Helicobacter pylori is a gram-negative, microaerophilic microorganism spiral in shape, which inhabits the gastric mucosa and is the primary causative agent of peptic ulcer disease (PUD). Ever since its discovery, H. pylori has been known as one of the most widely spread chronic bacterial infections in the global population with almost half of the world being infected by it [1]. It is significantly more common in developing nations where it is transmitted by socioeconomic factors, overcrowding, and sanitation conditions [2]. Reported prevalence rates are still dramatically high in Pakistan and other parts of the South Asia in contrast to the Western populations [3]. H. pylori infection is closely linked to the peptic ulcer disease which involves the mucosal defect in the duodenum or stomach. Virulence factors, including cytotoxin-associated gene A (CagA) and vacuolating cytotoxin A (VacA) cause the bacterium to cause chronic gastritis and mucosal inflammation, which results in epithelial damage and ulcer formation [4]. There is epidemiological evidence to indicate that about 70-90 percent of the duodenal and 50-70 percent of the gastric ulcers are associated with H. pylori infection [5]. Recurrent infection is a factor that not only causes ulcer recurrence but also predisposes gastric adenocarcinoma and mucosa-associated lymphoid tissue (MALT) lymphoma [6]. The standard treatment regimens of the H. pylori infection are usually combination therapy of proton pump inhibitor and two or more antibiotics, usually clarithromycin, amoxicillin, metronidazole, or levofloxacin [7]. These regimens have however lost their effectiveness in recent years following the increasing antimicrobial resistance [8]. The issue of clarithromycin resistance, especially, has become a significant determinant of treatment failure in most parts of the world [9]. There have also been worrying trends in the rates of metronidazole and levofloxacin resistance particularly in the low- and middle-income nations [10]. The concept of multidrug resistance (MDR), which is defined as resistance to two or more of the commonly used antibiotics provides a real challenge to the health of the population. MDR strains also decrease the success rates of eradication greatly and make therapeutic strategies difficult [11]. Resistance trends can be even stronger in areas where there is high antibiotic abuse and over-the-counter accessibility, as in Pakistan [12]. Improper prescriptive habits and unfinished treatment regimens are additional forms that increase the development of resistant strains [13]. The data on local surveillance of prevalence and pattern of antibiotic susceptibility trends will be critical in giving the guidance in empirical treatment. Even international guidelines are more in favor of regional-specific therapy development in lieu of universal first-line regimens [14]. Devoid of renewed local statistics, clinicians are more likely to prescribe unhelpful therapies, which will cause chronic infection and higher healthcare costs [15][16].

Objective

To determine the prevalence and multidrug resistance pattern of Helicobacter pylori among patients with peptic ulcer disease attending tertiary care hospitals in Pakistan.

This was a hospital-based cross-sectional analytical study conducted at Tertiary Care Hospitals across major cities of Pakistan March, 2023, to September, 2025, including 255 adult patients presenting with clinically suspected peptic ulcer disease.

Inclusion Criteria

• Patients aged ≥18 years with endoscopically confirmed gastric or duodenal ulcer.
• Patients undergoing diagnostic testing for pylori infection (biopsy-based and/or stool antigen test as per protocol).
• Patients who provided informed written consent.

Exclusion Criteria

• Patients who received pylori eradication therapy or antibiotics within the preceding 4 weeks.
• Patients with gastric malignancy or history of gastric surgery.
• Patients unwilling to participate or with incomplete diagnostic workup.

Data Collection

After obtaining written informed consent, demographic and clinical data including age, gender, smoking status, NSAID use, prior antibiotic exposure, and presenting symptoms were recorded using a structured proforma. Upper gastrointestinal endoscopy findings were documented, and gastric biopsies were obtained for rapid urease testing, histopathology, and culture where feasible. Antimicrobial susceptibility testing of H. pylori isolates was performed against commonly used antibiotics including clarithromycin, metronidazole, amoxicillin, levofloxacin, and tetracycline using standardized laboratory methods. Multidrug resistance was defined as resistance to two or more antibiotic classes used in standard eradication regimens. All data were entered into a secured database for analysis.

Statistical Analysis

Data were analyzed using SPSS version 26. Continuous variables were expressed as mean ± standard deviation (SD), and categorical variables were presented as frequency and percentage. The prevalence of H. pylori infection and antibiotic resistance rates were calculated as proportions. Associations between infection status and demographic or clinical variables were assessed using the chi-square test. A p-value of <0.05 was considered statistically significant.

A total of 255 patients with peptic ulcer disease were included, with a mean age of 41.6 ± 13.2 years. Nearly half of the participants (48.6%) were aged 31–50 years (40.7 ± 5.2 years), while 24.3% were 18–30 years (25.8 ± 3.4 years) and 27.1% were above 50 years (58.9 ± 6.8 years). Males constituted 59.6% of the cohort (mean age 42.3 ± 13.5 years), compared to 40.4% females (40.5 ± 12.8 years). Duodenal ulcers were more common (63.5%; mean age 39.8 ± 12.6 years) than gastric ulcers (36.5%; 44.9 ± 14.1 years). NSAID use was reported in 38.4% (46.2 ± 12.7 years), while 43.1% were smokers (44.1 ± 12.5 years), indicating substantial exposure to established ulcer risk factors.

Table 1. Baseline Demographic and Clinical Characteristics of Peptic Ulcer Disease Patients (N = 255)

The overall prevalence of Helicobacter pylori infection was 68.2% (174/255). Infected patients were significantly younger, with a mean age of 39.7 ± 12.8 years compared to 45.8 ± 13.4 years in non-infected patients (p = 0.002). Duodenal ulcer was strongly associated with infection (71.3% vs. 46.9%; p <0.001), whereas gastric ulcer was more common in non-infected patients (53.1% vs. 28.7%; p <0.001). NSAID use was significantly higher in the H. pylori-negative group (56.8% vs. 29.9%; p <0.001), suggesting a non-infectious etiology in those cases.

Table 2. Prevalence of Helicobacter pylori Infection Among PUD Patients (N = 255)

Overall prevalence of H. pylori infection: 174/255 (68.2%)

Among 174 culture-positive isolates, the highest resistance was observed to metronidazole (58.6%) with a mean MIC of 8.6 ± 3.2 µg/mL, followed by clarithromycin resistance at 44.8% (1.9 ± 0.8 µg/mL). Levofloxacin resistance was noted in 36.8% (2.3 ± 1.1 µg/mL), while resistance to amoxicillin and tetracycline remained comparatively low at 16.1% (0.9 ± 0.4 µg/mL) and 10.9% (0.6 ± 0.3 µg/mL), respectively.

Table 3. Antibiotic Resistance Pattern Among H. Pylori Positive Isolates (n = 174)

Multidrug resistance (resistance to ≥2 antibiotics) was identified in 36.8% of isolates, with a mean patient age of 42.7 ± 13.1 years (p = 0.012). Single-drug resistance was observed in 29.9% (40.2 ± 12.4 years), while 33.3% showed no resistance (37.4 ± 11.8 years). Dual resistance to clarithromycin and metronidazole was present in 27.6% (43.1 ± 12.9 years), and triple-drug resistance was detected in 12.1% (44.8 ± 13.6 years).

Table 4. Multidrug Resistance Pattern Among H. Pylori Isolates (n = 174)

This research showed that the prevalence of the disease in peptic ulcer patients was high with a prevalence rate of 68.2% of the patients having the infection, which was in line with the established burden of infection in the developing nations. Infected patients were also much younger (39.7 ± 12.8 years) than non-infected persons (45.8 ± 13.4 years), which indicated that the disease caused by H. pylori can develop earlier in life. There have been reports in the past of prevalence rates of between 60-75 in similar socioeconomic settings especially in patients, who have duodenal ulcers [17]. Duodenal ulcers were strongly associated with H. pylori positivity (71.3%), whilst gastric ulcers were more prevalent among H. pylori-negative patients, most of whom reported increased NSAID use (56.8%). This supports the evidence provided by other studies that showed that H. pylori has a stronger etiological relationship with duodenal ulcer disease and injury by NSAID is more widely associated with gastric ulcers [18]. The pattern in antibiotic resistance that is observed in the given study is alarming. The resistance rates to metronidazole (58.6%), clarithromycin (44.8%), and levofloxacin (36.8%), were the highest. The resistance to amoxicillin (16.1) and tetracycline (10.9) was still low. Such high rates of resistance to metronidazole and clarithromycin are not new in South Asia and such high resistance rates have been recorded in previous studies which were usually linked to the high rates of antibiotic misuse and their availability as over-the-counter drugs. The rate of clarithromycin resistance in this case goes beyond the 15 percent mark that the international guidelines recommend, and this is a cause of serious concern about the effectiveness of the common first-line therapy [19]. It is noteworthy that 36.8% of isolates were multidrug resistant and 27.6 and 12.1 were dual and triple resistant to clarithromycin and metronidazole respectively. Past studies have indicated that multidrug resistance rates are on the rise all over the world, especially in areas where people are exposed to antibiotics. The fact that older age is associated with resistance patterns in this study suggests that cumulative exposure to antibiotics may be among the causal factors noted in previous studies [20]. All in all, the results indicate a high prevalence of H. pylori infection and a concerning multidrug resistance pattern among patients with peptic ulcers. In line with the prior studies, the increased resistance rates jeopardize the efficacy of the empirical therapy and emphasize the necessity of region-specific surveillance and susceptibility-based treatment plans to enhance the eradication rates.

It is concluded that Helicobacter pylori infection remains highly prevalent among patients with peptic ulcer disease, with more than two-thirds of cases testing positive. The study reveals alarmingly high resistance rates to commonly used first-line antibiotics, particularly metronidazole and clarithromycin, as well as a substantial burden of multidrug-resistant strains. These findings indicate that standard empirical triple therapy may no longer be reliably effective in this setting. Continuous regional surveillance and implementation of susceptibility-guided treatment strategies are essential to improve eradication success and prevent further escalation of antimicrobial resistance.

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