Strategies In Overcoming Drug Resistances in H. Pylori Infection

Peptic ulcer disease (PUD) is a worldwide issue with a 5- to 10% lifetime risk of developing. The incidence rate of PUD gets reduced globally (around 51%) as a result of improved sanitary and hygienic practices as well as the adoption of effective therapeutic methods and the judicious use of non- steroidal anti- inflammatory medication (NSAIDs). Consequently, the prevalence, hospital admission, and death rate associated with peptic ulcers have decreased, according to epidemiological studies; however, the case death rate has stabilized at 5-10%[1] The gastrointestinal tracts (GI) mucosal lining gets damaged or ruptured in PUD which causes excessive amount of gastric acid or the pepsin to be secreted. theses symptoms usually occur in the close section of the duodenum and the stomach area [1]. PUD can be caused by many different factors. In those suffering from peptic disease, mucosal disruption may be to a high level of acid secreted in addition to nutrients or anxiety. the use of nonsteroidal anti- inflammatory drugs (NSAIDs) and an infection with H. pylori are the primary risk factors for gastric or peptic ulcers [1] Approximately half of the population of this country suffers with helicobacter pylori (H. pylori) which is the roots of the most common and determined pathogenic bacteria in the world. It is thought that the H. pylori cause stomach cancer, gastritis, peptic ulcer disease and functional dyspepsia [1-3] The first successful isolation of h. pylori to the intestinal biospsies of patient aith choronic antral gastric ulcers occurred in 1983(Isaacson and Wright, 1983). H. pylori is a spiral shaped 2-4 m long, 0.5-1 m wide microaerophilic, gram-negative bacteria [19]. H. pylori employ a variety of methods to infect the gastric mucosa and suffer in the strong gastric environment. these approaches include 1] leaving the acidic stomach lumen 2] adhering to the stomach epithelial cells 3] buffering the pH in the vicinity 4] developing biofilm 5] changing into a stress- resistance morphology (coccoid) which is possible but impossible to cultivate state and 6] bypassing the immune system of the host [20,21]. Additionally, the  development of carcinogenesis is linked to H. pylori toxins like vacuolating cytotoxin (VaCA) and the cytotoxin – associated gene A (CagA). the most prevalence characteristics of H. pylori linked to gastric infection

Routes of H.pylori infection

It is essential for gaining a deeper understanding of the pathways of transmission of infection with H.pylori with the goal for enhanced the protection of people against infection . there are three methods that H.pylori may infect people.

1 Individual to individual routes

2 Routes from animals to humans

3 Routes for food and water

1. Individual to individual routes

Person to person contact may outcome in the spread of H.pylori , particularly in families where mother -child transmission is common (Yang et al., 2023) .Research has demonstrated that H. pylori infections can happen in families in cluster.

2.  Routes from animal to humans

Human-to-animal transmission is considered to be a significant pathway for H.pylori infection. An organism called H.pylori can infect both people and animals. (Duan et al., 2023). Additionally evidence suggesting that milk and sheep milk may act as a vector for H.pylori infection in humans is provided by detection of the bacteria in milk, meat (beef, mutton) and other fresh foods (Hemmatinezhad et al., 2016; Shaaban et al., 2023)

3 .  Routes for food and water

Finally research indicates the food and water based infection H.pylori  infection .H. pylori feces can contaminate lake, river and ground water , all of which are crucial origins of drinking water  therefore its probable that absorbing water is how people obtained H.pylori

Pathogenesis of Helicobacter pylori

Three distinct stages of the h. pylori pathogenesis can be studied the attachment with and colonization in the gastrointestinal tract, the induction and envasion of the hosts immune response, and the eventual establishment of the disease.[19]

1 Dispersal and route of infection

H.pylori infections are typically acquired during childhood and frequently asymptomatic .however , about 30% of infected individual may exhibit symptoms of digestive disorders, including moderate- to- sever gastritis, peptic ulcer, and even stomach cancer. MALT lymphoma because H. pylori infections occur within a limited host range, horizontal as well as vertical transmission are common mode of transmission. The four main routes of H.pylori infection are the fecal-oral, gastric oral, oral-oral, and gastric routes, which can happen through ingesting of contaminated food, water, or during endoscopic procedures, even through the precise way of transmission remain unidentified.[20]

2. molecular mechanisms of infection

2.2.1 Attachment and colonization

The cellular process coordinating H.pylori infection has been advanced method essential for the bacteria beneficial the development in the stomach mucosa ,at the top during disease. Starting in h. pylori attachment and colonization strategies. The bacterial cells chemotaxis to the target site, that is mediated be a specific receptor available on the host cells- mostly members of the Tlp family [22] – is what is initially initiates the pathogens colonization. these receptors have been shown to be activated by an abundance of the chemical signals, such as urea, lactic acid, ROS species, and gastric juice, which facilitates chemical reaction epithelium[22]. Flagellar motility enables H.pylori to get around via the mucus the layer as well as come to stomach epithelium , contributing towards its capacity to strongly hold to while expand the mucosa [23]. Additionally, H.pylori can form biofilms, structured bacterial communities embedded in an exterior matrix, developing dedication and delivering defence towards the host defence’s and treatment therapies [24].

2.2.2 Production of virulence factor

This bacterium’s the causes depends upon a number of virulence factors, such as those that contribute in motility, adhesion, urease, and the production of cytotoxins [26].

2.2.3. CagA and VacA

Enhances patient compliance by lowering the frequency of dosing their smaller size and spherical shape allow them to be added into the body. after attaching itself, h. pylori interact with the host in a complex way by using a variety of virulence factors to control host cell signalling. the primary coordinator of these is the pathogenicity island known as CagA (cytotoxin associated gene A) [27], if CagA is injected into host cells through the type 4 secretion system (T4SS), it sets if a series of events that impair cellular processes and aid in the emergence of gastric pathologies [28]. Numerous pathways of cell signalling are activated as a result of the tyrosine phosphorylation of CagA [29]. This involves the phosphatidylinositol 3-Kinase (PI3K) mitogen activated protein kinase (MAPK) pathways being abnormally activated [30,31]. These pathways are dysregulated, which leads to alterations, in cellular morphology, loss of cell polarity, and the start of carcinogenic process. A defining characteristic of the pathogenesis caused by H. pylori is this cellular taking control of host cell signalling [32]. Another important virulence factor that has different impact on the host cells in the vacuolating toxin VacA[32]. As its name implies VacA forms and compromises the functioning of the gastric epithelial barrier.  Channels or pores that cause the cell membrane to become more permeable [26]. In the VacA alters the apoptotic pathways in host cells, resulting in pro – and antiapoptotic impact that vary based on the type of cell and related circumstances. Additionally, VacA toxin influences the structure and function of the host cells by aiding in the establishment of vacuoles with them [34]. Additionally, this toxin thwarts and subverts immune Reponses by influencing T cell and other unaffected cell function. 

Urease production and survival at low PH

Approximately 15% of all the proteins in the bacterium are made up of urease, one of the most frequently expressed proteins by the pathogen [42]. One of H. pylori distinguishing characteristics is the urease production, which is frequently utilized in the identification of additionally the infection [43]. Additionally, H. pylori demonstrate chemotactic activity towards specific chemoattractant, like urea and carbonates, which draw the bacterial cells to the areas of the stomach lining that have a greater pH. The family of H. pylori chemical receptors play a critical role in the desired chemotactic activity that encourages survival in an acidic stress environment [50].

Immune system modulation and induction of inflammatory responses

The hosts immune system and bacterial factors interact dynamically to produce an immune reaction to an infection with H.pylori [51] . The development of H. pylori infections is caused by an immune system response that is complicated and multifaceted [52]. H. pylori cause chronic inflammation in the stomach mucosa during the innate immune phase. This response is mediated by lipids and the peptidoglycan component of the cell wall and is characterized by the recruitment of immune cells, neutrophils, macrophages, lymphocytes, and pro- inflammatory cytokines (like interleukin -1beta, interleukin- 6, and tumour – necrosis factor -alpha) as well as the release of neutrophils, macrophages and lymphocytes [53,54]. Via pattern recognition receptors like toll-like receptors, innate immune activation triggered by bacterial components such as lipopolysaccharides and peptidoglycan intensify the inflammatory cascade [55]. This prepares the body for the immune systems adaptive response, in which CD4+T-helper cells are essential for fostering a Th1 response that secretes interferon -gamma. while B Cells create antibodies against H. pylori, regulatory cells are enlisted and help suppress the immune system [56,57].

Modulation of mucin production

The protective mucous layer in the stomach mucosa is impacted by H. pylori influence on mucin production. The mucins carbohydrate components function as a ligand to facilitate the bacterium’s attachment to the stomach mucosal lining [63]. The microorganism can change the production of the mucin genes and directly affected mucins such as MUC5A and MUC1. Causing mucin turnover to be blocked or hampered [64]. The modifications to mucin production have an impact on the mucosal glycocalyx’s composition, which influences adherence. And H.pylori colonization Additionally , this regulation of mucin synthesis helps to the capacity of the bacteria to overcome the hosts defences and causes enduring infections [65]. In the continuous efforts to manage the related diseases, understanding the subtleties of immune system control in infection by H.pylori is essential for developing treatments and preventive measures . The intricacies of this host – pathogen interaction are still being discovered by the ongoings research.

How it develops resistance in resistance H. pylori  

There are multiples ways that the organism helicobacter pylori have become resistance to antibiotics, but mutation in genes and horizontal transfer of genes are the main culprit, essentially H. pylori develop antibiotic resistance in the following ways

1. Chromosomal mutation

An overwhelming amount of research suggest that chromosomal changes [1]. have been the primary cause for H. pylori immunity to antibiotics. this mutation happens in particular genes linked to obstruction to various classes of antibiotics.

• Macrolides; 23S rRNA gene mutations [1] [2]
• Metronidazole rdxA and frxA gene mutations [1][2]
• Quinolones; gyrA gene mutations [1] [2]
• Rifamycin; rpoB gene mutations [1]
• Amoxicillin; pbp -1A gene mutations [1][2]
• Tetracycline; 16S rRNA gene mutation [1]

In H. pylori, small changes in those genes are a highly prevalence mechanism of resistance [1][2]. Resistance is conferred by the mutations lower capacity to bind antibiotics to their targets.  

2. Horizontal gene mutation

Transfer of antibiotics resistance may arise about genetic provides among district strains of H. pylori [1], through these are less frequent, studies conducted in vitro have demonstrated resistance genes can spread through transformation. [1]

3. Efflux pumps

Multidrug resistance (MDR) may occur compare to H. pylori efflux pump system triggering because they force antimicrobial agents out of the bacterial cells [5]. Gene mutation and efflux pump combine to increase resistance. 

4. Biofilm formation 

Because it can create biofilm, H. pylori is shielded from its hosts immune system and antibiotics [4], In addition to acting as a physical barrier, bio films cause bacteria to go dormant, which reduces their susceptibility to antibiotics. Chromosome mutation, horizontal transfer of genes, pumps for efflux, and biofilm creation have all performed together to give H, pylori resistance to the vast majority of first- line antibiotics used to eradicate the infection.[3][4]. To counter this growing g threat ongoing surveillance along with studies into novel treatment approaches are essential.

Resistance strains

Clarithromycin resistance

Globally clarithromycin resistance is a major concern, According to recent studies , resistance rates have gone up in a number of nations, including Australia ,Talwan , Belgium , Bulgaria , Italy , and Iran, for example, a study found that rates of antibiotics resistance ranged from 25% to 36% across different parts of Europe , and that resistance has been steadily rising over time, conversely[1][3] , stable resistance rates under 15% have been demonstrated in nations like France , Spain, and Russian .[1]. The bacterium Helicobacter pylori (H, pylori), which is well known for its involvement in gastrointestinal disorders, is the main cause of clarithromycin Aresistance. usually, the 23S ribosomal RNA gene has the particular mutations confer this resistance, specially at positions A2142G, A2142C, and A2143G theses mutations significantly decrease the binding efficacy of clarithromycin, which results in treatment failures.[1]

Specific bacteria and species

Bacteria name; Helicobacter pylori

Species; the primary species exhibiting clarithromycin resistance is helicobacter pylori

Common mutation associated with clarithromycin Resistance

1. A2142G mutation This mutation causes the 23S RNA genes position 2142to change from adenine (A) to guanine (G). In H. pylori strains resistant to clarithromycin, it is one of the most commonly found mutations and is linked to a high level of resistance (minimum inhibitory concentration, MIC >64mg/L).[1][2]
2. A2143G mutation This mutation, like A2142G, denotes a change in adenine (A) to guanine (G) at position 2143. It may confer a slightly lower level of resistance than A2142G, but it also frequently observed in resistance strains. [1][2]
3. A2142C mutation at position 214C, there is a mutation that causes adenine (A) to change to cytosine (C). Although it is not as A2142G and A2143G, it nevertheless plays a significant part in the development of clarithromycin resistance. [1][3]
4. Other notable mutations There are currently recognized mutation include;

• T2182C
• C2195T
• A2115G
• G2141A
• A2144T

Despite being less frequent, theses mutations might nevertheless be involved in resistance mechanism.[1][2][3]

Mechanism of resistance

The primary mechanism that these mutations confer resistance is via modification to the ribosomes RNAs clarithromycin binding site, The 23S rRNA peptidyl transferase region binds to clarithromycin, preventing the synthesis of new protein change in these areas cause the antibiotic binding affinity to be obstructed, which permitted H, pylori to persist in spite of these presence. [2][4]

Metronidazole Resistance  

Different regions demonstrate varying trends in metronidazole resistance. According to specific research, resistance levels in some populations- most especially those in Latin American and some parts of Asia – have reached 93.2%.[2] [3]. On the conjunction, resistance rates have decreased in other areas, emphasizing the need for specific monitoring to inform treatment choices.[3] one of the problems with treating infections brought on by different bacterial species is metronidazole resistance. An extensive summary of particular bacteria linked to metronidazole resistance along with an explanation of the resistance mechanisms, is provided below.

Specific bacteria associated with metronidazole resistance

1. Helicobacter pylori

• Mechanism of resistance
• Mutation in rdxA and fdxA Genes; The main cause of resistance in H, pylori is through to be mutations in the rdxA gene, that produce a nitro reductase necessary for metronidazole activation. Drug activation is reduced by mutation D59N and D59S, that can result in high resistance rates (up to 99%) [5][4]. The frxA gene may also modify the bacterial response to oxidation stress, which could further enhance resistance [4][5].

Prevalence of resistance; A study carried out in Bangladesh found that 72.7% of the clinical isolates of H, pylori [4] were resistance of metronidazole, according to another study, about 40% of H, pylori strains were resistant to metronidazole, which made treatment plans more difficult and required the use of combination regimens or other antibiotics [5].

2. Clostridioides difficile

Mechanism of resistance

Plasmid- mediated resistance; C. difficile’s metronidazole resistance has been linked to the pCD-METRO plasmid, which confers resistance by means of genes that facilitate drug inactivation.[1] The fact that this plasmid has been found in both toxic and non-toxic strains emphasizes its significance in the emergence of resistance to therapy.

Clinical cases; A patient’s development of metronidazole- resistant strains during treatment was documented in a case study. Underscoring the resistances therapeutic consequences.[1]

3. Bacteroides fragilis

Mechanism of resistance

Nim genes; The primary cause of Bacteroides fragilis metronidazole resistance among anaerobic bacteria is the presence of nim genes (e.g, nimA to nimK). The nitroimidazole reductases encoded by these genes have a capacity to inactivate metronidazole, resulting in various levels of resistance [2]. concerns concerning the effectiveness for treatment for infections brought on by this organism are raised by the rising frequency of resistance strains.

4. Enterococcus casseliflavus and Enterococcus gallinarum

Mechanism of resistance

Because these species produce nitroeductase, which convert metronidazole into inactive compounds, they show metronidazole resistance. The drugs antibacterial properties suffer by this metabolic alteration.[2] This resistance mechanism has been demonstrated in isolates from patients receiving repeated metronidazole treatments, suggesting a clinical connection between exposure to antibiotics and the appearance of resistance strains.

5. Giardia intestinalis; Mechanism in protozoan parasites such as Giardia intestinalis include decreased metronidazole uptake and raised efflux. Additionally connected have been modification to oxidoreductase- related metabolic pathways. [3][4] the need for alternate treatments is highlighted by recent studies showing an increase in G. intestinalis strains resistant to metronidazole.

Levofloxacin resistance

Levofloxacin resistance has also been observed to be rising, especially in Asia and Europe, where resistance rates have been found to be as high as 20% in certain studies,[1][2]. given the dependence on fluoroquinolones as second – line treatments for the eradication of H. pylori, this increase is cause for concern over levofloxacin resistance in the management of different bacterial infections is growing. An extensive summary of particular bacteria linked to levofloxacin resistance is provided below, along with the data on their mechanism, frequency.

Specific bacteria associate with levofloxacin resistance

1. Helicobacter pylori

Mechanism of resistance

Mutation in gyrA and gyrB genes; The main cause of resistance in H. Pylori is believed to be mutations in the gyrA gene, which produces a subunit of DNA gyrase, a Fluroquinone target. Specific mutations that have been strongly linked to levofloxacin resistance include N87K and D91N. [2] Additionally, there has been a strong correlation demonstrated between resistance and mutation in the gyrB gene, specifically at locus A343V. [2]

Prevalence of resistance; levofloxacin resistance rates among H. pylori isolates were found to be roughly 36% in some regions, according to a study [2]. Another study from Bangladesh found a significant correlation between levofloxacin resistance and mutations in gyrA underscoring the importance of susceptibility testing prior to starting treatment [2].

2. Escherichia coli

Mechanism of resistance;

 levofloxacin resistance in E. coli is frequently linked to mutation in the gyrA and parC genes. In particular, gyrA mutation at codon 83 and 87 are frequent [5]. Reduced susceptibility may also result from the existence of plasmid- medicated quinolone resistance (PMQR) genes, such as qnrS, which shield target enzyme from fluoroquninolone action.[5]

Prevalence of resistance; Depending on the region and local antibiotic usage patterns, studies have shown that clinical strains of E. coli have varied rates of levofloxacin resistance, ranging from 10 % to over 50%.[5]

3. Klebsiella pneumoniae

Mechanism of resistance; Klebsiella pneumoniae, like E, coli, primarily demonstrates levofloxacin resistance due to mutations in the gyrA in the gene. It has been determined that ser83Leu and Asp87Gly mutations are important causes of resistance [1]. Treatment options may become more complex when extended- spectrum beta-lactamases (ESBLs) are present.

Prevalence of resistance; levofloxacin resistance rates in pneumonia have been reported to exceed 40% especially in infections linked to healthcare facilities.[1]

4. Pseudomonas aeruginosa

Mechanism of resistance; Levofloxacin resistance in pseudomonas aeruginosa is frequently caused by mutations in the gyrA, gyrB, and parC genes. These genes can be mutated to change DNA gyrase and topoisomerase 4, two important targets of fluoroquinolones [6]. Moreover, efflux pumps are important in lowering drug concentration inside cells.

Prevalence of resistance; it has been estimated that 30% of patients have pseudomonas aeruginosa strains that are resistance to levofloxacin; patients who have chronic infection tend to have higher prevalence rates.[6]

5. Enterobacteriaceae

Mechanism of resistance; levofloxacin resistance in Enterobacteriaceae is associated with particular mutation in the gyrA genes quinolone resistance -determining regions (QRDR). for example, mutations like Asp87Asn and Ser83Tyr have been linked to higher resistance levels [1] by shielding target enzymes, PMQR genes like qnr can also promote higher-level resistance.

Prevalence of resistance; According to a study, the percentage of clinical isolates of Enterobacteriaceae that were resist to levofloxacin varied greatly, with some regions reporting as high as 50%.[1] 

Multidrug resistance

Concern over the rise in resistance to drugs H. pylori strains is growing, Numerous studies have stated cases of triple resistance to Levofloxacin, metronidazole, and clarithromycin as well as dual resistance to these three drugs, certain populations rates of triple resistance exceed 15% and dual resistance has reached 15% [1][2][3]. Due to their resistance to three or more classes of antimicrobial medication, multidrug- resistant (MDR) bacteria pose a serious threat public health. This summary offers comprehensive details on particular bacterial species linked to multidrug resistance, including their mechanisms, prevalence.

Specific bacteria and associated with multidrug resistance

1. Acinetobacter baumannii

Mechanism of resistance; A baumannii produces beta- lactamases, especially carbapenemases, which hydrolyse carbapenems and other beta- lactam, as one of its many antibiotic resistance strategies. Additionally, it has changes in outer permeability of the membrane that prevent drug uptake and uses efflux pumps to expel several classes of antibiotics.[1][2]

Prevalence of resistance; studies suggests that A. baumannii demonstrates elevated levels of resistance to multiple drugs. In fact, more than 70% of clinical isolates have been found to be resistant to multiple antibiotics, such as carbapenems and fluroquinolones.[3][4]

2. Escherichia coli

Mechanism of resistance; Resistance in Escherichia coli is frequently linked with the synthesis of plasmid- mediated quinolones resistance (PMQR) genes such as qnrS and extended-spectrum beta- lactamases (ESBLs). Fluoroquinolones resistance can be caused by mutations in the gyrA and parC genes.[5][6]

Prevalence of resistance; multidrug- resistant E. coli strains are growing more common, according to systematic review, with resistance rates for commonly used antibiotics reaching 50% in some areas.[7][8]

Strategies to overcome resistance H. pylori

Treatment strategies

1. Quadruple therapy

Bismuth based Regimens

These regimens are advised as first -line treatments, particularly in areas with high rates of metronidazole and clarithromycin as resistance. They consist of a bismuth as compound, a proton pump inhibitor (PPI), two types of antibiotics (typically metronidazole and tetracycline or amoxicillin), They demonstrated high removal rates (over 90%) if properly administered.[1][2][4].

AST, antibacterial susceptibility test; BQT, bismuth quadruple therapy; CoCTx, concomitant therapy

Figure 1. Simplified H. pylori treatment strategy.