scholarly journals Development of an Internal Control for Evaluation and Standardization of a Quantitative PCR Assay for Detection of Helicobacter pylori in Drinking Water

2007 ◽  
Vol 73 (22) ◽  
pp. 7380-7387 ◽  
Author(s):  
Keya Sen ◽  
Nancy A. Schable ◽  
Dennis J. Lye
Keyword(s):  
Helicobacter Pylori ◽  
Drinking Water ◽  
Quantitative Pcr ◽  
Water Samples ◽  
Sodium Thiosulfate ◽  
Qpcr Assay ◽  
E Coli ◽  
H Pylori ◽  
Labeled Probe ◽  
Treated Drinking Water

ABSTRACT Due to metabolic and morphological changes that can prevent Helicobacter pylori cells in water from growing on conventional media, an H. pylori-specific TaqMan quantitative PCR (qPCR) assay was developed that uses a 6-carboxyfluorescein-labeled probe (A. E. McDaniels, L. Wymer, C. Rankin, and R. Haugland, Water Res. 39:4808-4816, 2005). However, proper internal controls are needed to provide an accurate estimate of low numbers of H. pylori in drinking water. In this study, the 135-bp amplicon described by McDaniels et al. was modified at the probe binding region, using PCR mutagenesis. The fragment was incorporated into a single-copy plasmid to serve as a PCR-positive control and cloned into Escherichia coli to serve as a matrix spike. It was shown to have a detection limit of five copies, using a VIC dye-labeled probe. A DNA extraction kit was optimized that allowed sampling of an entire liter of water. Water samples spiked with the recombinant E. coli cells were shown to behave like H. pylori cells in the qPCR assay. The recombinant E. coli cells were optimized to be used at 10 cells/liter of water, where they were shown not to compete with 5 to 3,000 cells of H. pylori in a duplex qPCR assay. Four treated drinking water samples spiked with H. pylori (100 cells) demonstrated similar cycle threshold values if the chlorine disinfectant was first neutralized by sodium thiosulfate.

scholarly journals High frequency of Helicobacter pylori DNA in drinking water in Kermanshah, Iran, during June–November 2012

2013 ◽  
Vol 12 (3) ◽  
pp. 504-512 ◽  
Author(s):  
Alvandi Amirhooshang ◽  
Abiri Ramin ◽  
Aryan Ehsan ◽  
Rezaei Mansour ◽  
Bagherabadi Shahram
Keyword(s):  
Helicobacter Pylori ◽  
Drinking Water ◽  
Water Samples ◽  
Tap Water ◽  
Piping System ◽  
Detection Rates ◽  
High Prevalence ◽  
Polymerase Chain ◽  
H Pylori ◽  
Drinking Water Samples

To gain a better understanding of transmission and selecting appropriate measures for preventing the spread of Helicobacter pylori, the aim of this study was to investigate the prevalence of H. pylori in drinking water samples in Kermanshah, Iran. Drinking water samples were collected from around Kermanshah and filtered through 0.45 μm nitrocellulose filters. The bacterial sediment was subjected to DNA extraction and polymerase chain reaction (PCR) for H. pylori detection using newly designed primers targeted at the conserved region of the ureC gene. The overall detection rates for H. pylori DNA in the water samples were 56% (66/118) with a frequency of 36% (25/70) in tap water samples and 85% (41/48) in wells. The detection limit was 50 bacteria per liter of filtered water and a pure H. pylori DNA copy number of 6 per reaction. Based on the evidence we may suggest that recontamination occurred and H. pylori entered into the water piping system through cracked or broken pipes or was released from established H. pylori biofilms on pipes. In conclusion, a high prevalence of H. pylori was detected in drinking water samples that strengthens the evidence of H. pylori transmission through drinking water.

scholarly journals Frequency of H. Pylori infection in children with recurrent abdominal pain at a Tertiary Care Hospital.

2020 ◽  
Vol 27 (02) ◽  
pp. 237-241
Author(s):  
Asim Khurshid ◽  
Shahid Ishaq ◽  
Mushtaq Ahmad
Keyword(s):  
Helicobacter Pylori ◽  
Drinking Water ◽  
Disease Duration ◽  
Family Income ◽  
Tertiary Care Hospital ◽  
Tertiary Care ◽  
Recurrent Abdominal Pain ◽  
Male Gender ◽  
Care Hospital ◽  
H Pylori

Objectives: Recurrent abdominal pain (RAP) impacts quality of life of the children. RAP also hampers education and physical activity of the children. Current study was aimed to find out the frequency of Helicobacter pylori in children with RAP in our tertiary care hospital. Study Design: Descriptive, cross-sectional study. Setting:  Department of Pediatric Medicine, Nishtar Hospital, Multan, Period: From 27-12-2017 to 26-06-2018. Material & Methods: A total of 185 patients suffering from RAP, aged 2-12 years, with a disease duration > 3 months, were enrolled. Age of the children, gender, duration of illness, number of episodes of pain, maternal literacy, family income, residential status, source of drinking water and h.pylori infection were calculated in these children. Post stratification chi-square test was applied to see its effect on H. Pylori infection. Results: Of these 185 study cases, 101 (54.6 %) were male patients while 84 (45.4%) were female. Mean age of our study cases was 7.57 ± 1.93 years. Of A total of 95 (51.4%) children belonged to rural areas and 90 (48.6 %) to urban areas. Helicobacter pylori infection was noted in 103 (55.7%) of our study cases. When helicobacter pylori  was stratified with regards to study variables, male gender, age < 8 years, monthly family income <Rs. 35000, source of drinking water as Hand Pump and disease duration < 6 months turned out to be statistically significant (P value < 0.05). Conclusion: Frequency of H.pylori was high in children with RAP. Helicobacter pylori was significantly associated with male gender, younger age, poor socioeconomic status, source of drinking water and disease duration.

scholarly journals Evaluation of DNA extraction methods for quantifying Helicobacter pylori in water with PCR inhibitors

2021 ◽  
Author(s):  
Eun-Sook Lee ◽  
So-Yang Cha ◽  
Jong-Soon Jung
Keyword(s):  
Helicobacter Pylori ◽  
Real Time ◽  
Dna Extraction ◽  
Water Samples ◽  
Real Time Pcr ◽  
Heavy Rain ◽  
Extraction Methods ◽  
Pcr Inhibitors ◽  
H Pylori ◽  
Dna Extraction Methods

Abstract DNA extraction methods were evaluated to reduce PCR inhibitors and quantify Helicobacter pylori directly from water samples using real-time PCR. Three nucleic acid extraction methods were evaluated for different types of water samples. While the QIAamp DNA mini kit for tissue was suitable for DNA extraction from treated water, the QIAamp DNA stool mini kit was still efficient in analyzing samples from river water after heavy rain and with high concentration of PCR inhibitors. The FastDNA SPIN Kit for Soil could extract DNA effectively from microbes in river and stream waters without heavy rain. Immunomagnetic separation (IMS) was used prior to DNA extraction and was a useful tool for reducing PCR inhibitors in influent and stream samples. H. pylori in various waters could be quantified directly by real-time PCR while minimizing the effect of PCR inhibitors by an appropriate method through the evaluation of DNA extraction methods considering the characteristics of the matrix water. The findings of the present study suggest that the types or characteristics of water sample by source and precipitation are an important factor in detecting H. pylori and they can be applied when detecting and monitoring of other pathogens in water.

scholarly journals Antibacterial Activity of Geraniol in Combination with Standard Antibiotics against Staphylococcus aureus, Escherichia coli and Helicobacter pylori

2018 ◽  
Vol 13 (7) ◽  
pp. 1934578X1801300 ◽  
Author(s):  
Subrat Kumar Bhattamisra ◽  
Chew Hui Kuean ◽  
Lee Boon Chieh ◽  
Vivian Lee Yean Yan ◽  
Chin Koh Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Helicobacter Pylori ◽  
Antibacterial Activity ◽  
Synergistic Effect ◽  
Inhibitory Concentration ◽  
Therapeutic Potential ◽  
E Coli ◽  
Checkerboard Assay ◽  
H Pylori

The antibacterial activity of geraniol and its effect in combination with ampicillin, amoxicillin and clarithromycin against Staphylococcus aureus, Escherichia coli and Helicobacter pylori was tested. The minimum inhibitory concentrations (MICs) and combinatory effects of geraniol against the bacteria were assessed by using the modified broth microdilution and checkerboard assay, respectively. The combinatory effect is expressed as fractional inhibitory concentration index (FICI). The MIC of geraniol against S. aureus, E. coli and H. pylori was found to be 11200, 5600, and 7325 μg/mL, respectively. A significant synergistic effect was observed with geraniol and ampicillin against S. aureus with FICI in the range 0.19 to 0.32. Geraniol and ampicillin exhibited a partial synergistic effect against E. coli. A similar effect was observed with geraniol and clarithromycin against S. aureus. A partial synergistic effect was observed with clarithromycin and geraniol against H. pylori with the FICI value in the range 0.86 to 0.89. An additive effect was observed with geraniol and amoxicillin combination against H. pylori. However, the amoxicillin and clarithromycin dose was reduced by thirty-two fold when combined with geraniol against H. pylori. The anti- H. pylori effect of geraniol with clarithromycin and amoxicillin could be of potential interest in the treatment of H. pylori infection and associated ulcers in humans. Further, geraniol, in combination with other antibiotics, has substantial therapeutic potential against S. aureus and E.coli infection.

scholarly journals Development of a quantitative PCR assay for detection of redside shiner (Richardsonius balteatus) from environmental DNA

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
Marshal S. Hoy ◽  
Carl O. Ostberg
Keyword(s):  
Quantitative Pcr ◽  
Water Samples ◽  
Effective Means ◽  
Environmental Dna ◽  
Sympatric Species ◽  
Negative Control ◽  
Control Site ◽  
Qpcr Assay ◽  
Redside Shiner ◽  
Richardsonius Balteatus

Abstract Objective A quantitative PCR (qPCR) assay for the detection of redside shiner (Richardsonius balteatus) environmental DNA (eDNA) was designed as a side product of a larger project aimed at using eDNA to determine the presence and geographic extent of native and non-native fishes in the reservoirs and associated tributaries above the three mainstem dams (Ross, Diablo, Gorge) on the Skagit River, Washington, USA. The eDNA survey results can be used to help guide additional sampling efforts that include traditional sampling methods, such as electrofishing and netting. Results The redside shiner qPCR assay (RSSCOI_540-601) was validated by testing for sensitivity using redside shiner genomic DNA from three different populations and by testing for specificity against 30 potentially sympatric species. No non-target amplification was observed in our validation tests. We then evaluated the assay on field-collected water samples where there are known populations of redside shiner and a negative control site where the target species is known to be absent. The field-collected water samples tested positive at the redside shiner sites and tested negative at the negative control site. The assay could provide resource managers with an effective means for surveying and monitoring redside shiner populations.

Application of a molecular method for the detection of group A rotaviruses in raw and treated water

2004 ◽  
Vol 50 (1) ◽  
pp. 223-228 ◽  
Author(s):  
W.B. van Zyl ◽  
P.J. Williams ◽  
W.O.K. Grabow ◽  
M.B. Taylor
Keyword(s):  
Drinking Water ◽  
Water Samples ◽  
Viral Gastroenteritis ◽  
Treated Water ◽  
Source Of Infection ◽  
Drinking Waters ◽  
Group A Rotaviruses ◽  
Group A ◽  
Treated Drinking Water ◽  
Drinking Water Samples

Group A human rotaviruses (HRVs) are the most important aetiological agents of acute viral gastroenteritis in infants and young children in both developing and industrialised countries. Rotaviruses are resistant to many chemical disinfectants and reportedly survive well in treated tapwater and sewage. In this study a group A specific reverse transcriptase-polymerase chain reaction (RT-PCR) followed by a nested-PCR was applied for the detection of HRVs in raw and treated drinking-water samples drawn at a water reclamation plant. For a period of two years (July 2000 to June 2002), borehole, raw and treated drinking-water samples were collected weekly. Viruses were recovered from the water samples using a glass wool adsorption-elution technique followed by secondary concentration using precipitation with polyethylene glycol. In the first year of the study group A HRVs were detected in 11% sewage samples, 8% partially treated waters and 5% final treated drinking waters. The results of the second year of the study showed the presence of group A HRVs in 11% sewage and untreated surface water samples, 15% partially treated water and 6.5% final treated drinking waters. No HRVs were detected in the water samples from the boreholes. The presence of group A HRVs in treated drinking-water samples suggested that this water could be a potential source of infection to consumers. The data also implied that either the water treatment did not remove HRVs or the treated water was contaminated post-treatment.

scholarly journals Borehole water: a potential health risk to rural communities in South Africa

2018 ◽  
Vol 19 (1) ◽  
pp. 128-136 ◽  
Author(s):  
S. Taonameso ◽  
L. S. Mudau ◽  
A. N. Traoré ◽  
N. Potgieter
Keyword(s):  
Drinking Water ◽  
Rural Communities ◽  
Water Samples ◽  
Rural Areas ◽  
Total Coliform ◽  
Water Sources ◽  
Potential Health ◽  
Point Mapping ◽  
E Coli ◽  
Mapping Tool

Abstract Sporadic outbreaks of diarrhoea in children in the Vhembe rural areas could be an indication of contamination in drinking water sources. In areas where improved water sources are used, not all rural households experience the benefits of these improved water sources. Water samples were collected from boreholes in three wards in the Vhembe District to determine microbiological risks over a 5-month period. A Water Point Mapping tool was used to indicate the borehole distribution. Water samples were taken from each functional borehole and analysed for total coliform and Escherichia coli counts, electrical conductivity, pH and temperature. A multiplex PCR protocol was used for identification of pathogenic E. coli. A total of 125 boreholes were identified of which only 12 were functional. Seven boreholes tested positive for total coliforms and E. coli counts. Four boreholes (33.3%) tested positive for diarrhoeagenic E. coli. Fifty-eight percent (58%) of water samples were without health risks, 17% were low risk and 25% could cause infection according to the South African water quality standards. This study indicated the importance of the role of the Municipalities and the maintenance plans that need to ensure that all boreholes are functional and provide safe drinking water to the rural communities.

Helicobacter pylori in water systems for human use in Mexico City

2001 ◽  
Vol 43 (12) ◽  
pp. 93-98 ◽  
Author(s):  
M. Mazari-Hiriart ◽  
Y. López-Vidal ◽  
J. J. Calva
Keyword(s):  
Helicobacter Pylori ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Mexico City ◽  
Water Samples ◽  
Water Systems ◽  
Treated Wastewater ◽  
Rrna Gene ◽  
H Pylori ◽  
System 1

Helicobacter pylori infection is associated with peptic ulcers and gastric cancer in humans. Transmission of H. pylori is still not certain with some epidemiological data suggesting water as a possible transmission route. The objective of this study was to detect H. pylori 16S rRNA gene in five water systems in the Mexico City area. Samples were taken between 1997 and 2000 from extraction wells (system 1), from dams used as water sources, both pre- and post-treatment (systems 2 and 3), treated wastewater (system 4) and non-treated wastewater (system 5). Detection of the H. pylori 16S rRNA gene in water samples was carried out using nested PCR in 139 water samples and confirmed by using cagA gene detection by PCR-hybridisation. The results showed the presence of H. pylori in 58 (42%) of the water samples in total with a prevalence of 68% in system 1, 100% in system 2, 0% in system 3, 17% in system 4 and 20% in system 5. This first stage showed the presence of H. pylori in the tested water systems; nevertheless, viability of the microorganism in water and vegetables needs to be confirmed as well as demonstration of a relationship between human and environmental strains.

Protection against Helicobacter pylori infection in mice by intragastric vaccination with H. pylori antigens is achieved using a non-toxic mutant of E. coli heat-labile enterotoxin (LT) as adjuvant

Vaccine ◽  
1998 ◽  
Vol 16 (1) ◽  
pp. 33-37 ◽  
Author(s):  
Marta Marchetti ◽  
Michela Rossi ◽  
Valentina Giannelli ◽  
Marzia M Giuliani ◽  
Mariagrazia Pizza ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Helicobacter Pylori Infection ◽  
E Coli ◽  
Heat Labile ◽  
H Pylori

scholarly journals Bacterial killing in gastric juice – effect of pH and pepsin on Escherichia coli and Helicobacter pylori

2006 ◽  
Vol 55 (9) ◽  
pp. 1265-1270 ◽  
Author(s):  
H. Zhu ◽  
C. A. Hart ◽  
D. Sales ◽  
N. B. Roberts
Keyword(s):  
Escherichia Coli ◽  
Helicobacter Pylori ◽  
Gastric Juice ◽  
Bacterial Killing ◽  
E Coli ◽  
Test Organisms ◽  
H Pylori ◽  
Ph Range ◽  
K 12 ◽  
Rough Mutant

The susceptibility of Escherichia coli and Helicobacter pylori to pH and the effect of pepsin-mediated proteolysis were investigated. This was to establish the relative importance of their bacterial killing properties in gastric juice. Solutions in the pH range 1.5–7.4 with or without pig pepsin A were used, together with seven gastric juice samples obtained from patients undergoing routine gastric collection. Escherichia coli C690 (a capsulate strain), E. coli K-12 (a rough mutant) and Helicobacter pylori E5 were selected as the test organisms. Suspensions of bacteria (1×106 E. coli ml−1 and 1×108 H. pylori ml−1) were pre-incubated with test solutions at 37 °C for up to 2 h, and then cultured to establish the effect on subsequent growth. Survival of bacteria was diminished at pHs of less than 3.5, whereas killing required a pH of less than 2.5. Pre-incubation with pig pepsin at 0.5, 1.0 and 2.0 mg ml−1 at pH 3.5 reduced viable counts by 100 % for E. coli 690 and E. coli K-12 after 100 min incubation. With H. pylori, the viable counts decreased to 50 % of the control after 20 min incubation in 1 mg pepsin ml−1 at pH 2.5, 3.0 and 3.5. The gastric juices showed bactericidal activity at pH 3.5, and the rate of killing was juice dependent, with complete death of E. coli 690 occurring between 5 and 40 min post-incubation. Thus, killing of E. coli and H. pylori occurs optimally at pHs of less than 2.5. At pH 3.5, little effect is observed, whereas addition of pepsin alone or in gastric juice causes a marked increase in bacterial susceptibility, suggesting an important role for proteolysis in the killing of bacteria.

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