of reversible H+/K+-ATPase inhibition during the secretory stage of gastric parietal cells, achieving satisfactory effects within a few hours of administration[10]. Comparative studies of H. pylori eradication with P-CAB-based triple therapy and PPI-based triple therapy point to superior or non-inferior efficacy of the use of P-CABs, considering them as a successful first-line therapeutic alternative[139-141]. The pharmacokinetic properties of vonoprazan (one of the most studied P-CABs), such as speed of action, prolonged effect and reversibility of H+/K+-ATPase binding are also favorable characteristics for its clinical application[142]. Triple therapy could include clarithromycin, amoxicillin and 20 mg of vonoprazan twice a day, but a double therapy involving 1 g of amoxicillin every 8 hours and 20 mg of vonoprazan every 12 hours has also been studied and showed results not inferior to BQT[143].

The use of probiotics in association with the therapeutic regimens for eradication of H. pylori has also been a topic of extensive research, as both the bacteria and the antibiotics cause an impact on the gut microbiota. Yet, the data regarding this topic remains conflicting. For instance, some studies propose probiotics are only beneficial in alleviating side effects[11,144], while others suggest its use also increases eradication rates[145]. The recommendations for prescription are also unclear: While a meta-analysis by Zhang et al[146] found that using a single probiotic agent for 10 or more day retrieved better results, another meta-analysis, by Wang et al[147], demonstrated that the use of different probiotics combined, for more than 14 days, is more effective. In regard of the specific agents, the ones from the genus Lactobacilli and Saccharomyces seem to be the most promising[148,149]. Notably, Saccharomyces is the only recommended for pediatric use by the European Society for Paediatric Gastroenterology, Hepatology and Nutrition[150]. Even with the uncertain evidence, the prescription of probiotics has shown to be significantly beneficial, but more clinical studies investigating the potentiality of specific agents and strains are needed in order to formulate more specific guidelines for their use.

DISEASES ASSOCIATED WITH HELICOBACTER PYLORI INFECTION

H. pylori infection and chronic gastritis can evolve to different gastric and non-gastric conditions, associated with bacterial virulence, immunological responses and environmental factors. Figure 2 summarizes some of the proposed mechanisms involved in the development of gastric diseases associated to the infection, which will be further discussed.

Figure 2 Schematic summary of the underlying mechanisms to Helicobacter pylori-associated gastric diseases. IL-27: Interleukin 27; DupA: Duodenal ulcer promoting protein A; VacA: Vacuolating cytotoxin A; CagA: Cytotoxin-associated antigen A; MALT: Mucosa-associated lymphoid tissue.

Gastric and duodenal ulcers

H. pylori infection is the prevailing cause of gastric and duodenal peptic ulcers led by chronic gastritis[151], all of which fall under the larger category of peptic ulcer disease (PUD). While duodenal ulcers are clinically presented as a disruption to the mucosal surface of the duodenum that extends beyond the pre-epithelial, epithelial, and subepithelial layers[152], gastric ulcers consist of damage in the mucosal part of the stomach breaching the muscularis mucosa and exceeding 5 mm in diameter[153].

Knowing H. pylori has tropism to gastric type epithelium, the difference on the site of ulceration is deeply linked with the gastritis extent and distribution[6]. The duodenum is normally a hostile environment to H. pylori, but its induced inflammation and dysregulation of acid secretion is capable of lowering the pH in the duodenal bulb, precipitating the glycine conjugated bile acids and allowing the bacteria to infect mostly its metaplasic gastric mucosa[154]. Similarly, when the gastritis pattern predominates in the stomach’s antrum, H. pylori-induced inflammation disrupts the secretions of gastric acid-regulating hormones such as somatostatin, stimulating gastrin secretion[155] and leading to increased acid production directly via parietal cells and indirectly via histamine release from enterochromaffin-like cells[156] resulting in a hyperacidic gastric environment and a damaged tissue, prone to ulceration[157].

Several virulence factors may have a role in increasing the risk of peptic ulceration, such as the s1 and i1 types of VacA[24,158], and strains positive for the duodenal ulcer promoting gene A (dupA), due to more severe gastritis and increased expression of IL-8 in the mucosal lining. However, while some studies have linked dupA to a higher risk of developing duodenal ulcers and GC, others have failed to find consistent evidence supporting these associations[159-161]. Other determinants that contribute to the development of gastroduodenal ulcers include the use of non-steroidal anti-inflammatory drug (NSAIDs)[162], social and environmental factors such as older age[5,163], H. pylori infection within relatives[164] and even psychological stress, seeing it contributes to an increase in acid production[154,165-167].

Gastric and duodenal ulcers, within the spectrum of PUD, have distinct and non-specific symptoms. Thus, patients with gastric ulcers tend to have postprandial abdominal pain, nausea, vomiting, and weight loss due to aversion to food, a result of the intensification of pain after eating, while duodenal ulcers commonly present as nocturnal abdominal pain, hunger followed by bloating, and nausea[168]. The main complications of peptic ulcer disease are bleeding, presented as melena or hematemesis, and perforation, presented as a sudden onset of intense pain in the upper abdomen, and gastric outlet obstruction[168]. In untreated peptic ulcer disease, symptoms often recur due to a cycle of spontaneous healing followed by relapse, driven by the continued presence of underlying causes such as H. pylori infection or NSAID usage[168].

There is robust evidence through meta-analyses that eradicating H. pylori has positive outcomes in peptic ulcer disease of both stomach and duodenum, with or without complications[169-172]. More recent evidence suggests that in H. pylori-positive duodenal ulcers eradication therapy is superior to other medical treatments, but the same has not yet been proven for H. pylori-positive gastric ulcers[171]. Eradication of H. pylori is also related to lower recurrence of gastric and duodenal ulcers in patients who were not put on anti secretory therapy for management of the disease[169].

Gastric adenocarcinoma

As shown, H. pylori is a significant risk factor for gastric adenocarcinoma (GAC), driven by complex mechanisms involving both bacterial factors and host characteristics[173]. Based on epidemiological observations, Pelayo Correa proposed a model describing the cascade of events leading to intestinal-type gastric carcinogenesis: Chronic active gastritis and progresses to multifocal gastric atrophy, IM, low-grade dysplasia, high-grade dysplasia, and eventually gastric adenocarcinoma[174]. Although H. pylori infection is widely acknowledged as the most significant cause of GAC[175], metabolic, environmental, genomic and epigenetic factors all contribute to carcinogenesis[7], including family history, lifestyle factors (dietary habits, smoking, or alcohol consumption), and even Epstein-Barr virus infection[176], which explain why not all individuals infected with H. pylori develop GC.

Another determinant to whether or not a H. pylori-positive individual could evolve to GAC is an infection by more or less virulent strains. CagA-positive strains are highly correlated to GC, due to its capability of by altering host cellular signaling and creating a pro-oncogenic environment[177], and their infections result in more severe clinical outcomes and significantly elevate the risk of GC beyond that observed with H. pylori infection alone[178]. VacA is another key virulence factor of H. pylori-induced carcinogenesis, seeing it interferes with autophagy and allows bacteria to evade the host immune response, thereby contributing to chronic infection and GC development[179]. In this context, strains of H. pylori that are both CagA-positive and carry the VacA s1 and m1 genotypes are associated with a heightened risk of GC, although VacA s1m1 strains have been linked to both GC and peptic ulcer disease[180]. Nonetheless, other H. pylori virulence factors, such as HtrA[181], BabA[182], SabA[183], and OipA[184], all play critical roles in the pathogenesis of GC, by facilitating adhesion, colonization, enhancing inflammation and epithelial damage.

Gastric cancer is often asymptomatic in its early stages. When symptoms do appear, they typically include dysphagia, asthenia, indigestion, vomiting, weight loss, early satiety, and/or iron-deficiency anemia[185]. These highly unspecific symptoms frequently lead to delayed diagnosis, resulting in approximately 60% of GC patients being ineligible for curative treatment due to late-stage presentation or comorbidities[186].

Eradicating H. pylori is currently regarded as the most effective strategy for preventing gastric adenocarcinoma, including proximal GC[74], especially among those without premalignant gastric lesions at the start of treatment[187], but the limited impact on overall mortality highlights the need for further research into the broader benefits of eradication across varying clinical scenarios[187]. Even after successful eradication, progression to gastric adenocarcinoma may still occur, depending on factors such as the condition of the underlying gastric mucosa and the presence of other risk factors[74]. The viability of preventing rather than curing established GC through H. pylori eradication, particularly in cases involving CagA-positive strains, reiterates the "hit-and-run" mechanism of carcinogenesis[188].

Mucosa-associated lymphoid tissue lymphoma

A low-grade B-cell neoplasia, the gastric mucosa-associated lymphoid tissue lymphoma (GML) is evidently correlated with H. pylori infection[189]. Its pathogenesis relies on formation of mucosa-associated lymphoid tissue (MALT) through chronic inflammation, persistent antigenic stimulation and cytokine liberation[190,191]. T helper cells also play a crucial role in lymphoid tissue formation by supporting polyclonal B cells[192,193], which are capable of recognizing auto antigens from the gastric mucosa due to cross-reactivity with bacterial epitopes. As a result, polyclonal B cells increase in number in the region, heightening the risk of translocations and double-strand DNA mutations, culminating in the emergence of antigen-dependent marginal zone lymphomas[191,194,195].

In this context, the translocation t (11;18)(q21;q21) is observed in 10% to 50% of GML cases[196-198] and leads to the activation of NF-κB mediated by the disruption of a signalosome complex involving BCL10, CARD11, and MALT1. The latter, a fusion protein, is associated with advanced stages of MALT lymphoma[199-201]. Furthermore, chronic exposure to antigens induces the release of reactive oxygen species through immune responses, which contribute to the development of genetic alterations[202]. Consequently, during the evolution of lymphoid tissue, genetic and epigenetic changes may occur in genes linked to cancer development, such as tumor suppressor genes and oncogenes, ultimately leading to lymphoma. Among the most common alterations are the API2-MALT1 gene fusion and mutations in the TP53 and M4D88 genes, which are associated with lymphoma[191,203].

When evaluating the clinical features of GML, a wide variability of symptoms is associated with the condition, which may include indigestion, nausea, vomiting, abdominal pain and distension, as well as weight loss[204]. Its diagnosis is primarily achieved through analyses conducted via biopsy of the affected tissue[18-20]. Treatment, according to clinical guidelines, focuses on eradicating H. pylori, which results in lymphoma cure in over 75% of early-stage GML patients with H. pylori infection, and up to 29.3% of patients without H. pylori infection, who still achieve complete remission following bacterial eradication therapy despite the absence of an infection diagnosis[205-208].

These findings are conflicting, as it was anticipated that H. pylori eradication therapy would have no effect on remission in patients without the infection. This suggests the potential involvement of other bacteria in GML development, and that eradicating these bacteria may also lead to remission. Such outcomes have heightened researchers' interest in the correlation between non-H. pylori gastric helicobacters (NHPHs) and GML development[203]. These species differ from H. pylori both genetically and phenotypically, exhibiting urease positivity, diverse virulence factors, and varied flagellar structures[209,210]. However, in NHPH infections in mice, inflammatory processes associated with GML development were found to be remarkably similar to those seen in H. pylori infections, demonstrating significant similarities in the pathogenesis of these infections[211-215].

Extra-gastric manifestations

Besides the usually associated gastric diseases, H. pylori has shown correlation to extra-gastric conditions. As previously reviewed by Santos et al[216] the non-gastric manifestations found in literature can range from hematological and autoimmune to affecting different organ systems, although some still lack robust evidence to their correlations.

Iron deficiency anemia (IDA) and vitamin B12 deficiency are two of the most well-established extra-gastric manifestations of H. pylori infection, as chronic inflammation can lead to hypochlorhydria and reduced secretion of intrinsic factor, impairing iron and B12 absorption, respectively. Although these conditions are mostly correlated to cases of atrophic gastritis, they have also been observed in patients with milder inflammation[217]. Clinical trials and meta-analyses have confirmed that eradication therapy significantly improves serological parameters in H. pylori-positive individuals with IDA[218,219] and B12 deficiency[220], encouraging clinicians to test for H. pylori infection when presented to unexplained cases of these conditions.

Another condition correlated to H. pylori infection is immune thrombocytopenic purpura (ITP): An autoimmune disease associated with low platelet serum levels, allowing the occurrence of hemorrhagic lesions throughout the body. The mechanism of which H. pylori infections induces ITP is not completely elucidated yet, but the fact that H. pylori eradication in patients with ITP leads to satisfactory improvement in platelet serum levels strengthens the hypothesis that the bacterium influences the pathogenesis of this disease[221-223]. One of the theories regarding H. pylori influence is a molecular mimicry of CagA, that being, anti-CagA antibodies cross-reacting with platelet glycoprotein IIb/IIIa complex[224], opsonizing and facilitating phagocytosis by Kupffer cells and macrophages in the spleen, while also impairing megakaryopoiesis[225,226].

VACCINES IN DEVELOPMENT

Due to the high prevalence of H. pylori infection[227] and its association with the development of severe diseases[206,228,229], as well as other clinically significant pathologies, including extra-gastric conditions[216], the ineffective immune response in eradicating the bacterium in a substantial proportion of patients[230] and the frequent inefficacy of antibiotic therapy due to increasing bacterial resistance[231], there is an urgent need for the development of prophylactic and therapeutic vaccines against H. pylori.

Several bacterial antigens have been identified as promising targets for vaccine development, such as urease[12], CagA[232-235], and VacA[232,236], singled out as some of the most important virulence factors for H. pylori infection and colonization, as well as catalase, BabA, HspA, GGT, FliD, and others[12]. However, due to the challenges in achieving the desired protective effects, the development of a multi-epitope and recombinant vaccines is under consideration[237,238]. One study reported promising results, demonstrating immunoprotective effects and reduced inflammation associated with infection by using the cholera toxin B subunit combined with tandem copies of T- and B-cell epitopes from the UreA and UreB subunits of urease (CTB-UE), administered in a Mongolian gerbil model against H. pylori[237].

Another study involving 80 mice and employing the EpiVax/H. pylori vaccine-a multi-epitope approach combining DNA vaccination followed by a peptide-liposome formulation-showed efficacy and high interferon-γ production[239].

Helicovaxor utilized the recombinant expression of the HpaA adhesion antigen, either alone or in combination with colonization factor antigens from enterotoxigenic Escherichia coli, enhancing the immune response against HpaA[240]. Meanwhile, another vaccine, developed by the Murdoch Children's Research Institute and currently in preclinical stages, does not aim to eradicate the infection but rather to reduce inflammation. This vaccine employs HtrA, the sole serine protease produced by H. pylori, and has demonstrated inflammation inhibition in mice[12].

Some promising vaccines are administered orally, such as the probiotic vaccine that employs Lactococcus lactis containing the UreB-IL-2 protein. This vaccine elicits an immune response with the production of anti-urease antibodies and has demonstrated the ability to reduce H. pylori colonization, confirming UreB-IL-2 as a potent candidate for oral vaccines[241].

Several other published clinical studies have not achieved the anticipated success, as immunity was not effectively developed. Nevertheless, these studies remain of significant importance for guiding future research[12].

Currently, the website https://clinicaltrials.gov lists four completed studies on both prophylactic and therapeutic vaccines for H. pylori infections. Among these is a Phase I clinical trial by Imevax evaluating the safety of the therapeutic vaccine IMX101 in both infected patients and H. pylori-negative individuals. This vaccine incorporates the GGT antigen, an undisclosed H. pylori outer membrane protein, and a mucosal adjuvant. Another completed study, a Phase III trial conducted by the Jiangsu Province Centers for Disease Control and Prevention, investigates the safety of an oral recombinant H. pylori vaccine in Chinese children. The trials identified are summarized in Table 2.

Table 2 Helicobacter pylori vaccines currently in clinical trials - clinicalTrials.gov database.

Vaccine classification	Intervention/treatment	ClinicalTrials.gov ID	Phase	Status	Sponsor
Therapeutic vaccines	Biological: CTA control/biological: IMX101 vaccine	NCT03270800	1	Completed	ImevaX
Prophylactic vaccines	Biological: H. pylori vacines/biological: Placebo Vaccine	NCT00736476	1	Completed	Novartis Vaccines
	Biological: H. pylori vaccine/biological: Placebo	NCT02302170	3	Completed	Jiangsu Province Centers for Disease Control and Prevention
	Biological: H. pylori vaccine/biological: Placebo	NCT00613665	1	Completed	Novartis Vaccines

H. pylori: Helicobacter pylori.

CONCLUSION

Important advances have been made in understanding the complex interplay between H. pylori, its virulence mechanisms, host immune responses, and clinical outcomes, from the molecular mechanisms of virulence to the development of severe gastroduodenal diseases and extra-gastric associations. Despite progress in diagnostic methods and treatment regimens, the growing challenge of antibiotic resistance and limitations of empirical therapy underscore the importance of validating the use of susceptibility-guided treatment. Additionally, novel therapeutic strategies hold promise but require further validation before routine application. Understanding H. pylori pathogenesis in greater depth, alongside efforts to refine clinical management and preventive strategies, is essential to mitigate the burden of infection and its associated diseases worldwide.