Molecular Characterization and Mutational Analysis of Clarithromycin- and Levofloxacin-Resistance Genes in Helicobacter pylori from Gastric Biopsies in Southern Croatia.

Point mutations in the 23S rRNA, gyrA, and gyrB genes can confer resistance to clarithromycin (CAM) and levofloxacin (LVX) by altering target sites or protein structure, thereby reducing the efficacy of standard antibiotics in the treatment of Helicobacter pylori infections. Considering the confirmed primary CAM and LVX resistance in H. pylori infected patients from southern Croatia, we performed a molecular genetic analysis of three target genes (23S rRNA, gyrA, and gyrB) by PCR and sequencing, together with computational molecular docking analysis. In the CAM-resistant isolates, the mutation sites in the 23S rRNA gene were A2142C, A2142G, and A2143G. In addition, the mutations D91G and D91N in GyrA and N481E and R484K in GyrB were associated with resistance to LVX. Molecular docking analyses revealed that mutant H. pylori strains with resistance-related mutations exhibited a lower susceptibility to CAM and LVX compared with wild-type strains due to significant differences in non-covalent interactions (e.g., hydrogen bonds, ionic interactions) leading to destabilized antibiotic-protein binding, ultimately resulting in antibiotic resistance. Dual resistance to CAM and LVX was found, indicating the successful evolution of H. pylori resistance to unrelated antimicrobials and thus an increased risk to human health.

Helicobacter pylori Infection in Croatian Population: Knowledge, Attitudes and Factors Influencing Incidence and Recovery.

Basic and clinical knowledge about Helicobacter pylori infections has been improved in the past. However, the translation of this knowledge into public health intervention has remained poor. A survey based cross-sectional study was performed to assess the factors regarding the H. pylori infection in the general population. The survey was conducted using a previously developed questionnaire, adapted for the population in Croatia. Respondents (N = 1131) had a good knowledge score with a median of 4 out of 5 correct answers (interquartile range: 2-4). Senior participants had a lower frequency of high knowledge answers about H. pylori (43.1%) compared to younger (56.1%) and middle-aged participants (51.5%, p = 0.014). Rural participants had a higher frequency of low knowledge answers compared to urban and suburban ones (21.7% vs. 9.5% and 9.4%, p = 0.011). Only 315 participants (27.9%) were screened for the H. pylori infection, despite high support for the screening programs among the untested (74.7%) and tested (85.7%). Habits of smoking (p = 0.036) and coffee drinking (p = 0.008) were associated with more symptoms after eradication therapy. Further education is needed for the groups at risk for H. pylori infection, especially to raise the awareness of the importance of screening programs. More research is warranted to assess the effects of dietary changes on therapy outcomes.

Adherence to Maastricht V/Florence consensus report for the management of Helicobacter pylori infection among primary care physicians and medical students in Croatia: A cross-sectional study.

BACKGROUND: Helicobacter pylori (H pylori) eradication is becoming increasingly difficult. The aim of our study was to determine the knowledge of current guidelines and attitude in the diagnosis and treatment of H pylori infection in primary care physicians (PCPs) and medical students in Croatia. MATERIALS AND METHODS: A study was conducted among PCPs and medical students to evaluate adherence to Maastricht V consensus guidelines. Questionnaire was distributed by e-mail to 2338 PCPs offices in Croatia and to the medical students from the University of Split School of Medicine. Responses were collected electronically from June 22 to August 22, 2020. RESULTS: Two hundred forty-nine PCPs and 169 medical students were included in the study. Bismuth or non-bismuth-based quadruple therapy as first-line treatment for H pylori was the choice of 4.8% of PCPs and 13% of students, while 66.3% PCPs and 79.9% students would choose clarithromycin-based triple therapy. Bismuth-based quadruple therapy was the most preferred second line of treatment for 45.4% of PCPs and 34.9% of students. Only 2.8% PCPs and 7.1% of students would correctly recommend first and second line of treatment for H. pylori infection. A larger proportion of students than PCPs would prefer C13-urea breath test (50.3% vs 6.4%). Only 59.0% PCPs would treat for H pylori in all patients including the asymptomatic ones. Students more frequently recognized the link between H pylori and gastric cancer compared with PCPs (92.9% vs 73.5%). CONCLUSIONS: Primary care physicians and medical students' knowledge of H pylori guidelines are insufficient in Croatia and ask for additional training.

Fatal Hepatitis-Associated Aplastic Anemia in a Young Male.

Hepatitis-associated aplastic anemia is a rare syndrome in which bone marrow failure occurs within weeks to 1 year after attack of acute hepatitis. Studies suggest that cytotoxic T lymphocytes play a central role in bone marrow destruction, but the exact etiology remains unknown. Bone marrow transplantation or immunosuppressive therapy are primary curative options. We present a case of a young male who was admitted to the Department of Gastroenterology and Hepatology for acute hepatitis of an unknown cause. Liver biopsy revealed extensive inflammatory process with hepatocyte necrosis. Forty days later, new onset pancytopenia was identified. Bone marrow biopsy showed severe hypocellularity, and he was diagnosed with severe hepatitis-associated aplastic anemia. Treatment with cyclosporine was initiated, but with inadequate response, and pretransplant evaluation was started. Due to severe neutropenia, following alveotomy procedure, the patient developed deep neck infection with consequent airway obstruction. Despite urgent treatment, his condition deteriorated to sepsis with lethal outcome.

Signet Ring Carcinoma of the Appendix Presenting as Crohn's Disease in a Young Male.

Primary signet ring cell carcinoma is a rare event in surgery. It looks like acute appendicitis and it is difficult to diagnose it on clinical grounds alone. The diagnosis is always confirmed by histopathology of a surgically removed appendix. A young man, 22 years old, presented with vomiting, diarrhea, and cramps in his abdomen without abdominal tenderness (mild abdominal discomfort in the right lower abdominal quadrant without signs of peritoneal irritation) during the previous month. The first endoscopic results showed only changes of mucosa that could be attributed to endoscopic and clinical representation of Crohn's disease. A few days after the initiation of the therapy with aminosalicylates and corticosteroids, the patient went into ileus and was transferred to the Department of Surgery, where he underwent an emergency right-sided hemicolectomy with resection of the transversal colon and forming of an ileostoma. The first pathohistological diagnosis was pseudomembranous colitis. Because the patient's condition was deteriorating, a revision of the pathohistological diagnosis was done. After careful revision and extensive sampling, a signet ring cell carcinoma arising in the appendix with infiltration of the ileocecal region was found. Immunohistochemically, tumor cells were positive for CDX-2 CK7, CK20, CK19, and carcinoembryonic antigen and negative for chromogranin A. Sixteen isolated lymph nodes were negative. Although the patient had a disease that was localized to the appendix and ileocecal region with no apparent distal metastasis, his clinical condition was worsening rapidly and he died after 2 months. This case shows the aggressive biological behavior of the appendix signet ring cell carcinoma. Scrupulous histopathological examination of the appendix is an obligatory procedure. Elimination of the signet ring cell carcinoma from other carcinoma subtypes is of special importance as it has an exceptionally poor prognosis and is generally diagnosed in its advanced stages.

Diagnosis of Helicobacter pylori infection : A short review.

Helicobacter pylori infections represent an important factor in the pathogenesis of chronic gastritis, peptic ulcer, MALT lymphoma and gastric adenocarcinoma. The recently published Maastricht V/Florence consensus report indicated that the urea breath test using 13 C urea still remains the best non-invasive test to diagnose H. pylori infections with high sensitivity and specificity. Among the stool antigen tests, the ELISA monoclonal antibody test is a rational option. Effective therapy should be based only on susceptibility testing in regions with documented high clarithromycin resistance (>15%). Advanced high-resolution endoscopic technologies enable increased diagnostic accuracy for detection of H. pylori infections.

Acute, food-induced moderate elevation of plasma uric acid protects against hyperoxia-induced oxidative stress and increase in arterial stiffness in healthy humans.

We examined the effects of acute, food-induced moderate increase of plasma uric acid (UA) on arterial stiffness and markers of oxidative damage in plasma in healthy males exposed to 100% normobaric oxygen. Acute elevation of plasma UA was induced by consumption of red wine, combination of ethanol and glycerol, or fructose. By using these beverages we were able to separate the effects of UA, wine polyphenols and ethanol. Water was used as a control beverage. Ten males randomly consumed test beverages in a cross-over design over the period of 4 weeks, one beverage per week. They breathed 100% O(2) between 60(th) and 90(th)min of the 4-h study protocol. Pulse wave augmentation index (AIx) at brachial and radial arteries, plasma antioxidant capacity (AOC), thiobarbituric acid-reactive substances (TBARS), lipid hydroperoxides (LOOH) assessed by xylenol orange method, UA and blood ethanol concentrations were determined before and 60, 90, 120, 150 and 240 min after beverage consumption. Consumption of the beverages did not affect the AIx, TBARS or LOOH values during 60 min before exposure to hyperoxia, while AOC and plasma UA increased except in the water group. Significant increase of AIx, plasma TBARS and LOOH, which occurred during 30 min of hyperoxia in the water group, was largely prevented in the groups that consumed red wine, glycerol+ethanol or fructose. In contrast to chronic hyperuricemia, generally considered as a risk factor for cardiovascular diseases and metabolic syndrome, acute increase of UA acts protectively against hyperoxia-induced oxidative stress and related increase of arterial stiffness in large peripheral arteries.

The increase in human plasma antioxidant capacity after red wine consumption is due to both plasma urate and wine polyphenols.

By using red wine, dealcoholized red wine, polyphenols-stripped red wine, ethanol-water solution and water, the role of wine polyphenols and induction of plasma urate elevation on plasma antioxidant capacity was examined in humans (n=9 per beverage). Healthy males randomly consumed each beverage in a cross-over design. Plasma antioxidant capacity (measured by ferric reducing antioxidant power, FRAP), ethanol, catechin and urate concentrations were determined before and 30, 60, 90, 120 and 180 min after beverage intake. Dealcoholized red wine and polyphenols-stripped red wine induced similar increase in FRAP values which represented nearly half the effect of the original red wine. This indicates that consumption of red wine involves two separate mechanisms in elevation of plasma FRAP values and both wine phenols and plasma urate contribute to that effect.

Red wine induced modulation of vascular function: separating the role of polyphenols, ethanol, and urates.

By using red wine (RW), dealcoholized red wine (DARW), polyphenols-stripped red wine (PSRW), ethanol-water solution (ET), and water (W), the role of wine polyphenols, ethanol, and urate on vascular function was examined in humans (n = 9 per beverage) and on isolated rat aortic rings (n = 9). Healthy males randomly consumed each beverage in a cross-over design. Plasma ethanol, catechin, and urate concentrations were measured before and 30, 60 and 120 minutes after beverage intake. Endothelial function was assessed before and 60 minutes after beverage consumption by normalized flow-mediated dilation (FMD). RW and DARW induced similar vasodilatation in the isolated vessels whereas PSRW, ET, and W did not. All ethanol-containing beverages induced similar basal vasodilatation of brachial artery. Only intake of RW resulted in enhancement of endothelial response, despite similar plasma catechin concentration after DARW. The borderline effect of RW on FMD (P = 0.0531) became significant after FMD normalization (P = 0.0043) that neutralized blunting effect of ethanol-induced basal vasodilatation. Effects of PSRW and ET did not differ although plasma urate increased after PSRW and not after ET, indicating lack of urate influence on endothelial response. Acute vascular effects of RW, mediated by polyphenols, cannot be predicted by plasma catechin concentration only.

Malignant and non-malignant asbestos-related pleural and lung disease: 10-year follow-up study.

AIM: To examine the presence of radiologically visible lung and pleural changes in patients who were exposed to the asbestos dust, and to correlate the progression of these changes with the duration and intensity of exposure and smoking. We also evaluated possible correlation between non-malignant asbestos-related pleural abnormalities and the occurrence of malignant pleural mesothelioma. METHODS: Among 7,300 patients who visited our department between 1991 and 2000 due to non-specific respiratory symptoms, we selected 2,420 with chest X-rays indicating the possible existence of non-malignant asbestos-related diseases. The selected group was followed-up for progression of radiological changes and the development of malignant pleural mesothelioma, and the changes were correlated with the intensity and duration of exposure to asbestos dust and smoking. RESULTS: Radiological changes characteristic for non-malignant asbestos-related pleural disease or lung asbestosis were identified in 340 (14%) out of 2,420 examined patients, of whom 77 (22.6%) developed malignant pleural mesothelioma, as compared with 13 patients out of 2,080 (0.6%) without radiological signs of asbestosis or pleural changes. Twenty-three (29.9%) patients who presented with a progression of pleural disease and lung asbestosis had a very significant incidence of malignant pleural mesothelioma (p<0.001). We also found that 55 (71.4%) patients with the highest asbestos exposure level (grade 3) developed malignant pleural mesothelioma more often (p=0.044). No correlation was found between malignant pleural mesothelioma development and duration of asbestos exposure (p=0.149) or smoking habit (p=0.617). Professionally exposed patients were at 3.3-times higher relative risk (95% confidence interval, 2.28-4.75) than those who were not exposed to develop malignant pleural mesothelioma. CONCLUSIONS: The risk of developing lung asbestosis increased with the level of exposure to asbestos dust and smoking. The risk of developing pleural disease correlated with the intensity and duration of exposure, but not with smoking. The patients with progressive pleural and parenchymal changes are at particularly high risk of developing malignant pleural mesothelioma and must be under special surveillance.