Helicobacter pylori eradication in renal transplant candidates

Abstract Introduction: Treatment for Helicobacter pylori (H. pylori) infection is recommended in transplant candidates due to the association between this infection and gastrointestinal disorders, which could significantly increase morbidity after renal transplantation with the use of immunosuppression. The objective of this study was to analyze the rate of eradication of H. pylori after antimicrobial treatment in chronic kidney disease patients who are candidates for kidney transplantation. Methods: A multicenter prospective cohort study was conducted. All adult chronic kidney disease patients seen at our institution were included. In the pre-transplantation evaluation, 83 patients underwent an upper gastrointestinal endoscopy with 2 diagnostic methods to detect H. pylori: histology and the rapid urease test. In total, 33 patients with H. pylori infection received treatment with 20 mg omeprazole, 500 mg amoxicillin, and 500 mg clarithromycin once daily for 14 days. Another upper gastrointestinal endoscopy was performed 8 to 12 weeks after the end of treatment to check for healing. Results: The study showed a prevalence of H. pylori in 51 (61.4%) patients. Histology was positive in 50 (98%) patients and the rapid urease test was positive in 31 (60.8%). The infection eradication rate was 48.5% (16 patients). Conclusions: There was a high prevalence rate of H. pylori and a low eradication rate after the long-term antimicrobial triple scheme used. The association of the rapid urease test with gastric mucosa histology did not increase the detection rate of H. pylori.

Introduction: Treatment for Helicobacter pylori (H. pylori) infection is recommended in transplant candidates due to the association between this infection and gastrointestinal disorders, which could significantly increase morbidity after renal transplantation with the use of immunosuppression. The objective of this study was to analyze the rate of eradication of H. pylori after antimicrobial treatment in chronic kidney disease patients who are candidates for kidney transplantation. Methods: A multicenter prospective cohort study was conducted. All adult chronic kidney disease patients seen at our institution were included. In the pretransplantation evaluation, 83 patients underwent an upper gastrointestinal endoscopy with 2 diagnostic methods to detect H. pylori: histology and the rapid urease test. In total, 33 patients with H. pylori infection received treatment with 20 mg omeprazole, 500 mg amoxicillin, and 500 mg clarithromycin once daily for 14 days. Another upper gastrointestinal endoscopy was performed 8 to 12 weeks after the end of treatment to check for healing. Results: The study showed a prevalence of H. pylori in 51 (61.4%) patients. Histology was positive in 50 (98%) patients and the rapid urease test was positive in 31 (60.8%). The infection eradication rate was 48.5% (16 patients). Conclusions: There was a high prevalence rate of H. pylori and a low eradication rate after the long-term antimicrobial triple scheme used. The association of the rapid urease test with gastric mucosa histology did not increase the detection rate of H. pylori.

IntRoductIon
Helicobacter pylori (H. pylori) is a human pathogen with worldwide distribution 1 , responsible for the most prevalent bacterial infection currently known 2 . It is an etiologic agent of gastrointestinal tract comorbidities varying from mild to severe 3,4 . The prevalence of H. pylori ranges from 30 to 80% in several countries. Despite the high prevalence, clinical manifestations are rare in most patients who carry H. pylori in their gastrointestinal tracts, and only a minority of patients develop symptoms 5 .
The study of Homse and colaborators 6 showed that 100% of these patients present some form of high endoscopic alteration, many of which are potentially severe, including peptic ulcers, gastritis, erosive duodenitis, and gastric intestinal metaplasia 7 . H. pylori causes chronic gastric inflammation, which can progress into precancerous alterations such as atrophic gastritis and intestinal metaplasia 8 .
There are currently several methods to diagnose H. pylori. Some methods require a prior upper gastrointestinal endoscopy for access to the necessary material 2 . A biopsy of the gastric mucosa is required for some diagnostic methods such as: histology, culture, polymerase chain reaction, and the rapid urease test 2 . Histology is considered the gold standard method for diagnosis of infection by H. pylori. It also provides relevant information for the detection of numerous diseases of the esophagus and gastric mucosa 2 . The rapid urease test uses the ability of H. pylori to synthesize large amounts of urea as the basis for diagnosis and presents advantages such as low cost, availability, and high specificity, making it widely used in clinical practice.
Treatment for H. Pylori infection has been recommended in transplant candidates because of the association between this infection and gastrointestinal disorders such as peptic ulcers, gastric hyperplastic polyps, gastric adenomas, gastric cancers, and mucosa associated-lymphoid-type (MALT) lymphomas 9,10,11 . With the use of immunosuppression, these disorders could become serious, significantly increasing morbidity after renal transplant. The American College of Gastroenterology Guideline 12 recommends that clarithromycin triple therapy and bismuth quadruple therapy for H. pylori be administered for 14 days, similar to the current recommendations for prolonged treatments (10 to 14 days) 13,14 . Patients with the following characteristics were excluded: pregnant, those with a recent history (less than three months) of H. pylori infection, abdominal surgery, or high digestive hemorrhage, use of any antibiotic in the past 30 days, and history of allergies to any of the compounds of the therapeutic plan (omeprazole, amoxicillin, or clarithromycin) to be used in the treatment of H. pylori infection.
On the day of the procedure, the patients were interviewed regarding medications of continuous use and gastrointestinal tract symptoms. Upper gastrointestinal endoscopies were performed in the ambulatory service by a member of the Gastroenterology/Endoscopy service of the Evangelical Hospital of Londrina. Patients on continuous use of gastric protectors were advised to suspend the medication for 7 days before the endoscopy. Patients with H. pylori infections were treated daily with 20 mg omeprazole in the morning, in a fasting state, and 500 mg amoxicillin plus 500 mg clarithromycin at night, for 14 days. On hemodialysis days, patients were instructed to take the antibiotic after the sessions.
Data on general characteristics of the studied population were obtained: age, sex, ethnicity, origin, place of residence, and hemodialysis clinic. The following clinical data were collected through an interview: underlying disease, duration of hemodialysis prior to upper gastrointestinal endoscopy, associated comorbidities, previous and/or current smoking history, use of anti-hypertensives, use of gastric protectors, and presence of gastrointestinal tract symptoms in the three months prior to endoscopy. The data sources were registered in the patients' charts in the 6 dialysis units.
The first upper gastrointestinal endoscopy was performed at the time of pre-renal transplantation assessment at the clinic. The rapid urease test and histology, performed through gastric biopsy, were used to confirm H. pylori infection. A positive finding in one of these tests was indicative of H. pylori infection.
The biopsy was performed in two gastric regions: antrum and body. A third region was assessed in case of inflammation, a suggestive aspect of intestinal metaplasia or neoplasia. The fragments were fixed in 10% formalin and processed for histology and staining with Giemsa. Evaluations were performed by a single pathology laboratory.
The rapid urease test consisted of immersion of a gastric mucosa fragment of the antrum region into a vial containing urea and phenol red, an indicator of pH (H. pylori produces a urease enzyme that turns urea into carbon dioxide and ammonia, leading to an elevation in pH and color alteration to a shade of pink). The test was considered positive when the alteration occurred within two to sixty minutes.
Patients positive for H. pylori infection by at least one of the above methods received treatment according to the American College of Gastroenterology Guideline 12 . After one week of treatment, the patients were contacted by telephone to verify adherence to the treatment plan. At this point, the importance of completing treatment was reinforced. After completion of treatment, full compliance was confirmed when patients returned to see a nephrologist.

statistical analysis
The data were analyzed with the Windows' Medcalc program, version 18.0 (Medcalc Software, Ostend, Belgium).
The data are descriptively presented using simple frequencies (relative and absolute), means and deviation rates, or medians and interquartile range (IQR) depending on variable distribution. The data distribution was tested with the Shapiro-Wilk test. The frequencies were described as raw number or percentage, represented in contingency tables, and compared with the Fisher's exact test.

Results
In total, 83 patients with a median age of 47 years (IQR: 38 -56) were analyzed. The main underlying diseases were systemic arterial hypertension and diabetes mellitus, while the main dialysis type was hemodialysis. The median duration of dialysis before the upper gastrointestinal endoscopy was 14 months (IQR: 6 -48). Findings were positive in 96.4% of endoscopies. The most commonly found lesion was enanthematous pangastritis. Few patients had ulcers. Only one patient presented a precancerous lesion, which was Barrett's esophagus. No malignant lesions were detected. There were no associations between the endoscopic findings, symptomatology, and the presence of H. pylori based on the Fisher's exact test (Table 2).
Gastrointestinal symptoms were reported by 61.4% of patients. Epigastric pain did not occur in patients with endoscopic findings of ulcer. Nausea and pyrosis were frequent symptoms. There was a tendency for vomiting to be more associated to enantematous pangastritis (Table 3).
After 7 days of follow up by telephone contact, all patients presented good treatment compliance. Complete compliance was confirmed when patients returned to a nephrologist after 14 days of treatment; treatment did not include supervised pill-taking.

dIscussIon
A high prevalence of H. pylori was found in renal transplant candidates. The association of diagnostic methods was disadvantageous to the detection of H. pylori by upper gastrointestinal endoscopy. The eradication rate after using the triple scheme for 14 days was low in the included patients.
The eradication rate of H. pylori in the general population after treatment with the first line triple scheme (proton pump inhibitor, amoxicillin, and clarithromycin) has decreased in recent years, especially after the use of the shorter scheme (7 days) 15 . It was 93.5% in 2003 and by 2012, it had reduced to 78.8% 15 . Some meta-analyses show higher eradication rates with a longer treatment of 14 days 16,17,18,19 . Recent literature reviews demonstrated that the increase in triple therapy duration increased the H. pylori eradication rate (72.9 vs. 81.9%), independently of the type and dosage of antimicrobials 17 .
The eradication rate of H. pylori in chronic kidney disease patients varies according to the antimicrobial schemes used, whether triple, quadruple, or sequential. In the Seyyedand and collaborators study, the triple scheme with 30 mg lansoprazole twice a day, 250 mg clarithromycin twice a day, and 500 mg amoxicillin twice a day for 14 days, presented a 76.7% eradication rate for H. pylori 20   mg clarithromycin, and 1000 mg amoxicillin twice a day for 14 days, while the other group of 31 patients received the same drugs throughout the same period, but once a day. The eradication rate of both groups was 73% with no statistical difference between the two regimes (p= 0.973) 21 .
Although the extended time frame of the triple treatment was preferred by our patients, we found a lower eradication rate. This could have been due to poor compliance to the prolonged treatment or an unknown resistance to one of the medications used, especially clarithromycin. Resistance to clarithromycin is still the most common cause of triple therapy failure, and the period of treatment does not affect the high rates of clarithromycin resistance in the general population 13 . In a previous study that evaluated the 5 macro-regions of Brazil, bacterial resistance to clarithromycin varied from 15 to 20% 22 . The current study could not assess resistance to clarithromycin, since neither a culture, nor an antibiogram were conducted 13 . The influence of pre-therapeutic histological parameters on H. pylori eradication rate is still controversial. Georgopoulos and collaborators suggested that the coexistence of high scores of antral gastritis degree and activity with any degree of corpus gastritis may favorably affect the outcome of treatment, supporting the idea of facilitated diffusion of antibiotics in the inflamed mucosa 23 . However, in the current work, we found a lower eradication rate, although the most prevalent endoscopic finding was pangastritis and enema.
H. pylori is the most important etiological factor for gastric cancer. H. pylori causes chronic gastric inflammation that can progress to precancerous alterations in atrophic gastritis and in intestinal metaplasia. The risk of gastric cancer increases according to the extension and severity of these precancerous alterations. Eradication of H. pylori can induce resolution of gastric inflammation, stop progression of gastric mucosa damage, prevent additional damage to the DNA induced by H. pylori, improve gastric acid secretion, and restore normality of the internal environment 8 . Thus, we believe that treatment with an antimicrobial regime should be considered in patients with H. pylori and chronic kidney disease.
Patients with chronic kidney disease present a greater risk of gastroduodenal disorders. It is recommended that all hemodialysis and peritoneal dialysis patients be submitted to endoscopic evaluation to reduce the chances of developing peptic ulcers, especially in patients with a history of gastroduodenal bleeding or use of anticoagulants and/or non-steroidal anti-inflammatory drugs 24 . We used two diagnostic methods associated with upper gastrointestinal endoscopy to increase the probability of detection of H. pylori. The literature demonstrates high sensitivity and specificity of the rapid urease test, varying between 80 and 100% and 97 and 99%, respectively 16,25 . However, in the current study, H. pylori detection by the rapid urease test was low. A disadvantage of the test that could explain this situation is false-negative results due to the reduction in the activity of urease caused by recent use of antimicrobials, bismuth compounds, or PBIs, or because of achlorhydria. In addition, the presence of gastric bleeding from uremia reduces the sensitivity and specificity of the method. Furthermore, the amount of bacteria present affects the sensitivity of the test; quantities above 10,000 in the sample indicate positive results, while quantities below this can generate false negatives. The rapid urease test should be performed with a biopsy of one gastric region (antrum), differently from the histology from biopsy of two gastric regions (body and antrum). For now, we suggest that the association of both methods in our patients increased the evaluation costs and did not lead to an increase in detection rate.
Patients with chronic kidney disease have a high incidence of gastrointestinal diseases, even though the occurrence and type of symptoms can vary considerably between patients 11 . Many gastrointestinal symptoms such as anorexia, nausea, vomiting, and dyspepsia are common in patients with chronic kidney disease waiting for a renal transplant. These symptoms could be indicative of uremia or the result of medications and electrolyte imbalance, which makes it difficult to confidently predict the presence of a meaningful lesion in the superior gastrointestinal tract. However, many chronic kidney disease patients with peptic ulcers are asymptomatic 26,27 and can present significant complications before and after renal transplant, especially during the period of high immunosuppression 28 . This finding is very relevant, as an active peptic ulcer is a contraindication for renal transplant 29 and strongly supports the recommendation for upper gastrointestinal endoscopies before transplantation. There is no solid evidence on the role of routine triage with upper gastrointestinal endoscopy for H. pylori in asymptomatic candidates during evaluation before renal transplantation and no consensus between transplant centers 30 . In a recent study performed in the same renal transplant center, Homse and collaborators showed that lesions in the gastrointestinal tract were found in all analyzed patients, even though patients did not present symptoms 6 . It is believed that the findings of the study justify the recommendation for an upper gastrointestinal endoscopy in the preparation routine for renal transplantation in chronic kidney disease patients.

implicatiOns, strengths anD limitatiOns
This is one of the first studies to evaluate findings from upper gastrointestinal endoscopies for pre-transplant preparation and the efficiency of H. pylori treatment in renal transplant candidates. The verification of drug compliance by phone contact 7 days after the beginning of treatment was also considered a strength of the study. Complete compliance was confirmed when patients were seen by the nephrologist after the end of treatment, but supervised pill-taking was not performed. An association was found between H. pylori infection and upper gastrointestinal endoscopy findings in this population. Some limitations of the study should be considered, such as the low number of patients analyzed and the losses to follow up, which were partly due to the difficulties faced by patients to access transplant centers to repeat the upper gastrointestinal endoscopy. Therefore, our results should be interpreted with caution. Another limitation of the study was that no upper digestive endoscopy was performed after kidney transplantation. This was not included in the original study, as the coverage of its performance in asymptomatic patients is limited in the National Health System. Another limitation was that two of the drugs used against H. pylori are filtered during hemodialysis (amoxicillin and clarithromycin). To minimize this effect, patients were instructed to take these medications after hemodialysis.
There is no clinical protocol for a detailed endoscopic evaluation and eradication therapy for H. pylori in patients on dialysis. Therefore, future studies must be developed to confirm and expand the findings of the current study 24,31 . conclusIon A high prevalence of H. pylori was found in renal transplant candidates, and triple antimicrobial therapy applied over a long period had a low eradication rate. The performance of routine upper gastrointestinal endoscopy in pre-transplant evaluation detected gastrointestinal lesions in most patients and the endoscopic findings did not relate to symptoms. The association of the rapid urease test with gastric mucosa histology did not increase the detection rate of H. pylori.