Micro-broth dilution method with air-dried microplate for determining MICs of clarithromycin and amoxycillin for Helicobacter pylori isolates

2004 ◽  
Vol 53 (5) ◽  
pp. 403-406 ◽  
Author(s):  
Intetsu Kobayashi ◽  
Hiroe Muraoka ◽  
Takeshi Saika ◽  
Minoru Nishida ◽  
Toshio Fujioka ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Close Agreement ◽  
Reliable Method ◽  
Agar Plate ◽  
Dilution Method ◽  
Air Drying ◽  
H Pylori ◽  
Broth Dilution

MICs of clarithromycin and amoxycillin for 253 isolates of Helicobacter pylori were measured by an air-dried microplate method and compared with the results obtained by the agar plate dilution method. The air-dried microplate method is performed by coating each well of a 96-well microplate with the test antibiotic and air-drying it. There were no marked differences between the air-dried microplate method and agar plate dilution methods in the MIC50 and MIC90 values or MIC ranges of clarithromycin obtained for the 253 isolates of H. pylori. More specifically, the MICs of clarithromycin for 114 (45.1 %) of the 253 isolates were the same by the air-dried microplate method as the agar plate dilution method, and the differences in the MICs of clarithromycin for a further 114 isolates (45.1 %) varied within one twofold dilution. The MICs of amoxycillin by the former method were in close agreement with the MICs obtained by the latter method: MICs of amoxycillin for 199 (78.7 %) of the 253 isolates were the same by both methods, and the differences in the MICs of amoxycillin for 42 isolates (16.6 %) varied within one twofold dilution. These results indicate that the air-dried microplate method is a useful method for determination of MICs, because the results obtained were in close agreement with those obtained by the standard agar plate dilution method. The air-dried microplate method is, therefore, a convenient and reliable method for determining the MICs of clarithromycin and amoxycillin for H. pylori isolates.

scholarly journals Potential of FTIR-Spectroscopy for Drugs Screening against Helicobacter pylori

Antibiotics ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 897
Author(s):  
Pedro Sousa Sampaio ◽  
Cecília R. C. Calado
Keyword(s):  
Helicobacter Pylori ◽  
Ftir Spectroscopy ◽  
Partial Least Square ◽  
Least Square ◽  
New Drugs ◽  
Partial Least Square Regression ◽  
Dilution Method ◽  
Agar Dilution Method ◽  
Molecular Fingerprint ◽  
H Pylori

Helicobacter pylori colonizes the human stomach of half of the world’s population. The infection if not treated, persists through life, leading to chronic gastric inflammation, that may progress to severe diseases as peptic ulcer, gastric adenocarcinoma, and mucosa-associated lymphoid tissue lymphoma. The first line of treatment, based on 7 to 21 days of two antibiotics associated with a proton pump inhibitor, is, however, already failing most due to patient non-compliance that leads to antibiotic resistance. It is, therefore, urgent to screen for new and more efficient antimicrobials against this bacterium. In this work, Fourier Transform Infrared (FTIR) spectroscopy was evaluated to screen new drugs against H. pylori, in rapid (between 1 to 6 h), and high-throughput mode and based on a microliter volume processes in relation to the agar dilution method. The reference H. pylori strains 26,695 and J99, were evaluated against a peptide-based antimicrobial and the clinical antibiotic clarithromycin, respectively. After optimization of the assay conditions, as the composition of the incubation mixture, the time of incubation, and spectral pre-processing, it was possible to reproducibly observe the effect of the drug on the bacterial molecular fingerprint as pointed by the spectra principal component analysis. The spectra, obtained from both reference strains, after its incubation with drugs concentrations lower than the MIC, presented peak ratios statistically different (p < 0.05) in relation to the bacteria incubated with drugs concentrations equal or higher to the MIC. It was possible to develop a partial least square regression model, enabling to predict from spectra of both bacteria strains, the drug concentration on the assay, with a high correlation coefficient between predicted and experimental data (0.91) and root square error of 40% of the minimum inhibitory concentration. All this points to the high potential of the technique for drug screening against this fastidious growth bacterium.

scholarly journals Implication of the Structure of the Helicobacter pylori cag Pathogenicity Island in Induction of Interleukin-8 Secretion

2001 ◽  
Vol 69 (3) ◽  
pp. 1625-1629 ◽  
Author(s):  
C. Audibert ◽  
C. Burucoa ◽  
B. Janvier ◽  
J. L. Fauchère
Keyword(s):  
Helicobacter Pylori ◽  
Clinical Presentation ◽  
Blot Hybridization ◽  
Pathogenicity Island ◽  
Interleukin 8 ◽  
Middle Region ◽  
Dot Blot ◽  
Long Distance ◽  
H Pylori

ABSTRACT Helicobacter pylori virulence is associated with the presence of the cag pathogenicity island (PAI). Thecag PAI is involved in the ability to induce interleukin-8 (IL-8) secretion by human cells, which is implicated in the inflammatory response of the gastric mucosa to H. pyloriinfection. The aim of this study was to determine whether the genetic structure of the cag PAI is conserved and whether it is linked to IL-8 induction ability. Detection of specific markers (cagA, picB, cag13-cag14, virD4, and IS605) by PCR and dot blot hybridization and long-distance PCR determination of the presence of cagI, cagII, and the middle region of thecag PAI were performed on 153 strains isolated from adults suffering from ulcers (n = 79) or gastritis (n = 74). IL-8 induction ability was evaluated by coculture of the strains with HEp-2 cells. Eighty-three strains (54.3%) had an entire cag PAI, 12 strains (7.8%) had thecag PAI split in two, 49 strains (32%) had nocag PAI, and 9 strains exhibited other structural combinations. The presence of an entire cag PAI was statistically correlated with the presence of IS605(P = 0.006) and the ability to induce IL-8 secretion but not with clinical presentation of the infection. The structure of the cag PAI appears to be rather conserved and is related to the proinflammatory power of a strain. The existence of strains inducing IL-8 secretion regardless of the cag PAI structure suggests that this region is not the only requirement for IL-8 secretion.

scholarly journals New Site of Modification of 23S rRNA Associated with Clarithromycin Resistance of Helicobacter pylori Clinical Isolates

2002 ◽  
Vol 46 (12) ◽  
pp. 3765-3769 ◽  
Author(s):  
Carla Fontana ◽  
Marco Favaro ◽  
Silvia Minelli ◽  
Anna Angela Criscuolo ◽  
Antonio Pietroiusti ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
23S Rrna ◽  
Dilution Method ◽  
Rrna Gene ◽  
Functional Sites ◽  
Biopsy Specimens ◽  
Clarithromycin Resistance ◽  
Pcr Product ◽  
23S Rrna Gene ◽  
H Pylori

ABSTRACT Resistance of Helicobacter pylori to clarithromycin occurs with a prevalence ranging from 0 to 15%. This has an important clinical impact on dual and triple therapies, in which clarithromycin seems to be the better choice to achieve H. pylori eradication. In order to evaluate the possibility of new mechanisms of clarithromycin resistance, a PCR assay that amplified a portion of 23S rRNA from H. pylori isolates was used. Gastric tissue biopsy specimens from 230 consecutive patients were cultured for H. pylori isolation. Eighty-six gastric biopsy specimens yielded H. pylori-positive results, and among these 12 isolates were clarithromycin resistant. The latter were studied to detect mutations in the 23S rRNA gene. Sequence analysis of the 1,143-bp PCR product (portion of the 23S rRNA gene) did not reveal mutation such as that described at position 2142 to 2143. On the contrary, our findings show, for seven isolates, a T-to-C transition at position 2717. This mutation conferred a low level of resistance, equivalent to the MIC for the isolates, selected using the E-test as well as using the agar dilution method: 1 μg/ml. Moreover, T2717C transition is located in a highly conserved region of the 23S RNA associated with functional sites: domain VI. This fact has a strong effect on the secondary structure of the 23S RNA and on its interaction with macrolide. Mutation at position 2717 also generated an HhaI restriction site; therefore, restriction analysis of the PCR product also permits a rapid detection of resistant isolates.

scholarly journals Antimicrobial Sensitivity Pattern of Helicobacter pylori Isolates among Subgroup of Bangladeshi Patients

2014 ◽  
Vol 8 (2) ◽  
pp. 49-52
Author(s):  
MMSU Islam ◽  
Shamsun Nahar ◽  
Mst Naznin Sarker ◽  
ASM Salimullah ◽  
Mohammad Asadur Rahman ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Diarrhoeal Disease ◽  
Cross Sectional Study ◽  
Dilution Method ◽  
Agar Dilution Method ◽  
Cross Sectional ◽  
Antimicrobial Sensitivity ◽  
Sensitivity Pattern ◽  
H Pylori ◽  
Medical University

This cross sectional study was carried out at Bangabandhu Sheikh Mujib Medical University (BSMMU) and International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR,B) from July 2008 to September 2009. Aim of the study was to find out the antimicrobial susceptibility profile of Helicobacter pylori isolates from dyspeptic patients. Total 224 dyspeptic patients from Out Patient Department (OPD) of BSMMU were initially enrolled after informed written consent. After upper GI endoscopy 157 patients were finally included who had erosions, ulcers or atrophic changes in the stomach or duodenum. Two biopsy samples were taken from each of them. Samples were incubated at 37°C in a double gas incubator with 5%O2, 10%CO2 and 85%N2. Total 82 (52.23%) samples were found positive for H. pylori. Isolated organisms were then tested for sensitivity to Amoxicillin, Clarithromycin, Tetracycline, Levofloxacin and Metronidazole by Agar dilution method. Among 82 patients 51(62.2%) were male and 31(37.8) were female with a male:female ratio 1.6:1. Patients were categorized into two groups one having gastric or duodenal ulcer (30.5%) and other having no ulcer (69.5%). Among these isolates 92.7% were sensitive to Amoxicillin, 89% to Clarithromycin, 81.7% to Tetracycline, 80.5% to Levofloxacin and only 26.8% to Metronidazole. Beside these, 81.7% isolates were sensitive to both Amoxicillin and Clarithromycin, 74.4% to Amoxicillin and Tetracycline, 73.2% to Amoxicillin and Levofloxacin, 72% to Clarithromycin and Tetracycline, 59% to Clarithromycin and Levofloxacin and 51% to Tetracycline and Levofloxacin DOI: http://dx.doi.org/10.3329/fmcj.v8i2.20280 Faridpur Med. Coll. J. 2013;8(2): 49-52

scholarly journals Structure-biological activity relationship of synthetic trihydroxilated chalcones

1998 ◽  
Vol 29 (4) ◽  
pp. 307-310 ◽  
Author(s):  
Cristina M. Devia ◽  
Nora B. Pappano ◽  
Nora B. Debattista
Keyword(s):  
Staphylococcus Aureus ◽  
Biological Activity ◽  
Agar Plate ◽  
Dilution Method ◽  
Bacteriostatic Activity ◽  
Plate Method ◽  
Minimal Inhibitory Concentrations ◽  
Structure Activity ◽  
Relationship Of ◽  
Broth Dilution

The bacteriostatic activity of 2’,4’,2-trihydroxychalcone; 2’,4’,3-trihydroxychalcone and 2’,4’,4-trihydroxychalcone, prepared by condensation of 2,4-dihydroxyacetophenone and benzaldehyde substituted, against Staphylococcus aureus ATCC 25923 was assayed by agar plate method. The three compounds presented important inhibition halos. In order to elucidate structure-activity relationships, the minimal inhibitory concentrations against S. aureus were determined by the broth dilution method and the results obtained were compared to that of 2',4'-dihydroxychalcone. The sequence observed was: MIC 2’,4’,3-(OH)3 > MIC 2’,4’-(OH)2 > MIC 2’,4’,4-(OH)3 > > MIC 2’,4’,2-(OH)3. These results showed that the introduction of an electron donating group (-OH) in the aromatic B-ring causes an increase in bioactivity, and that the intensity of action depends on the position of the OH substitute.

scholarly journals HELICOBACTER PYLORI INFECTION;

2017 ◽  
Vol 24 (09) ◽  
pp. 1425-1430
Author(s):  
Tahira Tabassum ◽  
Ayesha Imtiaz ◽  
Aamir Sharif ◽  
Muhammad Akram
Keyword(s):  
Helicobacter Pylori ◽  
Age Groups ◽  
Helicobacter Pylori Infection ◽  
Step Test ◽  
Chromatographic Technique ◽  
Medical College ◽  
Education Levels ◽  
One Step ◽  
H Pylori

Objectives: To estimate the prevalence of Helicobacter pylori infection inpopulation of Sargodha and determination of possible risk factors. Design: Descriptive study.Place and Duration: University medical complex & Research Center, Sargodha Medical College,Sargodha from 01.01.2017 to 31.03.2017. Material and Methods: The study was conductedamong 486 clinically suspected individuals of Sargodha, Punjab, Pakistan. Data was obtainedby questionnaire and H. pylori antibodies were detected by H. pylori ‘One Step Test Device’based on immune-chromatographic technique. Results: A total of 486 individuals participatedin the study of which 327 (67.28%) were positive for H. pylori infection. The prevalence was66.97% in males and 33.2% in females (p = 0.00) and increased with increasing age (p =0.000). In the study, significant association was found between age groups, education levels,food habits, monthly income with H. pylori infection with p values 0.000, 0.000, 0.001, 0.041respectively while no significant association was seen for source of drinking water (p =0.321).Conclusion: The results of present study showed the prevalence of H. pylori in reported areaand confirmed that it is higher in developing countries. It is concluded that prevalence ofinfection with this bacterium can be minimized by increasing the hygenicity, improving the livingstandards and, of course, by educating the nation.

scholarly journals Helicobacter pylori antibiotic susceptibility patterns in Bangladesh: Emerging levofloxacin resistance

2016 ◽  
Vol 10 (03) ◽  
pp. 245-253 ◽  
Author(s):  
Hafeza Aftab ◽  
Muhammad Miftahussurur ◽  
Phawinee Subsomwong ◽  
Faruque Ahmed ◽  
AK Azad Khan ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Triple Therapy ◽  
Antibiotic Susceptibility ◽  
Resistance Rate ◽  
Resistance Pattern ◽  
Dilution Method ◽  
Agar Dilution Method ◽  
Peptic Ulcers ◽  
H Pylori ◽  
Resistance Rates

Introduction: The most recent study to report Helicobacter pylori antibiotic resistance rates in Bangladesh was published 15 years ago and did not include levofloxacin. We therefore aimed to determine the current antibiotic susceptibility of H. pylori to amoxicillin, clarithromycin, metronidazole, tetracycline and levofloxacin in Bangladesh. Methodology: This study included 133 consecutive patients who underwent endoscopy examination at Dhaka Medical College in November 2014. The serial two-fold agar dilution method was used to determine the minimum inhibitory concentrations of the five antibiotics. Results: Among 56 cultured strains, H. pylori showed high rates of resistance to clarithromycin and metronidazole (39.3% and 94.6%, respectively). Moreover, levofloxacin showed an emerging antimicrobial resistance pattern (66.1%), which was higher in patients with gastritis than that in those with peptic ulcers (p = 0.02). The resistance rate of levofloxacin was significantly higher in patients living in Dhaka city compared to those living in the village (p = 0.049). However, amoxicillin and tetracycline resistance rates were very low. Resistance to both metronidazole and levofloxacin was most commonly observed. Conclusions: The rates of resistance to clarithromycin, metronidazole, and levofloxacin were high in Bangladesh, which suggests that triple therapy based on these drugs may not be useful as first-line therapies in Bangladesh. Alternative strategies such as furazolidone-based triple therapy, bismuth-based quadruple therapies, or sequential therapy may be more effective for patients in in Bangladesh.

scholarly journals High Prevalence of Antibiotic Resistance in Iranian Helicobacter pylori Isolates: Importance of Functional and Mutational Analysis of Resistance Genes and Virulence Genotyping

2019 ◽  
Vol 8 (11) ◽  
pp. 2004 ◽  
Author(s):  
Nastaran Farzi ◽  
Abbas Yadegar ◽  
Amir Sadeghi ◽  
Hamid Asadzadeh Aghdaei ◽  
Sinéad Marian Smith ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Antibiotic Resistance ◽  
Rrna Genes ◽  
23S Rrna ◽  
Dilution Method ◽  
Agar Dilution Method ◽  
Gyra Gene ◽  
Therapeutic Regime ◽  
High Prevalence ◽  
H Pylori

The high prevalence of antibiotic resistance in Helicobacter pylori has become a great challenge in Iran. The genetic mutations that contribute to the resistance have yet to be precisely identified. This study aimed to investigate the prevalence of antibiotic resistance and virulence markers in Iranian H. pylori isolates and to analyze if there is any association between resistance and genotype. Antibiotic susceptibility patterns of 68 H. pylori isolates were investigated against metronidazole, clarithromycin, amoxicillin, rifampicin, ciprofloxacin, levofloxacin, and tetracycline by the agar dilution method. The frxA, rdxA, gyrA, gyrB, and 23S rRNA genes of the isolates were sequenced. The virulence genotypes were also determined using PCR. Metronidazole resistance was present in 82.4% of the isolates, followed by clarithromycin (33.8%), ciprofloxacin (33.8%), rifampicin (32.4%), amoxicillin (30.9%), levofloxacin (27.9%), and tetracycline (4.4%). Overall, 75% of the isolates were resistant to at least two antibiotics tested and considered as a multidrug resistance (MDR) phenotype. Most of the metronidazole-resistant isolates carried frameshift mutations in both frxA and rdxA genes, and premature termination occurred in positions Q5Stop and Q50Stop, respectively. Amino acid substitutions M191I, G208E, and V199A were predominantly found in gyrA gene of fluoroquinolone-resistant isolates. A2143G and C2195T mutations of 23S rRNA were found in four clarithromycin-resistant isolates. Interestingly, significant associations were found between resistance to metronidazole (MNZ) and cagA-, sabA-, and dupA-positive genotypes, with p = 0.0002, p = 0.0001, and p = 0.0001, respectively. Furthermore, a significant association was found between oipA “on” status and resistance to amoxicillin (AMX) (p = 0.02). The prevalence of H. pylori antibiotic resistance is high in our region, particularly that of metronidazole, clarithromycin, ciprofloxacin, and MDR. Simultaneous screening of virulence and resistance genotypes can help clinicians to choose the appropriate therapeutic regime against H. pylori infection.

Activity of Bulgarian propolis against 94 Helicobacter pylori strains in vitro by agar-well diffusion, agar dilution and disc diffusion methods

2005 ◽  
Vol 54 (5) ◽  
pp. 481-483 ◽  
Author(s):  
Lyudmila Boyanova ◽  
Galina Gergova ◽  
Rossen Nikolov ◽  
Sirigan Derejian ◽  
Elena Lazarova ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Growth Inhibition ◽  
Biological Effects ◽  
Diffusion Method ◽  
Dilution Method ◽  
Agar Dilution Method ◽  
Propolis Extract ◽  
Agar Dilution ◽  
H Pylori

Propolis exhibits antimicrobial, anti-inflammatory and other biological effects. The aim of this study was to evaluate the activity of 30 % ethanolic extract of Bulgarian propolis against 94 Helicobacter pylori strains by three methods. By the agar-well diffusion method, only 13.8 % of the strains exhibited no inhibition by 30 μl propolis extract (containing 9 mg propolis) and all isolates were inhibited to some extent by 90 μl of the extract (27 mg propolis) per well. The mean diameters of growth inhibition by 30, 60 or 90 μl propolis extract or 30 μl 96 % ethanol per well were 16.8, 19.2, 27.5 and 8.3 mm, respectively. The propolis extract was more active than the ethanol (P < 0.001). With 90 μl propolis extract per well, 69.4 % of the strains exhibited large diameters of growth inhibition (⩾20 mm) versus 26.6 % with 30 μl per well (P < 0.001). With moist propolis discs, inhibition was detected in more strains (92.1 %) than with dried discs (78.2 %, P < 0.05), with mean inhibitory diameters of 18.7 and 13.8 mm, respectively. By the agar dilution method, 100 and 300 μg propolis ml−1 inhibited the growth of 57.1 % and 76.2 %, respectively, of the 21 strains tested. In conclusion, Bulgarian propolis had a strong and dose-dependent activity against most of the H. pylori strains tested. Although the effect of propolis on H. pylori in vitro is promising, further microbiological, pharmacological and clinical trials are required.

scholarly journals Molecular Detection of Antibiotic Resistance in South African Isolates ofHelicobacter pylori

2013 ◽  
Vol 2013 ◽  
pp. 1-6 ◽  
Author(s):  
Nicoline F. Tanih ◽  
Roland N. Ndip
Keyword(s):  
Helicobacter Pylori ◽  
Antibiotic Resistance ◽  
South African ◽  
Molecular Detection ◽  
Rapid Diagnosis ◽  
Biopsy Specimens ◽  
H Pylori ◽  
Detection Of Mutations ◽  
Reverse Hybridisation Assay

Rapid diagnosis and treatment ofHelicobacter pylori(H. pylori) presents a challenge. We aimed at investigating the presence ofH. pylori, susceptibility profile, and associated mutations in an effort to validate the effectiveness of GenoType HelicoDR assay inH. pylorityping in our environment. Two hundred and fifty-four biopsy specimens were cultured and DNA extracted from seventy-eight positive cultures using the Qiagen DNA extraction kit. The GenoType Helico DR which employs reverse hybridisation was used to confirm the presence ofH. pylori, determination of its susceptibility to antimicrobials, and detection of mutations conferring resistance to clarithromycin and fluoroquinolones. The organism was isolated from 168/254 (66.1 %) of the specimens by culture. Of the 78 strains used for further investigation, 12/78 (15.38%) were resistant to clarithromycin while 66/78 (84.61%) were susceptible. For fluoroquinolone, 70/78 (89.74%) strains were susceptible while 8 (10.26%) were resistant. Mutations were observed in 17 strains with A2147G being the most prevalent; A2146C and D91N were the least. The reverse hybridisation assay is an easy and fast technique in confirming the presence ofH. pylori, its antimicrobial profile, and associated mutations. Analysis regarding the suitability of this assay forH. pylorityping is warranted in other regions.

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