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Sample records for qiaamp dna stool

  1. Comparison of QIAsymphony automated and QIAamp manual DNA extraction systems for measuring Epstein-Barr virus DNA load in whole blood using real-time PCR.

    Science.gov (United States)

    Laus, Stella; Kingsley, Lawrence A; Green, Michael; Wadowsky, Robert M

    2011-11-01

    Automated and manual extraction systems have been used with real-time PCR for quantification of Epstein-Barr virus [human herpesvirus 4 (HHV-4)] DNA in whole blood, but few studies have evaluated relative performances. In the present study, the automated QIAsymphony and manual QIAamp extraction systems (Qiagen, Valencia, CA) were assessed using paired aliquots derived from clinical whole-blood specimens and an in-house, real-time PCR assay. The detection limits using the QIAsymphony and QIAamp systems were similar (270 and 560 copies/mL, respectively). For samples estimated as having ≥10,000 copies/mL, the intrarun and interrun variations were significantly lower using QIAsymphony (10.0% and 6.8%, respectively), compared with QIAamp (18.6% and 15.2%, respectively); for samples having ≤1000 copies/mL, the two variations ranged from 27.9% to 43.9% and were not significantly different between the two systems. Among 68 paired clinical samples, 48 pairs yielded viral loads ≥1000 copies/mL under both extraction systems. Although the logarithmic linear correlation from these positive samples was high (r(2) = 0.957), the values obtained using QIAsymphony were on average 0.2 log copies/mL higher than those obtained using QIAamp. Thus, the QIAsymphony and QIAamp systems provide similar EBV DNA load values in whole blood. Copyright © 2011 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

  2. Comparison of QIAsymphony Automated and QIAamp Manual DNA Extraction Systems for Measuring Epstein-Barr Virus DNA Load in Whole Blood Using Real-Time PCR

    OpenAIRE

    Laus, Stella; Kingsley, Lawrence A.; Green, Michael; Wadowsky, Robert M.

    2011-01-01

    Automated and manual extraction systems have been used with real-time PCR for quantification of Epstein-Barr virus [human herpesvirus 4 (HHV-4)] DNA in whole blood, but few studies have evaluated relative performances. In the present study, the automated QIAsymphony and manual QIAamp extraction systems (Qiagen, Valencia, CA) were assessed using paired aliquots derived from clinical whole-blood specimens and an in-house, real-time PCR assay. The detection limits using the QIAsymphony and QIAam...

  3. Compatibility of DNA IQ™, QIAamp® DNA Investigator, and QIAsymphony® DNA Investigator® with various fingerprint treatments.

    Science.gov (United States)

    Lin, Sze-Wah; Ip, Stephen C Y; Lam, Tze-Tsun; Tan, Tung-Fai; Yeung, Wai-Lung; Tam, Wai-Ming

    2017-03-01

    Latent fingerprint and touch DNA are the two most important contact evidence for individualization in forensic science which provide complementary information that can lead to direct and unequivocal identification of the culprit. In order to retrieve useful information from both fingerprints and DNA, which are usually mingled together, one strategy is to perform fingerprint examination prior to DNA analysis since common DNA sampling technique such as swabbing could disturb or even destroy fingerprint details. Here, we describe the compatibility of three automatic DNA extraction systems, namely, DNA IQ™, QIAamp ® DNA Investigator, and QIAsymphony ® DNA Investigator ® , with respective to the effects of various fingerprint detection techniques. Our results demonstrate that Super Glue fingerprint treatment followed by DNA IQ™ extraction shows better effectiveness in DNA profiling. Aluminum powder dusting offers the least interference to the three DNA extraction systems above. Magnetic powder dusting, on the other hand, strongly impedes DNA recovery. Physical Developer is the most intrusive, which yields profiles with poor quality, including lower peak heights, poor peak height ratios, and poor intra-color balance. In terms of the choice of extraction method, DNA IQ™ system is recommended for sampling after fingerprint treatments, but not the two DNA Investigator systems.

  4. Evaluation of Patients with an Apparent False Positive Stool DNA Test: The Role of Repeat Stool DNA Testing.

    Science.gov (United States)

    Cooper, Gregory S; Markowitz, Sanford D; Chen, Zhengyi; Tuck, Missy; Willis, Joseph E; Berger, Barry M; Brenner, Dean E; Li, Li

    2018-03-07

    There is uncertainty as to the appropriate follow-up of patients who test positive on multimarker stool DNA (sDNA) testing and have a colonoscopy without neoplasia. To determine the prevalence of missed colonic or occult upper gastrointestinal neoplasia in patients with an apparent false positive sDNA. We prospectively identified 30 patients who tested positive with a commercially available sDNA followed by colonoscopy without neoplastic lesions. Patients were invited to undergo repeat sDNA at 11-29 months after the initial test followed by repeat colonoscopy and upper endoscopy. We determined the presence of neoplastic lesions on repeat evaluation stratified by results of repeat sDNA. Twelve patients were restudied. Seven patients had a negative second sDNA test and a normal second colonoscopy and upper endoscopy. In contrast, 5 of 12 subjects had a persistently positive second sDNA test, and 3 had positive findings, including a 3-cm sessile transverse colon adenoma with high-grade dysplasia, a 2-cm right colon sessile serrated adenoma with dysplasia, and a nonadvanced colon adenoma (p = 0.045). These corresponded to a positive predictive value of 0.60 (95% CI 0.17-1.00) and a negative predictive value of 1.00 (95% CI 1.00-1.00) for the second sDNA test. In addition, the medical records of all 30 subjects with apparent false positive testing were reviewed and no documented cases of malignant tumors were recorded. Repeat positive sDNA testing may identify a subset of patients with missed or occult colorectal neoplasia after negative colonoscopy for an initially positive sDNA. High-quality colonoscopy with careful attention to the right colon in patients with positive sDNA is critically important and may avoid false negative colonoscopy.

  5. Detection of Colorectal Cancer by a Quantitative Fluorescence Determination of DNA Amplification in Stool

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    Daniele Calistri

    2004-09-01

    Full Text Available DNA amplification of exfoliated cells in stool repre sents an inexpensive and rapid test, but has only 50% to 60% sensitivity. A new quantitative method, calle( fluorescence long DNA, was developed and validate( in our laboratory on stool obtained from 86 patient., with primary colorectal cancer and from 62 health individuals. It consists of the amplification of stoo DNA with fluorescence primers and the quantification of the amplification using a standard curve. Results are arbitrarily expressed in nanograms. The potential of thi new method compared to the conventional approact was analyzed in a subgroup of 94 individuals (51 patients and 38 healthy volunteers. In the presen series, DNA amplification analysis showed a specific ity of 97% and a sensitivity of only 50%. Conversely fluorescence DNA evaluation, using the best cutoff o 25 ng, showed a sensitivity of about 76% and a spec ificity of 93%. Similar sensitivity was observed regard less of Dukes stage, tumor location, and size, thu., also permitting the detection of early-stage tumors The present study seems to indicate that quantitative fluorescence DNA determination in stool successfully identifies colorectal cancer patients with a sensitivity comparable, if not superior, to that of multiple gene analysis but at a lower cost and in a shorter time.

  6. DNA extraction in Echinococcus granulosus and Taenia spp. eggs in dogs stool samples applying thermal shock.

    Science.gov (United States)

    Hidalgo, Alejandro; Melo, Angélica; Romero, Fernando; Hidalgo, Víctor; Villanueva, José; Fonseca-Salamanca, Flery

    2018-03-01

    The extraction of DNA in taeniid eggs shows complications attached to the composition of stool samples and the high resistance of eggs to degradation. The objective of this study was to test a method of DNA extraction in taeniid eggs by applying a thermal shock to facilitate the chemical-enzymatic degradation of these elements. A group of six tubes containing 1 ml of dog stool sample was spiked with eggs of Echinococcus granulosus and another group of six with Taenia pisiformis. Samples were floated with supersaturated sugar solution and centrifuged. The upper portion of each tube (500 μl) was aspirated and deposited in 1.5 ml tubes. Three tubes from each group were incubated at -20 °C and then at 90 °C, the remaining three from each group, incubated at room temperature. Proteinase K and lysis buffer were added to each tube and incubated for 12 h at 58 °C. The lysis effect was evaluated by microscopy at 3, 6 and 12 h and integrity by electrophoresis in 1% agarose gels. With the same experimental scheme, the thermal shock effect was evaluated in extractions of 1, 2, 3 and 4 eggs of each species and the DNA was quantified. Additionally, the protocol was applied in samples of 4 dogs diagnosed with natural infection by Taeniidae worms. Finally, all the extractions were tested by PCR amplification. Both E. granulosus and T. pisiformis eggs showed a similar response in the tests. In samples without treatment, the lysis effect was poor and showed no differences over time, but in those subjected to thermal shock, eggs degradation increased with time. In both treatments, there was no DNA loss integrity. The protocol applied to limited amounts of eggs yielded PCR products in 100% of the samples exposed to thermal shock, allowing PCR amplifications up to 1 egg. In non-exposed samples, the results were not replicable. However, DNA quantification showed low values in both treatments. In turn, DNA extractions with thermal shock in infected dog samples

  7. [On the use of FTA technology for collection, archieving, and molecular analysis of microsporidia dna from clinical stool samples].

    Science.gov (United States)

    Sokolova, O I; Dem'ianov, A V; Bovers, L S; Did'e, E S; Sokolova, Iu Ia

    2011-01-01

    The FTA technology was applied for sampling, archiving, and molecular analysis of the DNA isolated from stool samples to diagnose and identify microsporidia, the intracellular opportunistic parasites which induce malabsortion syndrome in immunosuppressed humans, particularly in patients with AIDS. Microsporidia DNA was successfully amplified in 6 of 50 stool samples of HIV-positive patients of the S. P. Botkin Memorial Infectious Disease Hospital (St. Petersburg) applied to FTA cards (FTA-Cars, Whatman Inc. Florham Park, NJ, USA). Amplicons (the fragments of rDNA) were directly sequenced, and microsporidia species--Encephalitozoon intestinalis, E. cuniculi, E. hellem, and Enterocytozoon bieneusi--were identified in Genbank by NCBI BLAST program. The FTA method of DNA immobilization is especially promising for epidemiological and field population studies which involve genotyping of microsporidia species and isolates.

  8. Microbial food safety: Potential of DNA extraction methods for use in diagnostic metagenomics

    DEFF Research Database (Denmark)

    Josefsen, Mathilde Hasseldam; Andersen, Sandra Christine; Christensen, Julia

    2015-01-01

    ) yielding protocols. The PowerLyzer PowerSoil DNA Isolation Kit performed significantly better than all other protocols tested. Selected protocols were modified, i.e., extended heating and homogenization, resulting in increased yields of total DNA. For QIAamp Fast DNA Stool Mini Kit (Qiagen) a 7-fold...... of the protocols to extract DNA was observed. The highest DNA yield was obtained with the PowerLyzer PowerSoil DNA Isolation Kit, whereas the FastDNA SPIN Kit for Feces (MP Biomedicals) resulted in the highest amount of PCR-amplifiable C. jejuni DNA....

  9. Detection of colorectal serrated polyps by stool DNA testing: comparison with fecal immunochemical testing for occult blood (FIT.

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    Russell I Heigh

    Full Text Available Precursors to 1/3 of colorectal cancer (CRC, serrated polyps have been under-detected by screening due to their inconspicuous, non-hemorrhagic, and proximal nature. A new multi-target stool DNA test (multi-target sDNA shows high sensitivity for both CRC and advanced adenomas. Screen detection of serrated polyps by this approach requires further validation. We sought to assess and compare noninvasive detection of sessile serrated polyps (SSP ≥ 1 cm by sDNA and an occult blood fecal immunochemical test (FIT.In a blinded prospective study, a single stool sample used for both tests was collected from 456 asymptomatic adults prior to screening or surveillance colonoscopy (criterion standard. All 29 patients with SSP ≥ 1 cm were included as cases and all 232 with no neoplastic findings as controls. Buffered stool samples were processed and frozen on receipt; Exact Sciences performed sDNA in batches using optimized analytical methods. The sDNA multi-marker panel targets methylated BMP3 (mBMP3 and NDRG4, mutant KRAS, β-actin, and hemoglobin. FIT (Polymedco OC-FIT Check was performed in separate lab ≤ 2 days post defecation and evaluated at cutoffs of 50 (FIT-50 and 100 ng/ml (FIT-100.MEDIAN AGES: cases 61 (range 57-77, controls 62 (52-70, p = NS. Women comprised 59% and 51%, p = NS, respectively. SSP median size was 1.2 cm (1-3 cm, 93% were proximal, and 64% had synchronous diminutive polyps. Among multi-target sDNA markers, mBMP3 proved highly discriminant for detection of SSP ≥ 1 cm (AUC = 0.87, p<0.00001; other DNA markers provided no incremental sensitivity. Hemoglobin alone showed no discrimination (AUC = 0.50, p = NS. At matched specificities, detection of SSP ≥ 1 cm by stool mBMP3 was significantly greater than by FIT-50 (66% vs 10%, p = 0.0003 or FIT-100 (63% vs 0%, p<0.0001.In a screening and surveillance setting, SSP ≥ 1 cm can be detected noninvasively by stool assay of exfoliated DNA markers, especially mBMP3. FIT appears to

  10. Digital quantification of gene methylation in stool DNA by emulsion-PCR coupled with hydrogel immobilized bead-array.

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    Liu, Yunlong; Wu, Haiping; Zhou, Qiang; Song, Qinxin; Rui, Jianzhong; Zou, Bingjie; Zhou, Guohua

    2017-06-15

    Aberrations of gene methylation in stool DNA (sDNA) is an effective biomarker for non-invasive colorectal cancer diagnosis. However, it is challenging to accurately quantitate the gene methylation levels in sDNA due to the low abundance and degradation of sDNA. In this study, a digital quantification strategy was proposed by combining emulsion PCR (emPCR) with hydrogel immobilized bead-array. The assay includes following steps: bisulfite conversion of sDNA, pre-amplification by PCR with specific primers containing 5' universal sequences, emPCR of pre-amplicons with beaded primers to achieve single-molecular amplification and identification of hydrogel embedding beads coated with amplicons. The sensitivity and the specificity of the method are high enough to pick up 0.05% methylated targets from unmethylated DNA background. The successful detection of hypermethylated vimentin gene in clinical stool samples suggests that the proposed method should be a potential tool for non-invasive colorectal cancer screening. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. The Stool DNA Test is More Accurate than the Plasma Septin 9 Test in Detecting Colorectal Neoplasia

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    Ahlquist, David A.; Taylor, William R.; Mahoney, Douglas W.; Zou, Hongzhi; Domanico, Michael; Thibodeau, Stephen N.; Boardman, Lisa A.; Berger, Barry M.; Lidgard, Graham P.

    2014-01-01

    Background & Aims Several noninvasive tests have been developed for colorectal cancer (CRC) screening. We compared the sensitivities of a multi-marker test for stool DNA (sDNA) and a plasma test for methylated Septin 9 (SEPT9) in identifying patients with large adenomas or CRC. Methods We analyzed paired stool and plasma samples from 30 patients with CRC and 22 with large adenomas from Mayo Clinic archives. Stool (n=46) and plasma (n=49) samples from age- and sex-matched patients with normal colonoscopy results were used as controls. The sDNA test is an assay for methylated BMP3, NDRG4, vimentin, and TFPI2; mutant KRAS; the β-actin gene, and quantity of hemoglobin (by the porphyrin method). It was performed blindly at Exact Sciences (Madison WI); the test for SEPT9 was performed at ARUP Laboratories (Salt Lake City UT). Results were considered positive based on the manufacturer's specificity cutoff values of 90% and 89%, respectively. Results The sDNA test detected adenomas (median 2 cm, range 1–5 cm) with 82% sensitivity (95% confidence interval [CI], 60%–95%); SEPT9 had 14% sensitivity (95% CI, 3%–35%; P=.0001). The sDNA test identified patients with CRC with 87% sensitivity (95% CI, 69%–96%); SEPT9 had 60% sensitivity (95% CI, 41%–77%; P=.046). The sDNA test identified patients with stage I–III CRC with 91% sensitivity (95% CI, 71%–99%); SEPT9 had 50% sensitivity (95% CI, 28%–72%; P=.013); for stage IV CRC, sensitivity values were 75% (95% CI, 35%–97%) and 88% (95% CI, 47%–100%), respectively (P=.56). False-positive rates were 7% for the sDNA test and 27% for SEPT9. Conclusions Based on analyses of paired samples, the sDNA test detects non-metastatic CRC and large adenomas with significantly greater levels of sensitivity than the SEPT9 test. These findings might be used to modify approaches for CRC prevention and early detection. PMID:22019796

  12. A novel nested multiplex polymerase chain reaction (PCR assay for differential detection of Entamoeba histolytica, E. moshkovskii and E. dispar DNA in stool samples

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    Parija Subhash C

    2007-05-01

    Full Text Available Abstract Background E. histolytica, a pathogenic amoeba, is indistinguishable in its cyst and trophozoite stages from those of non-pathogenic E. moshkovskii and E. dispar by light microscopy. We have developed a nested multiplex PCR targeting a 16S-like rRNA gene for differential detection of all the three morphologically similar forms of E. histolytica, E. moshkovskii and E. dispar simultaneously in stool samples. Results The species specific product size for E. histolytica, E. moshkovskii and E. dispar was 439, 553 and 174 bp respectively, which was clearly different for all the three Entamoeba species. The nested multiplex PCR showed a sensitivity of 94% and specificity of 100% for the demonstration of E. histolytica, E. moshkovskii and E. dispar DNA in stool samples. The PCR was positive for E. histolytica, E. moshkovskii and E. dispar in a total of 190 out of 202 stool specimens (94% sensitive that were positive for E. histolytica/E. dispar/E. moshkovskii by examination of stool by microscopy and/or culture. All the 35 negative control stool samples that were negative for E. histolytica/E. dispar/E. moshkovskii by microscopy and culture were also found negative by the nested multiplex PCR (100% specific. The result from the study shows that only 34.6% of the patient stool samples that were positive for E. histolytica/E. dispar/E. moshkovskii by examination of stool by microscopy and/or culture, were actually positive for pathogenic E. histolytica and the remaining majority of the stool samples were positive for non-pathogenic E. dispar or E. moshkovskii as demonstrated by the use of nested multiplex PCR. Conclusion The present study reports a new nested multiplex PCR strategy for species specific detection and differentiation of E. histolytica, E. dispar and E. moshkovskii DNA in stool specimens. The test is highly specific, sensitive and also rapid, providing the results within 12 hours of receiving stool specimens.

  13. Stool Culture

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    ... infections and may be identified with a stool culture. Some important examples include: Escherichia coli 0157:H7 and other toxin- ... the toxin-producing C. difficile will be performed. Examples of other less common causes include: ... of stool cultures that are reported as negative usually reflect the ...

  14. Stool Tests

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    ... clean, sealable container before taking to the laboratory. Plastic wrap can also be used to line the diaper of an infant or toddler who is not yet using the toilet. The stool should be collected into clean, dry plastic jars with screw-cap lids. You can get ...

  15. Diagnostic value of stool DNA testing for multiple markers of colorectal cancer and advanced adenoma: a meta-analysis.

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    Yang, Hua; Xia, Bing-Qing; Jiang, Bo; Wang, Guozhen; Yang, Yi-Peng; Chen, Hao; Li, Bing-Sheng; Xu, An-Gao; Huang, Yun-Bo; Wang, Xin-Ying

    2013-08-01

    The diagnostic value of stool DNA (sDNA) testing for colorectal neoplasms remains controversial. To compensate for the lack of large-scale unbiased population studies, a meta-analysis was performed to evaluate the diagnostic value of sDNA testing for multiple markers of colorectal cancer (CRC) and advanced adenoma. The PubMed, Science Direct, Biosis Review, Cochrane Library and Embase databases were systematically searched in January 2012 without time restriction. Meta-analysis was performed using a random-effects model using sensitivity, specificity, diagnostic OR (DOR), summary ROC curves, area under the curve (AUC), and 95% CIs as effect measures. Heterogeneity was measured using the χ(2) test and Q statistic; subgroup analysis was also conducted. A total of 20 studies comprising 5876 individuals were eligible. There was no heterogeneity for CRC, but adenoma and advanced adenoma harboured considerable heterogeneity influenced by risk classification and various detection markers. Stratification analysis according to risk classification showed that multiple markers had a high DOR for the high-risk subgroups of both CRC (sensitivity 0.759 [95% CI 0.711 to 0.804]; specificity 0.883 [95% CI 0.846 to 0.913]; AUC 0.906) and advanced adenoma (sensitivity 0.683 [95% CI 0.584 to 0.771]; specificity 0.918 [95% CI 0.866 to 0.954]; AUC 0.946) but not for the average-risk subgroups of either. In the methylation subgroup, sDNA testing had significantly higher DOR for CRC (sensitivity 0.753 [95% CI 0.685 to 0.812]; specificity 0.913 [95% CI 0.860 to 0.950]; AUC 0.918) and advanced adenoma (sensitivity 0.623 [95% CI 0.527 to 0.712]; specificity 0.926 [95% CI 0.882 to 0.958]; AUC 0.910) compared with the mutation subgroup. There was no significant heterogeneity among studies for subgroup analysis. sDNA testing for multiple markers had strong diagnostic significance for CRC and advanced adenoma in high-risk subjects. Methylation makers had more diagnostic value than mutation

  16. Comparison of DNA extraction methods for human gut microbial community profiling.

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    Lim, Mi Young; Song, Eun-Ji; Kim, Sang Ho; Lee, Jangwon; Nam, Young-Do

    2018-03-01

    The human gut harbors a vast range of microbes that have significant impact on health and disease. Therefore, gut microbiome profiling holds promise for use in early diagnosis and precision medicine development. Accurate profiling of the highly complex gut microbiome requires DNA extraction methods that provide sufficient coverage of the original community as well as adequate quality and quantity. We tested nine different DNA extraction methods using three commercial kits (TianLong Stool DNA/RNA Extraction Kit (TS), QIAamp DNA Stool Mini Kit (QS), and QIAamp PowerFecal DNA Kit (QP)) with or without additional bead-beating step using manual or automated methods and compared them in terms of DNA extraction ability from human fecal sample. All methods produced DNA in sufficient concentration and quality for use in sequencing, and the samples were clustered according to the DNA extraction method. Inclusion of bead-beating step especially resulted in higher degrees of microbial diversity and had the greatest effect on gut microbiome composition. Among the samples subjected to bead-beating method, TS kit samples were more similar to QP kit samples than QS kit samples. Our results emphasize the importance of mechanical disruption step for a more comprehensive profiling of the human gut microbiome. Copyright © 2017 The Authors. Published by Elsevier GmbH.. All rights reserved.

  17. Microbial diversity in fecal samples depends on DNA extraction method

    DEFF Research Database (Denmark)

    Mirsepasi, Hengameh; Persson, Søren; Struve, Carsten

    2014-01-01

    was to evaluate two different DNA extraction methods in order to choose the most efficient method for studying intestinal bacterial diversity using Denaturing Gradient Gel Electrophoresis (DGGE). FINDINGS: In this study, a semi-automatic DNA extraction system (easyMag®, BioMérieux, Marcy I'Etoile, France......BACKGROUND: There are challenges, when extracting bacterial DNA from specimens for molecular diagnostics, since fecal samples also contain DNA from human cells and many different substances derived from food, cell residues and medication that can inhibit downstream PCR. The purpose of the study...... by easyMag® from the same fecal samples. Furthermore, DNA extracts obtained using easyMag® seemed to contain inhibitory compounds, since in order to perform a successful PCR-analysis, the sample should be diluted at least 10 times. DGGE performed on PCR from DNA extracted by QIAamp DNA Stool Mini Kit DNA...

  18. Comparison of microscopy, ELISA, and real-time PCR for detection of Giardia intestinalis in human stool specimens

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    Beyhan, Yunus Emre; Taş Cengiz, Zeynep

    2017-08-23

    Background/aim: This study included patients who had digestive system complaints between August 2015 and October 2015. The research was designed to compare conventional microscopy with an antigen detection ELISA kit and the TaqMan-based real-time PCR (RT-PCR) technique for detection of Giardia intestinalis in human stool specimens. Materials and methods: Samples were concentrated by formalin-ether sedimentation technique and microscopic examinations were carried out on wet mount slides. A commercially available ELISA kit (Giardia CELISA, Cellabs, Brookvale, Australia) was used for immunoassay. DNA was extracted from fecal samples of about 200 mg using the QIAamp Fast DNA Stool Mini Kit (QIAGEN, Hilden, Germany) and the LightCycler Nano system (Roche Diagnostics, Mannheim, Germany) was used for the TaqMan-based RT-PCR assay. Results: A total of 94 stool samples, 38 of them diagnosed positive (40.4%) and 56 of them diagnosed negative by microscopy, were selected for evaluation by antigen detection and molecular assays. The prevalence of G. intestinalis infection was found as 46.8% (n: 44) and 79.8% (n: 75) by ELISA and RT-PCR, respectively. RT-PCR revealed by far the highest positivity rate compared to the other two methods. The difference between these methods was found to be statistically significant (P PCR, the sensitivity and specificity of microscopy and ELISA were 50.7% and 100% and 53.3% and 79%, respectively. Conclusion: RT-PCR seems to be much more sensitive and beneficial for rapid and accurate diagnosis of G. intestinalis in human stools.

  19. Validation of methylation-sensitive high-resolution melting (MS-HRM) for the detection of stool DNA methylation in colorectal neoplasms.

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    Xiao, Zhujun; Li, Bingsheng; Wang, Guozhen; Zhu, Weisi; Wang, Zhongqiu; Lin, Jinfeng; Xu, Angao; Wang, Xinying

    2014-04-20

    Methylation-sensitive high-resolution melting (MS-HRM) is a new technique for assaying DNA methylation, but its feasibility for assaying stool in patients with colorectal cancer (CRC) is unknown. First, the MS-HRM and methylation-specific PCR (MSP) detection limits were tested. Second, the methylation statuses of SFRP2 and VIM were analyzed in stool samples by MS-HRM, and in matching tumor and normal colon tissues via bisulfite sequencing PCR (BSP). Third, a case-control study evaluated the diagnostic sensitivity and specificity of MS-HRM relative to results obtained with MSP and the fecal immunochemical test (FIT). Finally, the linearity and reproducibility of MS-HRM were assessed. The detection limits of MS-HRM and MSP were 1% and 5%, respectively. The diagnostic sensitivities of MS-HRM (87.3%, 55/63) in stool and BSP in matching tumor tissue (92.1%, 58/63) were highly consistent (κ=0.744). The MS-HRM assay detected 92.5% (37/40) methylation in CRCs, 94.4% (34/36) in advanced adenomas, and 8.8% (5/57) in normal controls. The results of MS-HRM analysis were stable and reliable and showed fairly good linearity for both SFRP2 (PHRM shows potential for CRC screening. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Stool Gram stain

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    ... stool sample. The Gram stain method is sometimes used to quickly diagnose bacterial infections. How the Test is Performed You will need to collect a stool sample. There are many ways to collect the sample. You can catch the stool on plastic wrap that is loosely placed over the toilet bowl ...

  1. Stool C difficile toxin

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    ... toxin; Colitis - toxin; Pseudomembranous - toxin; Necrotizing colitis - toxin; C difficile - toxin ... be analyzed. There are several ways to detect C difficile toxin in the stool sample. Enzyme immunoassay ( ...

  2. Stool Color: When to Worry

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    Stool color: When to worry Yesterday, my stool color was bright green. Should I be concerned? Answers from Michael ... M.D. Stool comes in a range of colors. All shades of brown and even green are ...

  3. Bloody or tarry stools

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    ... small intestine Diverticulosis (abnormal pouches in the colon) Hemorrhoids (common cause of bright red blood) Inflammatory bowel ... have an exam even if you think that hemorrhoids are causing the blood in your stool. In ...

  4. Comparison of DNA extraction kits for detection of Burkholderia pseudomallei in spiked human whole blood using real-time PCR.

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    Nicole L Podnecky

    Full Text Available Burkholderia pseudomallei, the etiologic agent of melioidosis, is endemic in northern Australia and Southeast Asia and can cause severe septicemia that may lead to death in 20% to 50% of cases. Rapid detection of B. pseudomallei infection is crucial for timely treatment of septic patients. This study evaluated seven commercially available DNA extraction kits to determine the relative recovery of B. pseudomallei DNA from spiked EDTA-containing human whole blood. The evaluation included three manual kits: the QIAamp DNA Mini kit, the QIAamp DNA Blood Mini kit, and the High Pure PCR Template Preparation kit; and four automated systems: the MagNAPure LC using the DNA Isolation Kit I, the MagNAPure Compact using the Nucleic Acid Isolation Kit I, and the QIAcube using the QIAamp DNA Mini kit and the QIAamp DNA Blood Mini kit. Detection of B. pseudomallei DNA extracted by each kit was performed using the B. pseudomallei specific type III secretion real-time PCR (TTS1 assay. Crossing threshold (C T values were used to compare the limit of detection and reproducibility of each kit. This study also compared the DNA concentrations and DNA purity yielded for each kit. The following kits consistently yielded DNA that produced a detectable signal from blood spiked with 5.5×10(4 colony forming units per mL: the High Pure PCR Template Preparation, QIAamp DNA Mini, MagNA Pure Compact, and the QIAcube running the QIAamp DNA Mini and QIAamp DNA Blood Mini kits. The High Pure PCR Template Preparation kit yielded the lowest limit of detection with spiked blood, but when this kit was used with blood from patients with confirmed cases of melioidosis, the bacteria was not reliably detected indicating blood may not be an optimal specimen.

  5. Flushable reagent stool blood test

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    Stool occult blood test - flushable home test; Fecal occult blood test - flushable home test ... This test is performed at home with disposable pads. You can buy the pads at the drug store without ...

  6. Evaluating variation in human gut microbiota profiles due to DNA extraction method and inter-subject differences.

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    Wagner Mackenzie, Brett; Waite, David W; Taylor, Michael W

    2015-01-01

    The human gut contains dense and diverse microbial communities which have profound influences on human health. Gaining meaningful insights into these communities requires provision of high quality microbial nucleic acids from human fecal samples, as well as an understanding of the sources of variation and their impacts on the experimental model. We present here a systematic analysis of commonly used microbial DNA extraction methods, and identify significant sources of variation. Five extraction methods (Human Microbiome Project protocol, MoBio PowerSoil DNA Isolation Kit, QIAamp DNA Stool Mini Kit, ZR Fecal DNA MiniPrep, phenol:chloroform-based DNA isolation) were evaluated based on the following criteria: DNA yield, quality and integrity, and microbial community structure based on Illumina amplicon sequencing of the V4 region of bacterial and archaeal 16S rRNA genes. Our results indicate that the largest portion of variation within the model was attributed to differences between subjects (biological variation), with a smaller proportion of variation associated with DNA extraction method (technical variation) and intra-subject variation. A comprehensive understanding of the potential impact of technical variation on the human gut microbiota will help limit preventable bias, enabling more accurate diversity estimates.

  7. Evaluating variation in human gut microbiota profiles due to DNA extraction method and inter-subject differences

    Directory of Open Access Journals (Sweden)

    Brett eWagner Mackenzie

    2015-02-01

    Full Text Available The human gut contains dense and diverse microbial communities which have profound influences on human health. Gaining meaningful insights into these communities requires provision of high quality microbial nucleic acids from human fecal samples, as well as an understanding of the sources of variation and their impacts on the experimental model. We present here a systematic analysis of commonly used microbial DNA extraction methods, and identify significant sources of variation. Five extraction methods (Human Microbiome Project protocol, MoBio PowerSoil DNA Isolation Kit, QIAamp DNA Stool Mini Kit, ZR Fecal DNA MiniPrep, phenol:chloroform-based DNA isolation were evaluated based on the following criteria: DNA yield, quality and integrity, and microbial community structure based on Illumina amplicon sequencing of the V4 region of bacterial and archaeal 16S rRNA genes. Our results indicate that the largest portion of variation within the model was attributed to differences between subjects (biological variation, with a smaller proportion of variation associated with DNA extraction method (technical variation and intra-subject variation. A comprehensive understanding of the potential impact of technical variation on the human gut microbiota will help limit preventable bias, enabling more accurate diversity estimates.

  8. 21 CFR 868.6700 - Anesthesia stool.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Anesthesia stool. 868.6700 Section 868.6700 Food... DEVICES ANESTHESIOLOGY DEVICES Miscellaneous § 868.6700 Anesthesia stool. (a) Identification. An anesthesia stool is a device intended for use as a stool for the anesthesiologist in the operating room. (b...

  9. The effect of neutral and acidic oligosaccharides on stool viscosity, stool frequency and stool pH in preterm infants

    NARCIS (Netherlands)

    Westerbeek, E. A. M.; Hensgens, R. L.; Mihatsch, W. A.; Boehm, G.; Lafeber, H. N.; van Elburg, R. M.

    2011-01-01

    To determine the effect of neutral oligosaccharides [small-chain galacto-oligosaccharides/long-chain fructo-oligosaccharides (scGOS/lcFOS)] in combination with acidic oligosaccharides (pAOS) on stool viscosity, stool frequency and stool pH in preterm infants. In this explorative RCT, preterm infants

  10. Stools - pale or clay-colored

    Science.gov (United States)

    ... gov/ency/article/003129.htm Stools - pale or clay-colored To use the sharing features on this page, please enable JavaScript. Stools that are pale, clay, or putty-colored may be due to problems ...

  11. Evaluation of DNA extraction methods for the detection of Cytomegalovirus in dried blood spots

    Science.gov (United States)

    Koontz, D.; Baecher, K.; Amin, M.; Nikolova, S.; Gallagher, M.; Dollard, S.

    2015-01-01

    Background Dried blood spots (DBS) are collected universally from newborns and may be valuable for the diagnosis of congenital Cytomegalovirus (CMV) infection. The reported analytical sensitivity for DBS testing compared to urine or saliva varies greatly across CMV studies. The purpose of this study was to directly compare the performance of various DNA extraction methods for identification of CMV in DBS including those used most often in CMV studies. Study design Whatman® Grade 903 filter paper cards were spotted with blood samples from 25 organ transplant recipients who had confirmed CMV viremia. Six DNA extraction methods were compared for relative yield of viral and cellular DNA: 2 manual solution-based methods (Gentra Puregene, thermal shock), 2 manual silica column-based methods (QIAamp DNA Mini, QIAamp DNA Investigator), and 2 automated methods (M48 MagAttract Mini, QIAcube Investigator). DBS extractions were performed in triplicate followed by real-time quantitative PCR (qPCR). Results For extraction of both viral and cellular DNA, two methods (QIAamp DNA Investigator and thermal shock) consistently gave the highest yields, and two methods (M48 MagAttract Mini and QIAamp DNA Mini) consistently gave the lowest yields. There was an average 3-fold difference in DNA yield between the highest and lowest yield methods. Conclusion The choice of DNA extraction method is a major factor in the ability to detect low levels of CMV in DBS and can largely account for the wide range of DBS sensitivities reported in studies to date. PMID:25866346

  12. Bisulfite-Based DNA Methylation Analysis from Recent and Archived Formalin-Fixed, Paraffin Embedded Colorectal Tissue Samples.

    Science.gov (United States)

    Kalmár, Alexandra; Péterfia, Bálint; Hollósi, Péter; Wichmann, Barnabás; Bodor, András; Patai, Árpád V; Schöller, Andrea; Krenács, Tibor; Tulassay, Zsolt; Molnár, Béla

    2015-09-01

    We aimed to test the applicability of formalin-fixed and paraffin-embedded (FFPE) tissue samples for gene specific DNA methylation analysis after using two commercially available DNA isolation kits. Genomic DNA was isolated from 5 colorectal adenocarcinomas and 5 normal adjacent tissues from "recent", collected within 6 months, and "archived", collected more than 5 years ago, FFPE tissues using either High Pure FFPET DNA Isolation kit or QIAamp DNA FFPE Tissue kit. DNA methylation analysis of MAL, SFRP1 and SFRP2 genes, known to be hypermethylated in CRC, was performed using methylation-sensitive high resolution melting (MS-HRM) analysis and sequencing. QIAamp (Q) method resulted in slightly higher recovery in archived (HP: 1.22 ± 3.18 μg DNA; Q: 3.00 ± 4.04 μg DNA) and significantly (p < 0.05) higher recovery in recent samples compared to High Pure method (HP) (HP: 4.10 ± 2.91 μg DNA; Q: 11.51 ± 7.50 μg DNA). Both OD260/280 and OD260/230 ratios were lower, but still high in the High Pure isolated archived and recent samples compared to those isolated with QIAamp. Identical DNA methylation patterns were detected for all 3 genes tested by MS-HRM with both isolation kits in the recent group. However, despite of higher DNA recovery in QIAamp slightly more reproducible methylation results were obtained from High Pure isolated archived samples. Sequencing confirmed DNA hypermethylation in CRCs. In conclusion, reproducible DNA methylation patterns were obtained from recent samples using both isolation kits. However, long term storage may affect the reliability of the results leading to moderate differences between the efficiency of isolation kits.

  13. Undigested Pills in Stool Mimicking Parasitic Infection.

    Science.gov (United States)

    Mir, Fazia; Achakzai, Ilyas; Ibdah, Jamal A; Tahan, Veysel

    2017-01-01

    Background . Orally ingested medications now come in both immediate release and controlled release preparations. Controlled release preparations were developed by pharmaceutical companies to improve compliance and decrease frequency of pill ingestion. Case Report . A 67-year-old obese male patient presented to our clinic with focal abdominal pain that had been present 3 inches below umbilicus for the last three years. This pain was not associated with any trauma or recent heavy lifting. Upon presentation, the patient reported that for the last two months he started to notice pearly oval structures in his stool accompanying his chronic abdominal pain. This had coincided with initiation of his nifedipine pills for his hypertension. He reported seeing these undigested pills daily in his stool. Conclusion . The undigested pills may pose a cause of concern for both patients and physicians alike, as demonstrated in this case report, because they can mimic a parasitic infection. This can result in unnecessary extensive work-up. It is important to review the medication list for extended release formulations and note that the outer shell can be excreted whole in the stool.

  14. [Bristol Stool Chart: Prospective and monocentric study of "stools introspection" in healthy subjects].

    Science.gov (United States)

    Amarenco, G

    2014-09-01

    The Bristol Stool Chart (BSC) allows patients to identify their stool form using seven different images with accompanying written descriptors. Stool form was found to correlate better than stool frequency with whole-gut transit as measured by a radio-opaque marker study. This score is widely used in order to verify the presence of a constipation and to evaluate the therapeutic impact of various treatments. In our clinical practice, we was strongly surprised by the facility and the great precision of the patients to report their stool form, meaning that they usually and daily verify these stools. We wanted to precise the goals of a such attitude. Two questionnaires were proposed to healthy and voluntary subjects. Q1 was supposedly presented in order to verify the sensibility of a French version of BSC in a healthy population. Thus, Q1 precised the difficulties or not to understand pictures and written descriptors, asked about exhaustive analysis by means of BSC of stool form and bowel condition. All subjects with history of ano-rectal disorders or specific treatment for bowel dysfunction were excluded. After Q1 fulfilled, Q2 was proposed to the subjects. Q2 was designed to precise the goals of the patient when he look at his stool and the frequency of such an investigation. Finally a specific question concerning the subject opinion about this behavior in terms of bothersome, shame, or metaphysic interrogation. Eighty-five healthy subjects were recruited (42 female and 43 male). Mean age was 37.2 (sd = 15.7). Mean score of BCS was 2.07 (sd =1.05) (2.07 for female and 1.81 for male, P = 0.22). Number of categories of stool form was only 1 in 40%, 2 categories in 31%, 3 in 19%, 4 in 10%. Presence of a constipation defined by category 1 or 2 was found in 17% (23% in F, 12% in M, P = 0.075). Precision of BSC was noted as excellent in 68%, moderated in 18% and poor in 14%. BSC was considered as easy to use in 75%. Frequency of inspection of feces was systematic for 37%, 1

  15. Culturing Stool Specimens for Campylobacter spp., Pennsylvania, USA

    Science.gov (United States)

    M’ikanatha, Nkuchia M.; Dettinger, Lisa A.; Perry, Amanda; Rogers, Paul; Reynolds, Stanley M.

    2012-01-01

    In 2010, we surveyed 176 clinical laboratories in Pennsylvania regarding stool specimen testing practices for enteropathogens, including Campylobacter spp. Most (96.3%) routinely test for Campylobacter spp. In 17 (15.7%), a stool antigen test is the sole method for diagnosis. We recommend that laboratory practice guidelines for Campylobacter spp. testing be developed. PMID:22377086

  16. Review: Diagnostic accuracy of PCR-based detection tests for Helicobacter Pylori in stool samples.

    Science.gov (United States)

    Khadangi, Fatemeh; Yassi, Maryam; Kerachian, Mohammad Amin

    2017-12-01

    Although different methods have been established to detect Helicobacter pylori (H. pylori) infection, identifying infected patients is an ongoing challenge. The aim of this meta-analysis was to provide pooled diagnostic accuracy measures for stool PCR test in the diagnosis of H. pylori infection. In this study, a systematic review and meta-analysis were carried out on various sources, including MEDLINE, Web of Sciences, and the Cochrane Library from April 1, 1999, to May 1, 2016. This meta-analysis adheres to the guidelines provided by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses report (PRISMA Statement). The clinical value of DNA stool PCR test was based on the pooled false positive, false negative, true positive, and true negative of different genes. Twenty-six of 328 studies identified met the eligibility criteria. Stool PCR test had a performance of 71% (95% CI: 68-73) sensitivity, 96% (95% CI: 94-97) specificity, and 65.6 (95% CI: 30.2-142.5) diagnostic odds ratio (DOR) in diagnosis of H. pylori. The DOR of genes which showed the highest performance of stool PCR tests was as follows: 23S rRNA 152.5 (95% CI: 55.5-418.9), 16S rRNA 67.9 (95%CI: 6.4-714.3), and glmM 68.1 (95%CI: 20.1-231.7). The sensitivity and specificity of stool PCR test are relatively in the same spectrum of other diagnostic methods for the detection of H. pylori infection. In descending order of significance, the most diagnostic candidate genes using PCR detection were 23S rRNA, 16S rRNA, and glmM. PCR for 23S rRNA gene which has the highest performance could be applicable to detect H. pylori infection. © 2017 John Wiley & Sons Ltd.

  17. Development of the Brussels Infant and Toddler Stool Scale ('BITSS'): protocol of the study

    NARCIS (Netherlands)

    Vandenplas, Yvan; Szajewska, Hania; Benninga, Marc; Di Lorenzo, Carlo; Dupont, Christophe; Faure, Christophe; Miqdadi, Mohamed; Osatakul, Seksit; Ribes-Konickx, Carmen; Saps, Miguel; Shamir, Raanan; Staiano, Annamaria; Franckx, Johan; Green, Robin; Hegar, Badriul; Lemmens, Roel; Salvatore, Silvia; Vieira, Mario; Verghote, Marc; Xinias, Ioannis

    2017-01-01

    The Bristol Stool Form Scale (BSS) which consists of 7 photographs of different stool forms allows assessment of stool consistency (scale 1 for hard lumps to scale 7 for watery stools), in an objective manner in adults. The BSS is also sometimes used to characterise the stools of infants and young

  18. Real time PCR to detect the environmental faecal contamination by Echinococcus multilocularis from red fox stools.

    Science.gov (United States)

    Knapp, Jenny; Millon, Laurence; Mouzon, Lorane; Umhang, Gérald; Raoul, Francis; Ali, Zeinaba Said; Combes, Benoît; Comte, Sébastien; Gbaguidi-Haore, Houssein; Grenouillet, Frédéric; Giraudoux, Patrick

    2014-03-17

    The oncosphere stage of Echinococcus multilocularis in red fox stools can lead, after ingestion, to the development of alveolar echinococcosis in the intermediate hosts, commonly small mammals and occasionally humans. Monitoring animal infection and environmental contamination is a key issue in public health surveillance. We developed a quantitative real-time PCR technique (qPCR) to detect and quantify E. multilocularis DNA released in fox faeces. A qPCR technique using a hydrolysis probe targeting part of the mitochondrial gene rrnL was assessed on (i) a reference collection of stools from 57 necropsied foxes simultaneously investigated using the segmental sedimentation and counting technique (SSCT) (29 positive for E. multilocularis worms and 28 negative animals for the parasite); (ii) a collection of 114 fox stools sampled in the field: two sets of 50 samples from contrasted endemic regions in France and 14 from an E. multilocularis-free area (Greenland). Of the negative SSCT controls, 26/28 were qPCR-negative and two were weakly positive. Of the positive SSCT foxes, 25/29 samples were found to be positive by qPCR. Of the field samples, qPCR was positive in 21/50 (42%) and 5/48 (10.4%) stools (2 samples inhibited), originating respectively from high and low endemic areas. In faeces, averages of 0.1 pg/μl of DNA in the Jura area and 0.7 pg/μl in the Saône-et-Loire area were detected. All qPCR-positive samples were confirmed by sequencing. The qPCR technique developed here allowed us to quantify environmental E. multilocularis contamination by fox faeces by studying the infectious agent directly. No previous study had performed this test in a one-step reaction. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Optimal screening and donor management in a public stool bank.

    Science.gov (United States)

    Kazerouni, Abbas; Burgess, James; Burns, Laura J; Wein, Lawrence M

    2015-12-17

    Fecal microbiota transplantation is an effective treatment for recurrent Clostridium difficile infection and is being investigated as a treatment for other microbiota-associated diseases. To facilitate these activities, an international public stool bank has been created, which screens donors and processes stools in a standardized manner. The goal of this research is to use mathematical modeling and analysis to optimize screening and donor management at the stool bank. Compared to the current policy of screening active donors every 60 days before releasing their quarantined stools for sale, costs can be reduced by 10.3 % by increasing the screening frequency to every 36 days. In addition, the stool production rate varies widely across donors, and using donor-specific screening, where higher producers are screened more frequently, also reduces costs, as does introducing an interim (i.e., between consecutive regular tests) stool test for just rotavirus and C. difficile. We also derive a donor release (i.e., into the system) policy that allows the supply to approximately match an exponentially increasing deterministic demand. More frequent screening, interim screening for rotavirus and C. difficile, and donor-specific screening, where higher stool producers are screened more frequently, are all cost-reducing measures. If screening costs decrease in the future (e.g., as a result of bringing screening in house), a bottleneck for implementing some of these recommendations may be the reluctance of donors to undergo serum screening more frequently than monthly.

  20. Assessment of Commonly Used Pediatric Stool Scales: A pilot study

    OpenAIRE

    Saps, M.; Nichols-Vinueza, D.; Dhroove, G.; Adams, P.; Chogle, A.

    2013-01-01

    Background: The Bristol Stool Form Scale (BSFS) and a modified child-friendly version (M-BSFS) are frequently used in clinical practice and research. These scales have not been validated in children. 3-D stool scale models may be better adapted to the child's development. Aims: To assess the usefulness of the BSFS, M-BSFS, and a newly developed 3-D stool scale in children. Methods: Fifty children were asked to rank the picture cards of the BSFS and 3-D models from hardest to softest and...

  1. Development of a polymerase chain reaction applicable to rapid and sensitive detection of Clonorchis sinensis eggs in human stool samples

    Science.gov (United States)

    Cho, Pyo Yun; Na, Byoung-Kuk; Mi Choi, Kyung; Kim, Jin Su; Cho, Shin-Hyeong; Lee, Won-Ja; Lim, Sung-Bin; Cha, Seok Ho; Park, Yun-Kyu; Pak, Jhang Ho; Lee, Hyeong-Woo; Hong, Sung-Jong; Kim, Tong-Soo

    2013-01-01

    Microscopic examination of eggs of parasitic helminths in stool samples has been the most widely used classical diagnostic method for infections, but tiny and low numbers of eggs in stool samples often hamper diagnosis of helminthic infections with classical microscopic examination. Moreover, it is also difficult to differentiate parasite eggs by the classical method, if they have similar morphological characteristics. In this study, we developed a rapid and sensitive polymerase chain reaction (PCR)-based molecular diagnostic method for detection of Clonorchis sinensis eggs in stool samples. Nine primers were designed based on the long-terminal repeat (LTR) of C. sinensis retrotransposon1 (CsRn1) gene, and seven PCR primer sets were paired. Polymerase chain reaction with each primer pair produced specific amplicons for C. sinensis, but not for other trematodes including Metagonimus yokogawai and Paragonimus westermani. Particularly, three primer sets were able to detect 10 C. sinensis eggs and were applicable to amplify specific amplicons from DNA samples purified from stool of C. sinensis-infected patients. This PCR method could be useful for diagnosis of C. sinensis infections in human stool samples with a high level of specificity and sensitivity. PMID:23916334

  2. Rapid quantification of semen hepatitis B virus DNA by real-time polymerase chain reaction

    Science.gov (United States)

    Qian, Wei-Ping; Tan, Yue-Qiu; Chen, Ying; Peng, Ying; Li, Zhi; Lu, Guang-Xiu; Lin, Marie C.; Kung, Hsiang-Fu; He, Ming-Ling; Shing, Li-Ka

    2005-01-01

    AIM: To examine the sensitivity and accuracy of real-time polymerase chain reaction (PCR) for the quantification of hepatitis B virus (HBV) DNA in semen. METHODS: Hepatitis B viral DNA was isolated from HBV carriers’ semen and sera using phenol extraction method and QIAamp DNA blood mini kit (Qiagen, Germany). HBV DNA was detected by conventional PCR and quantified by TaqMan technology-based real-time PCR (quantitative polymerase chain reaction (qPCR)). The detection threshold was 200 copies of HBV DNA for conventional PCR and 10 copies of HBV DNA for real time PCR per reaction. RESULTS: Both methods of phenol extraction and QIAamp DNA blood mini kit were suitable for isolating HBV DNA from semen. The value of the detection thresholds was 500 copies of HBV DNA per mL in the semen. The viral loads were 7.5 × 107 and 1.67 × 107 copies of HBV DNA per mL in two HBV infected patients’ sera, while 2.14 × 105 and 3.02 × 105 copies of HBV DNA per mL in the semen. CONCLUSION: Real-time PCR is a more sensitive and accurate method to detect and quantify HBV DNA in the semen. PMID:16149152

  3. Prevalence of Helicobacter pylori in children by noninvasive stool ...

    African Journals Online (AJOL)

    Prevalence of Helicobacter pylori in children by noninvasive stool Antigen Enzyme Immunoassay. Augustine O. Ebonyi, Emeka Ejeliogu, Stanley T. Odigbo, Martha Omoo Ochoga, Stephen Oguche, Anejo-Okopi A. Joseph ...

  4. Assessment of commonly used pediatric stool scales: a pilot study.

    Science.gov (United States)

    Saps, M; Nichols-Vinueza, D; Dhroove, G; Adams, P; Chogle, A

    2013-01-01

    The Bristol Stool Form Scale (BSFS) and a modified child-friendly version (M-BSFS) are frequently used in clinical practice and research. These scales have not been validated in children. 3-D stool scale models may be better adapted to the child's development. To assess the usefulness of the BSFS, M-BSFS, and a newly developed 3-D stool scale in children. Fifty children were asked to rank the picture cards of the BSFS and 3-D models from hardest to softest and to match the pictures with descriptors for each stool type. Thirty percent of the children appropriately characterized the stools as hard, loose, or normal using the BSFS vs. 36.6% with the 3-D model (p=0.27). Appropriate correlation of stools as hard, loose, or normal consistency using the BSFS vs. the 3-D model by age group was: 6 to 11-year-olds, 27.5% vs. 33.3% (p=0.58) and 12 to 17-year-olds, 32.1% vs. 39.5% (p=0.41). Thirty-three percent correlated the BSFS pictures with the correct BSFS words, 46% appropriately correlated with the M-BSFS words, and 46% correlated the 3-D stool models with the correct wording. The BSFS and M-BSFS that are widely used as stool assessment instruments are not user-friendly for children. The 3-D model was not found to be better than the BSFS and the M-BSFS. Copyright © 2013 Asociación Mexicana de Gastroenterología. Published by Masson Doyma México S.A. All rights reserved.

  5. Rapid diagnosis of diarrhea caused by Shigella sonnei using dipsticks; comparison of rectal swabs, direct stool and stool culture.

    Directory of Open Access Journals (Sweden)

    Claudia Duran

    Full Text Available BACKGROUND: We evaluated a dipstick test for rapid detection of Shigella sonnei on bacterial colonies, directly on stools and from rectal swabs because in actual field situations, most pathologic specimens for diagnosis correspond to stool samples or rectal swabs. METHODOLOGY/PRINCIPAL FINDINGS: The test is based on the detection of S. sonnei lipopolysaccharide (LPS O-side chains using phase I-specific monoclonal antibodies coupled to gold particles, and displayed on a one-step immunochromatographic dipstick. A concentration as low as 5 ng/ml of LPS was detected in distilled water and in reconstituted stools in 6 minutes. This is the optimal time for lecture to avoid errors of interpretation. In distilled water and in reconstituted stools, an unequivocal positive reaction was obtained with 4 x 10(6 CFU/ml of S. sonnei. The specificity was 100% when tested with a battery of Shigella and different unrelated strains. When tested on 342 rectal swabs in Chile, specificity (281/295 was 95.3% (95% CI: 92.9% - 97.7% and sensitivity (47/47 was 100%. Stool cultures and the immunochromatographic test showed concordant results in 95.5 % of cases (328/342 in comparative studies. Positive and negative predictive values were 77% (95% CI: 65% - 86.5% and 100% respectively. When tested on 219 stools in Chile, Vietnam, India and France, specificity (190/198 was 96% (95% CI 92%-98% and sensitivity (21/21 was 100%. Stool cultures and the immunochromatographic test showed concordant results in 96.3 % of cases (211/219 in comparative studies. Positive and negative predictive values were 72.4% (95% CI 56.1%-88.6% and 100 %, respectively. CONCLUSION: This one-step dipstick test performed well for diagnosis of S. sonnei both on stools and on rectal swabs. These data confirm a preliminary study done in Chile.

  6. Recovery of DNA and fingerprints from touched documents.

    Science.gov (United States)

    Sewell, Jonathan; Quinones, Ignacio; Ames, Carole; Multaney, Bryan; Curtis, Stuart; Seeboruth, Haj; Moore, Stephen; Daniel, Barbara

    2008-09-01

    This study investigated the various factors affecting DNA profiling from DNA recovered from fingerprints deposited on paper before and after fingerprint enhancement treatments. The DNeasy plant mini kit (QIAGEN) was found to improve DNA recovery from paper by over 150% compared with the QIAamp mini kit. A significant decrease in the amount of DNA recovered was observed following treatment with DFO and/or Ninhydrin. This decrease in yield did not have a comparably significant effect on the quality of the SGM Plus profiles. Furthermore, this study found that whilst certain paper types, such as newspaper, magazine and filter paper allowed for the good recovery of DNA, common office paper and white card, strongly interfered with the recovery of DNA resulting in poor quality profiles.

  7. A simple and cost-effective method of DNA extraction from small formalin-fixed paraffin-embedded tissue for molecular oncologic testing.

    Science.gov (United States)

    Snow, Anthony N; Stence, Aaron A; Pruessner, Jonathan A; Bossler, Aaron D; Ma, Deqin

    2014-01-01

    Extraction of DNA from formalin-fixed, paraffin-embedded (FFPE) tissue is a critical step in molecular oncologic testing. As molecular oncology testing becomes more important for prognostic and therapeutic decision making and tissue specimens become smaller due to earlier detection of suspicious lesions and the use of fine needle aspiration methods for tissue collection, it becomes more challenging for the typical molecular pathology laboratory to obtain reliable test results. We developed a DNA extraction method to obtain sufficient quantity and high quality genomic DNA from limited FFPE tissue for molecular oncology testing using a combination of H&E stained slides, a matrix capture method and the Qiagen DNA column. THREE DNA EXTRACTION METHODS WERE COMPARED: our standard procedure of manually scraping tissue from unstained slides followed by DNA extraction using the QIAamp FFPE column (Qiagen, Valencia, CA), a glue capture method (Pinpoint Solution, Zymo Research Corp, Inc) on H&E stained slides followed by DNA extraction using either the QIAamp column or the column included with the Pinpoint kit (Zymo Research). The DNA extraction protocol was optimized. Statistical analysis was performed using the paired two-sample student's t-test. The combination of the matrix capture method with the QIAamp column gave an equivalent amount of DNA as our standard extraction method using the unstained slides and a 4.6-fold higher DNA yield than using the Zymo column included in the Pinpoint Slide Solution kit. Several molecular tests were performed and DNA purified using the new method gave the same results as for the previous methods. Using H&E stained slides allows visual confirmation of tumor cells during microdissection. The Pinpoint solution made removal of specific tissue from the slides easier and reduced the risk of contamination and tissue loss. This DNA extraction method is simple, cost-effective, and blends with our current workflow requiring no additional equipment.

  8. Identification and antibiotic sensitivity test of bacteria from stools of ...

    African Journals Online (AJOL)

    One hundred and fifty stool samples from 65 female and 85 male patients with acute diarrhoea from the Central Hospital, Agbor (Nigeria) were examined to ascertain the likelihood of cholera outbreak in Agbor. The samples were preserved in Carey-Blair semi-solid medium, inoculated directly on blood agar, McConkey agar ...

  9. Improved amplification efficiency on stool samples by addition of spermidine and its use for non-invasive detection of colorectal cancer

    KAUST Repository

    Roperch, Jean-Pierre

    2015-05-29

    Background Using quantitative methylation-specific PCR (QM-MSP) is a promising method for colorectal cancer (CRC) diagnosis from stool samples. Difficulty in eliminating PCR inhibitors of this body fluid has been extensively reported. Here, spermidine is presented as PCR facilitator for the detection of stool DNA methylation biomarkers using QM-MSP. We examined its effectiveness with NPY, PENK and WIF1, three biomarkers which we have previously shown to be of relevance to CRC. Results We determined an optimal window for the amplification of the albumin (Alb) gene (100 ng of bisulfite-treated stool DNA added of 1 mM spermidine) at which we report that spermidine acts as a PCR facilitator (AE = 1680%) for SG RT-PCR. We show that the amplification of methylated PENK, NPY and WIF1 is considerably facilitated by QM-MSP as measured by an increase of CMI (Cumulative Methylation Index, i.e. the sum of the three methylation values) by a factor of 1.5 to 23 fold in individual samples, and of 10 fold in a pool of five samples. Conclusions We contend that spermidine greatly reduces the problems of PCR inhibition in stool samples. This observed feature, after validation on a larger sampling, could be used in the development of stool-based CRC diagnosis tests.

  10. A Novel Stool PCR Test for Helicobacter pylori May Predict Clarithromycin Resistance and Eradication of Infection at a High Rate.

    Science.gov (United States)

    Beckman, Erin; Saracino, Ilaria; Fiorini, Giulia; Clark, Courtney; Slepnev, Vladimir; Patel, Denise; Gomez, Clarissa; Ponaka, Reddy; Elagin, Vecheslav; Vaira, Dino

    2017-08-01

    Clarithromycin-based regimens are commonly used as a first-line therapy for Helicobacter pylori -positive patients; however, resistance to clarithromycin has led to treatment failures. The aim of this study was to evaluate the feasibility of using stool samples to detect the presence of H. pylori DNA while concurrently detecting mutations associated with resistance to clarithromycin. For this purpose, total DNA was extracted from 294 raw stool specimens from H. pylori -positive and -negative patients. TaqMan real-time PCR amplification was used to detect the presence of H. pylori as well as to predict the phenotype of the organism and the related outcome for patients treated with clarithromycin. Clarithromycin resistance was determined upon analysis of the PCR result. Patients were also tested by a urea breath test and were subjected to esophagogastroduodenoscopy, followed by histology, culture, and a rapid urease test, in order to obtain a consensus patient infection status. Of 294 total stool samples, 227 were deemed true positive. The sensitivity of H. pylori detection by PCR was 93.8%. Of 213 true-positive samples that were sequenced, 36.2% showed point mutations associated with clarithromycin resistance (A2142C, A2142G, A2143G). The final correlation of the mutant genotypes as determined by sequencing with the eradication of infection was 86%. We found that Helicobacter pylori DNA can be detected in human stool specimens with high sensitivity and can therefore be used to determine the presence of the bacterium without obtaining a biopsy sample. Moreover, genotypic resistance to clarithromycin can be predicted without obtaining a biopsy sample, facilitating the choice of the right therapeutic approach. Copyright © 2017 American Society for Microbiology.

  11. Detection of Cryptosporidium sp infection by PCR and modified acid fast staining from potassium dichromate preserved stool

    Directory of Open Access Journals (Sweden)

    Agnes Kurniawan

    2009-09-01

    Full Text Available Aim To identify the frequency of Cryptosporidium infection in children below 3 years old by examining concentrated long term preserved stool using PCR detection of 18S rRNA gene and compared with modified acid fast staining technique.Methods Hundred eighty eight stools from children ≤ 3 years old were stored for 13 months in 2.5% K2Cr2O7 solution at 40C. Cryptosporidium oocysts were isolated by water-ether concentration technique. The concentrates were smeared onto object glass and stained with modified acid fast staining, and the rest of the concentrates were DNA extracted by freezing and thawing cycles and proteinase K digestion, then direct PCR was done to detect 18S rRNA gene.Result The proportion of positive stools for Cryptosporidium sp by acid fast staining from concentrated stools and 18S rRNA PCR were 4.8% and 34.6% respectively, which showed statistically significant difference.Conclusion The frequency of Cryptosporidium infection among children ≤ 3 years old was very high and stool storage in K2Cr2O7 for 13 months did not affect the PCR result. High prevalence of Cryptosporidium infection indicated high transmission in that area and the potential to be transmitted to other individuals such as the immunocompromised. (Med J Indones 2009;18:147-52Key words: 18S rRNA, cryptosporidiosis

  12. DNA extraction and barcode identification of development stages of forensically important flies in the Czech Republic.

    Science.gov (United States)

    Olekšáková, Tereza; Žurovcová, Martina; Klimešová, Vanda; Barták, Miroslav; Šuláková, Hana

    2018-04-01

    Several methods of DNA extraction, coupled with 'DNA barcoding' species identification, were compared using specimens from early developmental stages of forensically important flies from the Calliphoridae and Sarcophagidae families. DNA was extracted at three immature stages - eggs, the first instar larvae, and empty pupal cases (puparia) - using four different extraction methods, namely, one simple 'homemade' extraction buffer protocol and three commercial kits. The extraction conditions, including the amount of proteinase K and incubation times, were optimized. The simple extraction buffer method was successful for half of the eggs and for the first instar larval samples. The DNA Lego Kit and DEP-25 DNA Extraction Kit were useful for DNA extractions from the first instar larvae samples, and the DNA Lego Kit was also successful regarding the extraction from eggs. The QIAamp DNA mini kit was the most effective; the extraction was successful with regard to all sample types - eggs, larvae, and pupari.

  13. Evaluation and In-House Validation of Five DNA Extraction Methods for PCR-based STR Analysis of Bloodstained Denims

    Directory of Open Access Journals (Sweden)

    Henry Perdigon

    2004-06-01

    Full Text Available One type of crime scene evidence commonly submitted for analysis is bloodstain on denim. However, chemicals (e.g., indigo used to produce denim materials may co-purify with DNA and hence, affect subsequent DNA analysis. The present study compared five methods (e.g., standard organic, organic with hydrogen peroxide (H2O2, modified FTA™, organic/Chelex®-Centricon®, and QIAamp® DNA Mini Kit-based procedures for the isolation of blood DNA from denim. A Short Tandem Repeat (STR-based analysis across two to nine STR markers, namely, HUMvWA, HUMTH01, D8S306, HUMFES/FPS, HUMDHFRP2, HUMF13A01, HUMFGA, HUMTPOX, and HUMCSF1PO, was used to evaluate successful amplification of blood DNA extracted from light indigo, dark indigo, indigo-sulfur, pure indigo, sulfur-top, and sulfur-bottom denim materials. The results of the present study support the utility of organic/Chelex®-Centricon® and QIAamp® Kit procedures in extracting PCR-amplifiable DNA from five different types of denim materials for STR analysis. Furthermore, a solid-based method using FTA™ classic cards was modified to provide a simple, rapid, safe, and cost-effective procedure for extracting blood DNA from light, dark indigo and pure indigo denim materials. However, DNA eluted from bloodstained sulfur-dyed denims (e.g., sulfur-top and sulfur-bottom using FTA™ procedure was not readily amplifiable.

  14. Management of stool land revenue in Ghana: A study of the Nkawie ...

    African Journals Online (AJOL)

    Journal of Science and Technology (Ghana) ... of stool lands, poor record keeping which often results in multiple sales and chieftaincy disputes ... Given the constitutional importance of stool lands, this research investigates the impact of stool ... To this end, the research assessed the performance of key stakeholders like the

  15. Moellerella wisconsensis, a new genus and species of Enterobacteriaceae found in human stool specimens.

    Science.gov (United States)

    Hickman-Brenner, F W; Huntley-Carter, G P; Saitoh, Y; Steigerwalt, A G; Farmer, J J; Brenner, D J

    1984-04-01

    The name Moellerella wisconsensis is proposed for a group of the family Enterobacteriaceae previously called enteric group 46. The species name, wisconsensis, was coined because six of the nine strains were isolated in Wisconsin. M. wisconsensis strains were negative for indole production, Voges-Proskauer, H2S production, urea, phenylalanine deaminase, lysine and ornithine decarboxylases, arginine dihydrolase, gas production from D-glucose, acid production from trehalose, and motility; the strains were positive for methyl red, citrate (Simmons), and acid production from lactose and raffinose and resistant to colistin. DNAs from five strains of M. wisconsensis were highly related (80 to 93% in reactions assayed on hydroxyapatite at 60 degrees C and 78 to 97% at 75 degrees C) to 32P-labeled DNA of the proposed type strain (CDC 2896-78, ATCC 35017). Labeled DNA from this type strain was only 2 to 32% related (at 60 degrees C) to DNA from 49 strains of named and unnamed species of Enterobacteriaceae. Eight of nine M. wisconsensis strains were isolated from human stool samples. Clinical information on one strain was available, and it was found to be associated with a case of diarrhea. On MacConkey agar, colonies of M. wisconsensis were bright red with precipitated bile around them and thus were indistinguishable from Escherichia coli colonies. Future studies should focus on the isolation of this new organism and its relationship to human disease.

  16. Selection of the most suitable method for the extraction of DNA from foods and feeds for species identification

    Directory of Open Access Journals (Sweden)

    Michaela Nesvadbová

    2010-01-01

    Full Text Available High quality and purity of DNA isolated from food and feed is essential for species identification and has unpredictable influences an effect of analysis. In this study, the efficiency of eight different methods for DNA isolation was investigated. For DNA extraction, the raw chicken meet, ham, sausages, tinned lunch meat, pate, tinned feeds for dogs, complete granulated feeds for dogs and chicken flour were used. Kits of several different producers, i.e.: NucleoSpin Food (Marchery-Nagel, Wizard Genomic DNA Purification Kit (Promega, Invisorb Spin Food Kit I (Invitek, Wizard SV Genomic DNA Purification System (Promega, JetQuick Tissue DNA Spin Kit (Genomed, RNA Blue (Top-Bio, JetQuick Blood & Cell Culture Kit (Genomed, QIAamp DNA Mini Kit and QIAamp DNA Blood Mini Kit (Qiagen were employed in the study. Gel agarose electrophoresis for primary verification of DNA quality was performed. The isolates were subsequently assessed for quantity and quality using by spectrophotometer Nanodrop 2000 (Thermo Scientific. To verify of template usability and quality of isolated DNA, the polymerase chain reaction (PCR was used.Differences between isolated DNA from tinned products and meat, ham, sausage, granulated dog feed and chicken flour were found. In tinned food and feed, the DNA was more degraded, DNA content and DNA purity was lower and also PCR amplification was the most difficult. Overall DNA yield and quality have important influence on PCR products amplification. The best results were obtained with NucleoSpin Food and JetQuick Tissue DNA Spin Kit. DNA extracted by these methods proved highest yields, purity and template quality in all foods and feeds and the results of PCR analysis are excellent reproducible. Analyses showed that results depended on different food or feed using and dif­fe­rent isolation system.The results of this work will be utilized to choose the suitable isolating kit for educational course, which is designed for students and

  17. Disposal of children's stools and its association with childhood diarrhea in India.

    Science.gov (United States)

    Bawankule, Rahul; Singh, Abhishek; Kumar, Kaushalendra; Pedgaonkar, Sarang

    2017-01-05

    Children's stool disposal is often overlooked in sanitation programs of any country. Unsafe disposal of children's stool makes children susceptible to many diseases that transmit through faecal-oral route. Therefore, the study aims to examine the magnitude of unsafe disposal of children's stools in India, the factors associated with it and finally its association with childhood diarrhea. Data from the third round of the National Family Health Survey (NFHS-3) conducted in 2005-06 is used to carry out the analysis. The binary logistic regression model is used to examine the factors associated with unsafe disposal of children's stool. Binary logistic regression is also used to examine the association between unsafe disposal of children's stool and childhood diarrhea. Overall, stools of 79% of children in India were disposed of unsafely. The urban-rural gap in the unsafe disposal of children's stool was wide. Mother's illiteracy and lack of exposure to media, the age of the child, religion and caste/tribe of the household head, wealth index, access to toilet facility and urban-rural residence were statistically associated with unsafe disposal of stool. The odds of diarrhea in children whose stools were disposed of unsafely was estimated to be 11% higher (95% CI: 1.01-1.21) than that of children whose stools were disposed of safely. An increase in the unsafe disposal of children's stool in the community also increased the risk of diarrhea in children. We found significant statistical association between children's stool disposal and diarrhea. Therefore, gains in reduction of childhood diarrhea can be achieved in India through the complete elimination of unsafe disposal of children's stools. The sanitation programmes currently being run in India must also focus on safe disposal of children's stool.

  18. Evaluation of Four Automated Protocols for Extraction of DNA from FTA Cards

    DEFF Research Database (Denmark)

    Stangegaard, Michael; Børsting, Claus; Ferrero-Miliani, Laura

    2013-01-01

    protocols on three different instruments. The extraction processes were repeated up to six times with the same pieces of FTA cards. The sample material on the FTA cards was either blood or buccal cells. With the QIAamp DNA Investigator and QIAsymphony DNA Investigator kits, it was possible to extract DNA...... from the FTA cards in all six rounds of extractions in sufficient amount and quality to obtain complete short tandem repeat (STR) profiles on a QIAcube and a QIAsymphony SP. With the PrepFiler Express kit, almost all the extractable DNA was extracted in the first two rounds of extractions. Furthermore......, we demonstrated that it was possible to successfully extract sufficient DNA for STR profiling from previously processed FTA card pieces that had been stored at 4 °C for up to 1 year. This showed that rare or precious FTA card samples may be saved for future analyses even though some DNA was already...

  19. A loop-mediated isothermal amplification (LAMP assay for early detection of Schistosoma mansoni in stool samples: a diagnostic approach in a murine model.

    Directory of Open Access Journals (Sweden)

    Pedro Fernández-Soto

    2014-09-01

    Full Text Available Human schistosomiasis, mainly due to Schistosoma mansoni species, is one of the most prevalent parasitic diseases worldwide. To overcome the drawbacks of classical parasitological and serological methods in detecting S. mansoni infections, especially in acute stage of the disease, development of cost-effective, simple and rapid molecular methods is still needed for the diagnosis of schistosomiasis. A promising approach is the loop-mediated isothermal amplification (LAMP technology. Compared to PCR-based assays, LAMP has the advantages of reaction simplicity, rapidity, specificity, cost-effectiveness and higher amplification efficiency. Additionally, as results can be inspected by the naked eye, the technique has great potential for use in low-income countries.A sequence corresponding to a mitochondrial S. mansoni minisatellite DNA region was selected as a target for designing a LAMP-based method to detect S. mansoni DNA in stool samples. We used a S. mansoni murine model to obtain well defined stool and sera samples from infected mice with S. mansoni cercariae. Samples were taken weekly from week 0 to 8 post-infection and the Kato-Katz and ELISA techniques were used for monitoring the infection. Primer set designed were tested using a commercial reaction mixture for LAMP assay and an in house mixture to compare results. Specificity of LAMP was tested using 16 DNA samples from different parasites, including several Schistosoma species, and no cross-reactions were found. The detection limit of our LAMP assay (SmMIT-LAMP was 1 fg of S. mansoni DNA. When testing stool samples from infected mice the SmMIT-LAMP detected S. mansoni DNA as soon as 1 week post-infection.We have developed, for the first time, a cost-effective, easy to perform, specific and sensitive LAMP assay for early detection of S. mansoni in stool samples. The method is potentially and readily adaptable for field diagnosis and disease surveillance in schistosomiasis-endemic areas.

  20. Multiplex PCR detection of Cryptosporidium sp, Giardia lamblia and Entamoeba histolytica directly from dried stool samples from Guinea-Bissauan children with diarrhoea.

    Science.gov (United States)

    Mero, Sointu; Kirveskari, Juha; Antikainen, Jenni; Ursing, Johan; Rombo, Lars; Kofoed, Poul-Erik; Kantele, Anu

    2017-09-01

    In developing countries, diarrhoea is the most common cause of death for children under five years of age, with Giardia lamblia, Cryptosporidium and Entamoeba histolytica as the most frequent pathogenic parasites. Traditional microscopy for stool parasites has poor sensitivity and specificity, while new molecular methods may provide more accurate diagnostics. In poor regions with sample storage hampered by uncertain electricity supply, research would benefit from a method capable of analysing dried stools. A real-time multiplex PCR method with internal inhibition control was developed for detecting Giardia lamblia, Cryptosporidium hominis/parvum and Entamoeba histolytica directly from stool specimens. Applicability to dried samples was checked by comparing with fresh ones in a small test material. Finally, the assay was applied to dried specimens collected from Guinea-Bissauan children with diarrhoea. The PCR's analytical sensitivity limit was 0.1 ng/ml for G. lamblia DNA, 0.01 ng/ml for E. histolytica DNA and 0.1 ng/ml for Cryptosporidium sp. In the test material, the assay performed similarly with fresh and dried stools. Of the 52 Guinea-Bissauan samples, local microscopy revealed a parasite in 15%, while PCR detected 62% positive for at least one parasite: 44% of the dried samples had Giardia, 23% Cryptosporidium and 0% E. histolytica. Our new multiplex real-time PCR for protozoa presents a sensitive method applicable to dried samples. As proof of concept, it worked well on stools collected from Guinea-Bissauan children with diarrhoea. It provides an epidemiological tool for analysing dried specimens from regions poor in resources.

  1. Quality control of parasitology stool examination in Tabriz clinical laboratories

    Directory of Open Access Journals (Sweden)

    shahram Khademvatan

    2011-06-01

    Full Text Available The purpose of quality control program was to make doctors and laboratory personnel trust in laboratory results and consequently increasing confidence in laboratory achievements. The quality assurance means raising the level of quality in all tests that lead to raising the level of work efficiency and laboratories including minimum expense for society and minimum time for lab personnel. This study aimed to assess and determine the accuracy and precision of results in Tabriz medical diagnostic laboratories. Materials and Methods: In this retrospective study, 790 stool samples were selected randomly and tested by standard methods.Student t- test, SPSS software and sensitivity and accuracy formulas were used for data analysis. Results: The sensitivity was 62%, 22% and 8% with 95% confidence intervals for worm's eggs, protozoan cysts and trophozoite detection respectively. Conclusion: To elevate quality assurance in clinical diagnostic laboratory, monitoring and check of the laboratories by standard methods continually should be done.

  2. Treatment guidelines for primary nonretentive encopresis and stool toileting refusal.

    Science.gov (United States)

    Kuhn, B R; Marcus, B A; Pitner, S L

    1999-04-15

    Nonretentive encopresis refers to inappropriate soiling without evidence of fecal constipation and retention. This form of encopresis accounts for up to 20 percent of all cases. Characteristics include soiling accompanied by daily bowel movements that are normal in size and consistency. An organic cause for nonretentive encopresis is rarely identified. The medical assessment is usually normal, and signs of constipation are noticeably absent. A full developmental and behavioral assessment should be made to establish that the child is ready for intervention to correct encopresis and to identify any barriers to success, particularly disruptive behavior problems. Successful interventions depend on the presence of soft, comfortable bowel movements and addressing toilet refusal behavior. Daily scheduled positive toilet sits are recommended. Incentives may be used to reinforce successful defecation during these sits. A plan for management of stool withholding should be agreed on by the parents/caretakers and the family physician before intervention.

  3. Clinical illnesses associated with isolation of dysgonic fermenter 3 from stool samples.

    OpenAIRE

    Blum, R N; Berry, C D; Phillips, M G; Hamilos, D L; Koneman, E W

    1992-01-01

    The clinical significance of the fastidious organism DF-3 isolated from stool cultures is unclear. We sought to improve our understanding of this organism and to further define its association with human disease. Stool cultures for DF-3 were obtained from three sources: an ongoing study of enteric pathogens in patients infected with the human immunodeficiency virus, a screening procedure in which all stool samples submitted for Clostridium difficile toxin assay were cultured for DF-3, and sto...

  4. Value of Tropheryma whipplei quantitative polymerase chain reaction assay for the diagnosis of Whipple disease: usefulness of saliva and stool specimens for first-line screening.

    Science.gov (United States)

    Fenollar, Florence; Laouira, Sonia; Lepidi, Hubert; Rolain, Jean-Marc; Raoult, Didier

    2008-09-01

    Whipple disease (WD) is a chronic infectious disease caused by Tropheryma whipplei. WD DNA has been found in stool and saliva specimens from patients and asymptomatic carriers. A total of 4418 samples that were sent to our center for determination of WD were tested by a T. whipplei-specific quantitative polymerase chain reaction (PCR) based on repetitive sequences. Definite WD was diagnosed in 71 patients, including 55 patients with classic WD (defined by positive results of periodic acid-Schiff staining and/or specific immunohistochemistry of small-bowel biopsy specimens) and 16 patients with localized WD (including patients with endocarditis, neurologic infection, and uveitis). Of the persons without WD, 2.3% had stool specimens positive for T. whipplei by PCR and 0.2% had saliva specimens positive for T. whipplei by PCR. Diagnosis of WD was likely in patients with positive results of both PCR of saliva specimens and PCR of stool specimens (positive predictive value, 95.2%). When the bacterial load was >10(4) colony-forming units per g of stool, the positive predictive value was 100%. A negative result of PCR of a saliva or stool specimen had a negative predictive value of 99.2% for classic WD. For localized WD, positive results of both PCR of saliva specimens and PCR of stool specimens had a sensitivity of 58% (compared with 94% for classic WD). The positive predictive value of testing of blood, cerebrospinal fluid, and urine specimens was 100% for each, and the positive predictive value for testing of duodenal biopsy specimens was 97.5%. T. whipplei-specific quantitative PCR of saliva and stool specimens should be performed as first-line noninvasive screening for WD. When the results for both types of specimens are positive, diagnosis of classic WD should be highly suspected, especially if a high bacterial load is detected. Because PCR of saliva and stool specimens lacks sensitivity for determination of localized WD, invasive samples should be tested on the

  5. Stool microbiome and metabolome differences between colorectal cancer patients and healthy adults

    Science.gov (United States)

    In this study we used stool profiling to identify intestinal bacteria and metabolites that are differentially represented in humans with colorectal cancer (CRC) compared to healthy controls to identify how microbial functions may influence CRC development. Stool samples were collected from healthy a...

  6. How to: Establish and run a stool bank.

    Science.gov (United States)

    Terveer, E M; van Beurden, Y H; Goorhuis, A; Seegers, J F M L; Bauer, M P; van Nood, E; Dijkgraaf, M G W; Mulder, C J J; Vandenbroucke-Grauls, C M J E; Verspaget, H W; Keller, J J; Kuijper, E J

    2017-12-01

    Since 2013, several stool banks have been developed following publications reporting on clinical success of 'faecal microbiota transplantation' (FMT) for recurrent Clostridium difficile infections (CDI). However, protocols for donor screening, faecal suspension preparation, and transfer of the faecal suspension differ between countries and institutions. Moreover, no European consensus exists regarding the legislative aspects of the faecal suspension product. Internationally standardized recommendations about the above mentioned aspects have not yet been established. In 2015, the Netherlands Donor Feces Bank (NDFB) was founded with the primary aim of providing a standardized product for the treatment of patients with recurrent CDI in the Netherlands. Standard operation procedures for donor recruitment, donor selection, donor screening, and production, storage, and distribution of frozen faecal suspensions for FMT were formulated. Our experience summarized in this review addresses current donor recruitment and screening, preparation of the faecal suspension, transfer of the faecal microbiota suspension, and the experiences and follow-up of the patients treated with donor faeces from the NDFB. Copyright © 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  7. Direct quantification of cell-free, circulating DNA from unpurified plasma.

    Science.gov (United States)

    Breitbach, Sarah; Tug, Suzan; Helmig, Susanne; Zahn, Daniela; Kubiak, Thomas; Michal, Matthias; Gori, Tommaso; Ehlert, Tobias; Beiter, Thomas; Simon, Perikles

    2014-01-01

    Cell-free DNA (cfDNA) in body tissues or fluids is extensively investigated in clinical medicine and other research fields. In this article we provide a direct quantitative real-time PCR (qPCR) as a sensitive tool for the measurement of cfDNA from plasma without previous DNA extraction, which is known to be accompanied by a reduction of DNA yield. The primer sets were designed to amplify a 90 and 222 bp multi-locus L1PA2 sequence. In the first module, cfDNA concentrations in unpurified plasma were compared to cfDNA concentrations in the eluate and the flow-through of the QIAamp DNA Blood Mini Kit and in the eluate of a phenol-chloroform isoamyl (PCI) based DNA extraction, to elucidate the DNA losses during extraction. The analyses revealed 2.79-fold higher cfDNA concentrations in unpurified plasma compared to the eluate of the QIAamp DNA Blood Mini Kit, while 36.7% of the total cfDNA were found in the flow-through. The PCI procedure only performed well on samples with high cfDNA concentrations, showing 87.4% of the concentrations measured in plasma. The DNA integrity strongly depended on the sample treatment. Further qualitative analyses indicated differing fractions of cfDNA fragment lengths in the eluate of both extraction methods. In the second module, cfDNA concentrations in the plasma of 74 coronary heart disease patients were compared to cfDNA concentrations of 74 healthy controls, using the direct L1PA2 qPCR for cfDNA quantification. The patient collective showed significantly higher cfDNA levels (mean (SD) 20.1 (23.8) ng/ml; range 5.1-183.0 ng/ml) compared to the healthy controls (9.7 (4.2) ng/ml; range 1.6-23.7 ng/ml). With our direct qPCR, we recommend a simple, economic and sensitive procedure for the quantification of cfDNA concentrations from plasma that might find broad applicability, if cfDNA became an established marker in the assessment of pathophysiological conditions.

  8. LUMINEX®: a new technology for the simultaneous identification of five Entamoeba spp. commonly found in human stools.

    Science.gov (United States)

    Santos, Helena Lúcia Carneiro; Bandyopadhyay, Kakali; Bandea, Rebecca; Peralta, Regina Helena Saramago; Peralta, José Mauro; Da Silva, Alexandre Januário

    2013-03-15

    Six species of the genus Entamoeba, i.e., E. histolytica, E. dispar, E. moshkovskii, E. polecki, E. coli, and E. hartmanii can be found in human stools. Among these, only E. histolytica is considered to be pathogenic, causing intestinal and extra-intestinal disease, but it is morphologically identical to E. dispar and E. moshkovskii. In general, E. polecki, E. coli, and E. hartmanii can be differentiated morphologically from E. histolytica, but some of their diagnostic morphologic features may overlap creating issues for the differential diagnosis. Moreover, the previous inability to differentiate among Entamoeba species has limited epidemiologic information on E histolytica. The objective of this study was to develop a rapid, high-throughput screening method using Luminex technique for the simultaneous detection and differentiation of Entamoeba species. PCR amplification was performed with biotinylated Entamoeba sp 18S rRNA gene primers, designed to amplify a fragment ranging from 382 to 429 bp of the Entamoeba spp studied. Regions of this fragment that could differentiate among E. histolytica, E. moshkovskii, E. dispar, E. hartmanii and E. coli were selected to design hybridization probes to link to Luminex beads. The assay was standardized with cloned DNA samples of each species and evaluated with 24 DNA extracts from samples obtained from individuals diagnosed with these amebas in their stools. Using this approach we were able to correctly identify E. histoltyica, E. dispar, E hartmanni, E. coli and E. moshkovskii in all specimens studied. From twenty four samples tested by microscopy, PCR/DNA Sequencing and real-time PCR, 100% agreed with PCR-Luminex assay for identification of E. dispar, E. moshkovskii, E. hartmanni, E. histolytica, and E. coli. These results show that this method could be used in the diagnostic detection of Entamoeba spp in fecal samples. This diagnostic test was useful to clearly distinguish E histolytica from other species and also to

  9. An Optimized DNA Analysis Workflow for the Sampling, Extraction, and Concentration of DNA obtained from Archived Latent Fingerprints.

    Science.gov (United States)

    Solomon, April D; Hytinen, Madison E; McClain, Aryn M; Miller, Marilyn T; Dawson Cruz, Tracey

    2018-01-01

    DNA profiles have been obtained from fingerprints, but there is limited knowledge regarding DNA analysis from archived latent fingerprints-touch DNA "sandwiched" between adhesive and paper. Thus, this study sought to comparatively analyze a variety of collection and analytical methods in an effort to seek an optimized workflow for this specific sample type. Untreated and treated archived latent fingerprints were utilized to compare different biological sampling techniques, swab diluents, DNA extraction systems, DNA concentration practices, and post-amplification purification methods. Archived latent fingerprints disassembled and sampled via direct cutting, followed by DNA extracted using the QIAamp® DNA Investigator Kit, and concentration with Centri-Sep™ columns increased the odds of obtaining an STR profile. Using the recommended DNA workflow, 9 of the 10 samples provided STR profiles, which included 7-100% of the expected STR alleles and two full profiles. Thus, with carefully selected procedures, archived latent fingerprints can be a viable DNA source for criminal investigations including cold/postconviction cases. © 2017 American Academy of Forensic Sciences.

  10. Stool frequency recording in severe acute malnutrition ('StoolSAM'); an agreement study comparing maternal recall versus direct observation using diapers.

    Science.gov (United States)

    Voskuijl, Wieger; Potani, Isabel; Bandsma, Robert; Baan, Anne; White, Sarah; Bourdon, Celine; Kerac, Marko

    2017-06-07

    Approximately 50% of the deaths of children under the age of 5 can be attributed to undernutrition, which also encompasses severe acute malnutrition (SAM). Diarrhoea is strongly associated with these deaths and is commonly diagnosed solely based on stool frequency and consistency obtained through maternal recall. This trial aims to determine whether this approach is equivalent to a 'directly observed method' in which a health care worker directly observed stool frequency using diapers in hospitalised children with complicated SAM. This study was conducted at 'Moyo' Nutritional Rehabilitation Unit, Queen Elizabeth Central Hospital, Malawi. Participants were children aged 5-59 months admitted with SAM. We compared 2 days of stool frequency data obtained with next-day maternal-recall versus a 'gold standard' in which a health care worker observed stool frequency every 2 h using diapers. After study completion, guardians were asked their preferred method and their level of education. We found poor agreement between maternal recall and the 'gold standard' of directly observed diapers. The sensitivity to detect diarrhoea based on maternal recall was poor, with only 75 and 56% of diarrhoea cases identified on days 1 and 2, respectively. However, the specificity was higher with more than 80% of children correctly classified as not having diarrhoea. On day 1, the mean stool frequency difference between the two methods was -0.17 (SD; 1.68) with limits of agreement (of stool frequency) of -3.55 and 3.20 and, similarly on day 2, the mean difference was -0.2 (SD; 1.59) with limits of agreement of -3.38 and 2.98. These limits extend beyond the pre-specified 'acceptable' limits of agreement (±1.5 stool per day) and indicate that the 2 methods are non-equivalent. The higher the stool frequency, the more discrepant the two methods were. Most primary care givers strongly preferred using diapers. This study shows lack of agreement between the assessment of stool frequency in SAM

  11. Consistency of direct microscopic examination and ELISA in detection of Giardia in stool specimen among children

    Directory of Open Access Journals (Sweden)

    Zohreh Torabi

    2014-09-01

    Full Text Available Objective: To investigate the consistency of direct microscopic examination and ELISA for determination of Giadia in stool specimen. Method: Study population consisted of children with any clinical symptoms of Giardia infestation since last two weeks. Fresh stool specimen was collected from each child. The stools specimens were assessed by two methods of direct microscopic examination and ELISA.The degree of agreement between direct stool exam and ELISA was calculated by Cohen's kappa coefficient. Results: In this study, 124 children with age range 2-12 years were investigated. A total of 64 (61.7% and 79 (65.7% of children had Giardia by direct stool exam and ELISA test respectively. There was association between frequency of constipation and Giardia infection (P=0.036. The Cohen's kappa coefficient calculated for degree of agreement between direct stool exam and ELISA showed κ=0.756 (P<0.001. Conclusions: The frequency of Giardia infection in symptomatic children was high and there was high agreement rate between ELISA and direct stool smear.

  12. Using a commercially available DNA extraction kit to obtain high quality human genomic DNA suitable for PCR and genotyping from 11-year-old saliva saturated cotton spit wads

    Directory of Open Access Journals (Sweden)

    Hudziak James J

    2008-12-01

    Full Text Available Abstract Background We sought to describe the integrity of human genomic DNA extracted from saliva saturated cotton spit wads stored at -20°C for approximately 11 years. 783 spit wad samples were collected from an ADHD sample population (Vermont Family Study during 1996–2000. Human genomic DNA was extracted from the spit wads using a commercially available kit; QIAamp DNA Blood Midi Kit (Qiagen, Inc., Valencia, CA. with a few modifications. Results The resulting DNA yield was more than adequate for genetic analysis and ranged from approximately 1 μg to a total of 80 μg (mean 17.3 μgs ± 11.9 μgs. A260/A280 ratios for the human genomic DNA extracted from the spit wads was consistently within the generally acceptable values of 1.7–2.0, with the lowest purity being 1.70, and a mean value of 1.937 ± 0.226 for the 783 samples. The DNA also was suitable for PCR reactions as evidenced by the amplification of the serotonin-transporter-linked polymorphic region, 5HTTLPR. 5HTTLPR is a functional polymorphism in the promoter region of the serotonin transporter gene (HTT, SLC6A4, or SERT, consisting of two intensively studied alleles. 770 of the 783 samples (98.3% produced fragments after PCR of the expected size with primers specific for 5HTTLPR. Conclusion High quality and abundant genomic DNA can be successfully retrieved from saliva saturated cotton spit wads using the commercially available kit, QIAamp DNA Blood Midi Kit from Qiagen, Inc. Furthermore, the DNA can be extracted in less than 3 hours and multiple samples can be processed simultaneously thus reducing processing time.

  13. Antimicrobial resistance of bacterial enteropathogens isolated from stools in Madagascar.

    Science.gov (United States)

    Randrianirina, Frederique; Ratsima, Elisoa Hariniana; Ramparany, Lova; Randremanana, Rindra; Rakotonirina, Hanitra Clara; Andriamanantena, Tahiry; Rakotomanana, Fanjasoa; Rajatonirina, Soatiana; Richard, Vincent; Talarmin, Antoine

    2014-02-25

    Diarrheal diseases are a major public health problem in developing countries, and are one of the main causes of hospital admissions in Madagascar. The Pasteur Institute of Madagascar undertook a study to determine the prevalence and the pathogenicity of bacterial, viral and protozoal enteropathogens in diarrheal and non-diarrheal stools of children aged less than 5 years in Madagascar. We present here the results of the analysis of antimicrobial susceptibility of the bacteria isolated during this study. The study was conducted in the community setting in 14 districts of Madagascar from October 2008 to May 2009. Conventional methods and PCR were used to identify the bacteria; antimicrobial susceptibility was determined using an agar diffusion method for enterobacteriaceae and MICs were measured by an agar dilution method for Campylobacter sp. In addition to the strains isolated during this study, Salmonella sp and Shigella sp isolated at the Pasteur Institute of Madagascar from 2005 to 2009 were included in the analysis to increase the power of the study. Twenty-nine strains of Salmonella sp, 35 strains of Shigella sp, 195 strains of diarrheagenic E. coli, 203 strains of C. jejuni and 71 strains of C. coli isolated in the community setting were tested for antibiotic resistance. Fifty-five strains of Salmonella sp and 129 strains of Shigella sp isolated from patients referred to the Pasteur Institute of Madagascar were also included in the study. Many E. coli and Shigella isolates (around 80%) but fewer Salmonella isolates were resistant to ampicillin and trimethoprim/sulfamethoxazole. A small proportion of strains of each species were resistant to ciprofloxacin and only 3% of E. coli strains presented a resistance to third generation cephalosporins due to the production of extended-spectrum beta-lactamases. The resistance of Campylobacter sp to ampicillin was the most prevalent, whereas less than 5% of isolates were resistant to each of the other antibiotics. The

  14. High-Quality and -Quantity DNA Extraction from Frozen Archival Blood Clots for Genotyping of Single-Nucleotide Polymorphisms

    DEFF Research Database (Denmark)

    Bank, Steffen; Nexø, Bjørn Andersen; Andersen, Vibeke

    2013-01-01

    the efficiency of commercial purification kits for extracting DNA from long-term frozen clotted blood. Methods: Serum tubes with clotted blood were stored at −20°C for 1 to 2.5 years before DNA extraction. DNA was extracted from 10 blood clot samples using PureGene (Qiagen) with and without glycogen, the QIAamp...... with a median of 0.65 μg (range 0.5–2.6 μg) pr 300 μL total blood. Conclusion: The yield obtained by the different commercial kits varied considerably. Our work demonstrates that high-quality and -quantity DNA can be extracted with the Maxwell 16 Blood purification kit (Promega) from cryopreserved blood clots...

  15. Stool frequency recording in severe acute malnutrition ('StoolSAM'); an agreement study comparing maternal recall versus direct observation using diapers

    NARCIS (Netherlands)

    Voskuijl, Wieger; Potani, Isabel; Bandsma, Robert; Baan, Anne; White, Sarah; Bourdon, Celine; Kerac, Marko

    2017-01-01

    Background: Approximately 50% of the deaths of children under the age of 5 can be attributed to undernutrition, which also encompasses severe acute malnutrition (SAM). Diarrhoea is strongly associated with these deaths and is commonly diagnosed solely based on stool frequency and consistency

  16. Stool consistency and stool frequency are excellent clinical markers for adequate colon preparation after polyethylene glycol 3350 cleansing protocol: a prospective clinical study in children.

    Science.gov (United States)

    Safder, Shaista; Demintieva, Yulia; Rewalt, Mary; Elitsur, Yoram

    2008-12-01

    Colon preparation for a colonoscopy in children is a difficult task because of the unpalatable taste and large volume of cleansing solution that needs to be consumed to ensure a clean colon. Consequently, an unprepared colon frequently occurs in routine practices, which causes early termination and a repeated procedure. (1) To assess the effectiveness of polyethylene glycol solution (PEG 3350) in preparing the colon of children scheduled for a colonoscopy and (2) to investigate clinical markers associated with an adequate colon preparation before a colonoscopy. A total of 167 children scheduled for a colonoscopy. In a prospective study, children scheduled for a colonoscopy were given PEG 3350 solution (1.5 g/kg per day, up to 100 g/d) over a 4-day preparation period. Each day, a simple questionnaire that documents the amount of liquid consumed, adverse effects, and the number and consistency of stool was completed by the parents. After a colonoscopy procedure, the colon preparation was assigned a number grade. The data were later assessed and were compared to determine the association between the grade of cleansing and the frequency and/or consistency of stool during preparation. Colon preparation was completed in 149 children, 133 of whom were adequately prepared. Inadequate preparation was found in 16 children; the procedure was terminated prematurely in 2 of these patients because of unacceptable conditions. No significant adverse effects were noted. A number of >or=5 stools/d, and liquid stool consistency in the last 2 days of preparation were associated with adequate colon preparation. PEG 3350 solution is safe, efficacious, and tolerable for children. Stool frequency and consistency in the last 2 days of preparation were excellent markers (positive predictive value 91%-95%), which predict an adequately clean colon before a colonoscopy in children.

  17. Oral diosmectite reduces stool output and diarrhea duration in children with acute watery diarrhea.

    Science.gov (United States)

    Dupont, Christophe; Foo, Jimmy Lee Kok; Garnier, Philippe; Moore, Nicholas; Mathiex-Fortunet, Hèlène; Salazar-Lindo, Eduardo

    2009-04-01

    Diosmectite is a clay used to treat children with acute watery diarrhea. However, its effects on stool output reduction, the key outcome for pediatric antidiarrheal drugs, have not been shown. Two parallel, double-blind studies of diosmectite efficacy on stool reduction were conducted in children 1 to 36 months old in Peru (n = 300) and Malaysia (n = 302). Inclusion criteria included 3 or more watery stools per day for less than 72 hours and weight/height ratios of 0.8 or greater. Exclusion criteria were the need for intravenous rehydration, gross blood in stools, fever higher than 39 degrees C, or current treatment with antidiarrheal or antibiotic medications. Rotavirus status was determined. Diosmectite dosage was 6 g/day (children 1-12 months old) or 12 g/day (children 13-36 months old), given for at least 3 days, followed by half doses until complete recovery. Patients were assigned randomly to groups given diosmectite or placebo, in addition to oral rehydration solution (World Health Organization). Children in each study had comparable average ages and weights. The frequencies of rotavirus infection were 22% in Peru and 12% in Malaysia. Similar amounts of oral rehydration solution were given to children in the diosmectite and placebo groups. Stool output was decreased significantly by diosmectite in both studies, especially among rotavirus-positive children. In pooled data, children had a mean stool output of 94.5 +/- 74.4 g/kg of body weight in the diosmectite group versus 104.1 +/- 94.2 g/kg in the placebo group (P = .002). Diarrhea duration was reduced by diosmectite, which was well tolerated. These results show that diosmectite significantly decreased stool output in children with acute watery diarrhea, especially those who were rotavirus-positive.

  18. High frequency of parasitic and viral stool pathogens in patients with active ulcerative colitis: report from a tropical country.

    Science.gov (United States)

    Banerjee, Debabrata; Deb, Rachana; Dar, Lalit; Mirdha, Bijay R; Pati, Sunil K; Thareja, Sandeep; Falodia, Sushil; Ahuja, Vineet

    2009-01-01

    Diarrhoeal relapses in patients with ulcerative colitis (UC) may be associated with enteric infections and its diagnosis may lessen avoidable exposure to corticosteroids and/or immunosuppressants. The purpose of this study was to assess the frequency of stool pathogens (parasitic and viral) in patients with active UC. This prospective cross-sectional study included 49 consecutive patients (32 M, 17 F, mean age 35.8+/-12 years) with active UC. Three stool samples were collected from each patient and examined for parasitic infection. Rectal biopsies were obtained during sigmoidoscopy to demonstrate cytomegalovirus (CMV) inclusion bodies and to conduct qualitative polymerase chain reaction (PCR) for CMV and herpes simplex virus (HSV) DNA detection. Median duration of illness was 3.9+/-3.7 years and 83.7% of the patients had moderate to severe disease. The prevalence of parasitic infections in UC was 12%. The organisms isolated were Strongyloides stercoralis in 4%, Ankylostoma duodenale in 4%, Cryptosporidium in 2% and Entamoeba histolytica in 2% of the patients. The prevalence of CMV and HSV in rectal biopsies using qualitative PCR was 8% and 10%, respectively. No predictive factor was identified with CMV superinfection in patients with active UC. In India there is a high prevalence of parasitic and viral infections in patients with active UC. The results of the study suggest that, in tropical countries with a known high prevalence of parasitic diseases, aggressive evaluation for parasitic and viral infections should be carried out, as early identification and prompt treatment of such infections can improve the clinical course of patients with active UC.

  19. Modification of stool's water content in constipated infants: management with an adapted infant formula

    Directory of Open Access Journals (Sweden)

    Alvarez Marina M

    2011-05-01

    Full Text Available Abstract Background Constipation is a common occurrence in formula-fed infants. The aim of this preliminary study was to evaluate the impact of a formula with high levels of lactose and magnesium, in compliance with the official regulations, on stool water content, as well as a parental assessment of constipation. Materials and methods Thirty healthy term-born, formula-fed infants, aged 4-10 weeks, with functional constipation were included. All infants were full-term and fed standard formula. Exclusion criteria were preterm and/or low birth weight, organic constipation, being breast fed or fed a formula specially designed to treat constipation. Stool composition was measured by near-infrared reflectance analysis (NIRA and parents answered questions about crying associated with defecation and stool consistency at baseline and after two weeks of the adapted formula. Results After 2 weeks of the adapted formula, stool water content increased from 71 +/- 8.1% to 84 +/- 5.9%, (p Conclusions This preliminary study suggests that an adapted formula with high levels of lactose and magnesium increases stool water content and improves symptoms of constipation in term-born, formula-fed infants. A larger randomized placebo-controlled trial is indicated.

  20. Evaluation of four automated protocols for extraction of DNA from FTA cards.

    Science.gov (United States)

    Stangegaard, Michael; Børsting, Claus; Ferrero-Miliani, Laura; Frank-Hansen, Rune; Poulsen, Lena; Hansen, Anders J; Morling, Niels

    2013-10-01

    Extraction of DNA using magnetic bead-based techniques on automated DNA extraction instruments provides a fast, reliable, and reproducible method for DNA extraction from various matrices. Here, we have compared the yield and quality of DNA extracted from FTA cards using four automated extraction protocols on three different instruments. The extraction processes were repeated up to six times with the same pieces of FTA cards. The sample material on the FTA cards was either blood or buccal cells. With the QIAamp DNA Investigator and QIAsymphony DNA Investigator kits, it was possible to extract DNA from the FTA cards in all six rounds of extractions in sufficient amount and quality to obtain complete short tandem repeat (STR) profiles on a QIAcube and a QIAsymphony SP. With the PrepFiler Express kit, almost all the extractable DNA was extracted in the first two rounds of extractions. Furthermore, we demonstrated that it was possible to successfully extract sufficient DNA for STR profiling from previously processed FTA card pieces that had been stored at 4 °C for up to 1 year. This showed that rare or precious FTA card samples may be saved for future analyses even though some DNA was already extracted from the FTA cards.

  1. Maintaining Breast Cancer Specimen Integrity and Individual or Simultaneous Extraction of Quality DNA, RNA, and Proteins from Allprotect-Stabilized and Nonstabilized Tissue Samples

    LENUS (Irish Health Repository)

    Mee, Blanaid C.

    2011-12-29

    The Saint James\\'s Hospital Biobank was established in 2008, to develop a high-quality breast tissue BioResource, as a part of the breast cancer clinical care pathway. The aims of this work were: (1) to ascertain the quality of RNA, DNA, and protein in biobanked carcinomas and normal breast tissues, (2) to assess the efficacy of AllPrep® (Qiagen) in isolating RNA, DNA, and protein simultaneously, (3) to compare AllPrep with RNEasy® and QIAamp® (both Qiagen), and (4) to examine the effectiveness of Allprotect® (Qiagen), a new tissue stabilization medium in preserving DNA, RNA, and proteins. One hundred eleven frozen samples of carcinoma and normal breast tissue were analyzed. Tumor and normal tissue morphology were confirmed by frozen sections. Tissue type, tissue treatment (Allprotect vs. no Allprotect), extraction kit, and nucleic acid quantification were analyzed by utilizing a 4 factorial design (SPSS PASW 18 Statistics Software®). QIAamp (DNA isolation), AllPrep (DNA, RNA, and Protein isolation), and RNeasy (RNA isolation) kits were assessed and compared. Mean DNA yield and A260\\/280 values using QIAamp were 33.2 ng\\/μL and 1.86, respectively, and using AllPrep were 23.2 ng\\/μL and 1.94. Mean RNA yield and RNA Integrity Number (RIN) values with RNeasy were 73.4 ng\\/μL and 8.16, respectively, and with AllPrep were 74.8 ng\\/μL and 7.92. Allprotect-treated tissues produced higher RIN values of borderline significance (P=0.055). No discernible loss of RNA stability was detected after 6 h incubation of stabilized or nonstabilized tissues at room temperature or 4°C or in 9 freeze-thaw cycles. Allprotect requires further detailed evaluation, but we consider AllPrep to be an excellent option for the simultaneous extraction of RNA, DNA, and protein from tumor and normal breast tissues. The essential presampling procedures that maintain the diagnostic integrity of pathology specimens do not appear to compromise the quality of molecular isolates.

  2. Effects of olestra and sorbitol consumption on objective measures of diarrhea: impact of stool viscosity on common gastrointestinal symptoms.

    Science.gov (United States)

    McRorie, J; Zorich, N; Riccardi, K; Bishop, L; Filloon, T; Wason, S; Giannella, R

    2000-02-01

    The aim of this study was to determine the effects of olestra and sorbitol consumption on three accepted objective measures of diarrhea (stool output >250 g/day, liquid/watery stools, bowel movement frequency >3/day), and how stool composition influences reports of common gastrointestinal symptoms. A double-blind, placebo-controlled study compared the effects of sorbitol (40 g/day in candy), a poorly absorbed sugar-alcohol with known osmotic effects, with those of olestra (20 or 40 g/day in potato chips), a nonabsorbed fat, on objective measures of stool composition and GI symptoms. Sixty-six subjects resided on a metabolic ward for 12 days: 2 days lead-in, 4 days baseline, 6 days treatment. Sorbitol 40 g/day resulted in loose/liquid stools within 1-3 h of consumption. In contrast, olestra resulted in a dose-responsive stool softening effect after 2-4 days of consumption. Subjects reported "diarrhea" when mean stool apparent viscosity (peak force (PF), g) decreased from a perceived "normal" (mean +/- SE, 1355 +/- 224 g PF; firm stool) to loose (260 +/- 68 g PF) stool. Mean apparent viscosity of stool during treatment: placebo, 1363 +/- 280 g (firm); olestra 20 g/day 743 +/- 65 g (soft); olestra 40 g/day, 563 +/- 105 g (soft); and sorbitol 40 g/day, 249 +/- 53 g (loose). Of the 1098 stool samples collected, 38% (419/1098) were rated by subjects as "diarrhea," yet only 2% of treatment days (all in the sorbitol treatment group) met commonly accepted criteria for a clinical diarrhea. Sorbitol, but not olestra, increased the severity of abdominal cramping, urgency and nausea compared to placebo. Olestra consumption, at levels far in excess of normal snacking conditions, resulted in a gradual stool softening effect after several days of consumption, did not meet any of the three objective measures of diarrhea, and did not increase GI symptoms. Sorbitol consumption, at only 80% of the dose requiring a "laxative effect" information label, resulted in rapid onset loose

  3. Comparison of polycarbonate and cellulose acetate membrane filters for isolation of Campylobacter concisus from stool samples

    DEFF Research Database (Denmark)

    Linde Nielsen, Hans; Engberg, Jørgen; Ejlertsen, Tove

    2013-01-01

    One thousand seven hundred ninety-one diarrheic stool samples were cultivated for Campylobacter spp. We found a high prevalence of Campylobacter concisus with use of a polycarbonate filter (n = 114) compared to a cellulose acetate filter (n = 79) (P polycarbonate filter is superior...

  4. Not Your Run-of-the-Mill Art-Room Stools

    Science.gov (United States)

    Chrzanowski, Rose-Ann C.

    2010-01-01

    An art room should be a garden of visual stimulation, born of creativity, inquiry, critical thinking and intellectual conversation--and a little collaboration is not a bad thing either! When the author unpacked the new stools for her art room at the high school, she envisioned something more beautiful than the brown masonite circles that…

  5. The isolation of Moellerella wisconsensis from stool samples in the U.K.

    Science.gov (United States)

    Marshall, A R; Al-Jumaili, I J; Bint, A J

    1986-01-01

    Three strains of Moellerella wisconsensis were isolated from a total of 400 stool specimens screened for this organism by means of a new selective medium developed in this laboratory. This is the first report of the isolation of this organism in the U.K. The exact role of M. wisconsensis in causing diarrhoea remains to be elucidated.

  6. Differential clinical features and stool findings in shigellosis and amoebic dysentery

    NARCIS (Netherlands)

    Speelman, P.; McGlaughlin, R.; Kabir, I.; Butler, T.

    1987-01-01

    To obtain information that could assist the clinician to differentiate between shigellosis and amoebic dysentery, we compared clinical features and stool findings in 58 adult male patients in Bangladesh. Mean values indicated that patients with invasive amoebiasis were older and had a longer

  7. Detection of Salmonella typhi by nested polymerase chain reaction in blood, urine, and stool samples

    NARCIS (Netherlands)

    Hatta, Mochammad; Smits, Henk L.

    2007-01-01

    A nested polymerase chain reaction (PCR) specific for Salmonella enterica serovar Typhi was used for the detection of the pathogen in blood, urine, and stool samples from 131 patients with clinical suspicion of typhoid fever. The sensitivity of blood culture, the PCRs with blood, urine, and feces,

  8. Validation of the 3-day rule for stool bacterial tests in Japan.

    Science.gov (United States)

    Kobayashi, Masanori; Sako, Akahito; Ogami, Toshiko; Nishimura, So; Asayama, Naoki; Yada, Tomoyuki; Nagata, Naoyoshi; Sakurai, Toshiyuki; Yokoi, Chizu; Kobayakawa, Masao; Yanase, Mikio; Masaki, Naohiko; Takeshita, Nozomi; Uemura, Naomi

    2014-01-01

    Stool cultures are expensive and time consuming, and the positive rate of enteric pathogens in cases of nosocomial diarrhea is low. The 3-day rule, whereby clinicians order a Clostridium difficile (CD) toxin test rather than a stool culture for inpatients developing diarrhea >3 days after admission, has been well studied in Western countries. The present study sought to validate the 3-day rule in an acute care hospital setting in Japan. Stool bacterial and CD toxin test results for adult patients hospitalized in an acute care hospital in 2008 were retrospectively analyzed. Specimens collected after an initial positive test were excluded. The positive rate and cost-effectiveness of the tests were compared among three patient groups. The adult patients were divided into three groups for comparison: outpatients, patients hospitalized for ≤3 days and patients hospitalized for ≥4 days. Over the 12-month period, 1,597 stool cultures were obtained from 992 patients, and 880 CD toxin tests were performed in 529 patients. In the outpatient, inpatient ≤3 days and inpatient ≥4 days groups, the rate of positive stool cultures was 14.2%, 3.6% and 1.3% and that of positive CD toxin tests was 1.9%, 7.1% and 8.5%, respectively. The medical costs required to obtain one positive result were 9,181, 36,075 and 103,600 JPY and 43,200, 11,333 and 9,410 JPY, respectively. The 3-day rule was validated for the first time in a setting other than a Western country. Our results revealed that the "3-day rule" is also useful and cost-effective in Japan.

  9. Clinical illnesses associated with isolation of dysgonic fermenter 3 from stool samples.

    Science.gov (United States)

    Blum, R N; Berry, C D; Phillips, M G; Hamilos, D L; Koneman, E W

    1992-02-01

    The clinical significance of the fastidious organism DF-3 isolated from stool cultures is unclear. We sought to improve our understanding of this organism and to further define its association with human disease. Stool cultures for DF-3 were obtained from three sources: an ongoing study of enteric pathogens in patients infected with the human immunodeficiency virus, a screening procedure in which all stool samples submitted for Clostridium difficile toxin assay were cultured for DF-3, and stool samples submitted specifically for DF-3 culture. Retrospective clinical data were obtained from chart reviews of patients with positive cultures. Antimicrobial susceptibility testing and cell wall fatty acid analysis were performed for each DF-3 isolated. Eight isolates of DF-3 were obtained over a period of 8 months. All patients either had severe underlying disease or were immunocompromised, including three patients coinfected with human immunodeficiency virus and two patients with inflammatory bowel disease. The spectrum of clinical disease ranged from chronic diarrhea with a well-defined response to therapy for DF-3 to an asymptomatic carrier state. Cell wall fatty acid analysis of these isolates demonstrated a consistent pattern with a large peak of 12-methyltetradecanoate. DF-3, a fastidious gram-negative coccobacillus, can be recovered from stool cultures of immunocompromised patients by using selective media. The presence of 12-methyltetradecanoate in cell wall fatty acid analysis assists in identification. The increased use of a selective medium-(cefoperazone-vancomycin-amphotericin B) in the evaluation of diarrhea in immunocompromised hosts, including persons with inflammatory bowel disease, may better define the association of DF-3 with human gastrointestinal disease.

  10. New ultrasonographic evaluation of stool and/or gas distribution for treatment of chronic constipation.

    Science.gov (United States)

    Manabe, Noriaki; Kamada, Tomoari; Hata, Jiro; Haruma, Ken

    2018-03-01

    The first aim of this study was to develop a new ultrasonographic method (US) to evaluate stool and/or gas distribution. The second aim was to apply this method to compare stool and/or gas distribution between healthy subjects and patients with chronic constipation and evaluate whether US parameters could be an alternative to the colonic transit time (CTT). We enrolled seven healthy volunteers (four men, three women; mean age 29.3 ± 5.2 years) who underwent US and computed tomography (CT) on the same day to evaluate the reproducibility of US results. We then enrolled 268 patients with chronic constipation (94 men, 174 women; mean age 63.3 ± 4.2 years) and 66 age- and sex-matched healthy subjects (controls). The transverse diameters of four segments of the colon [ascending (AC), transverse (TC), descending (DC), and sigmoid (SC)] and the rectum (R) were measured, and their stool and/or gas distribution was evaluated using the constipation index (CI) [AC + TC + DC + SC + R/5] and left/right (L/R) distribution [(DC + SC)/(AC + TC)]. The CTT was assessed using radiopaque markers. All healthy subjects underwent US and CT successfully, with a sufficiently high reproducibility coefficient for this method and significant correlation between the US and CT parameters. The stool and/or gas distribution evaluated by US showed a significant difference in one of the US parameters between healthy subjects and patients, and the CI was an indirect indicator for the CTT. These findings may assist physicians evaluate stool and/or gas distribution of patients with chronic constipation, which is an indirect indicator for CTT.

  11. Agreement Between Home-Based Measurement of Stool Calprotectin and ELISA Results for Monitoring Inflammatory Bowel Disease Activity

    NARCIS (Netherlands)

    Heida, Anke; Knol, Mariska; Kobold, Anneke Muller; Bootsman, Josette; Dijkstra, Gerard; van Rheenen, Patrick F

    2017-01-01

    BACKGROUND & AIMS: An increasing number of physicians use repeated measurements of stool calprotectin to monitor intestinal inflammation in patients with inflammatory bowel diseases (IBDs). A lateral flow-based rapid test allows patients to measure their own stool calprotectin values at home. The

  12. Determination of enterotoxigenic Clostridium perfringens by detecting of the cpa and cpe genes in stool samples of human origin, associated to gastrointestinal disease

    International Nuclear Information System (INIS)

    Oropeza Barrios, Gletty

    2014-01-01

    A molecular methodology is provided to the Centro Nacional de Referencia de Bacteriologia (CNRB) of the Instituto Costarricense de Investigacion y Ensenanza en Nutricion y Salud. An opportune diagnosis is realized of enterotoxigenic Clostridium perfringens in stool samples of sporadic cases and cases associated to foodborne disease outbreaks. DNA extraction of the white microorganism was performed through the methodology implemented in the CNRB. The technique of polymerase chain reaction (PCR) were adapted and standardized to establish the identification of C. perfringens to species level and detection of cpe gene coding for enterotoxin. The sensitivity of the method was determined in a selective culture medium for C. perfringens (Tryptose sulfite cycloserine Agar). A detection limit of about 2,3 x 10 4 CFU/ml was reached for the cpe gene and at least 2,8 x 10 2 CFU/ml for the cpa gene. Retrospective analysis of 61 samples of diarrheal stool suspicious by C. perfringens is performed to evaluate the efficacy of the technique. Three outbreaks caused by C. perfringens were identified and a 10% of positivity in the samples were obtained analyzed during the period between July 2012-March 2014 [es

  13. Stool Softeners

    Science.gov (United States)

    ... 50 Plus® (as a combination product containing Docusate, Sennosides) ... Gentle Strength® (as a combination product containing Docusate, Sennosides) ... Gentlax S® (as a combination product containing Docusate, Sennosides)

  14. Brazilian Portuguese translation, cross-cultural adaptation and reproducibility assessment of the modified Bristol Stool Form Scale for children.

    Science.gov (United States)

    Jozala, Debora Rodrigues; Oliveira, Isabelle Stefan de Faria; Ortolan, Erika Veruska Paiva; Oliveira Junior, Wilson Elias de; Comes, Giovana Tuccille; Cassettari, Vanessa Mello Granado; Self, Mariella Marie; Lourenção, Pedro Luiz Toledo de Arruda

    2018-03-15

    To translate and culturally adapt the modified Bristol Stool Form Scale for children into Brazilian Portuguese, and to evaluate the reproducibility of the translated version. The stage of translation and cross-cultural adaptation was performed according to an internationally accepted methodology, including the translation, back-translation, and pretest application of the translated version to a sample of 74 children to evaluate the degree of understanding. The reproducibility of the translated scale was assessed by applying the final version of Brazilian Portuguese modified Bristol Stool Form Scale for children to a sample of 64 children and 25 healthcare professionals, who were asked to correlate a randomly selected description from the translated scale with the corresponding representative illustration of the stool type. The final version of Brazilian Portuguese modified Bristol Stool Form Scale for children were evidently reproducible, since almost complete agreement (k>0,8) was obtained among the translated descriptions and illustrations of the stool types, both among the children and the group of specialists. The Brazilian Portuguese modified Bristol Stool Form Scale for children was shown to be reliable in providing very similar results for the same respondents at different times and for different examiners. The Brazilian Portuguese modified Bristol Stool Form Scale for children is reproducible; it can be applied in clinical practice and in scientific research in Brazil. Copyright © 2018 Sociedade Brasileira de Pediatria. Published by Elsevier Editora Ltda. All rights reserved.

  15. Molecular characterization of diarrheagenic Escherichia coli strains from stools samples and food products in Colombia

    OpenAIRE

    Rúgeles, Laura Cristina; Bai, Jing; Martínez, Aída Juliana; Vanegas, María Consuelo; Gómez-Duarte, Oscar Gilberto

    2010-01-01

    The prevalence of diarrheagenic E. coli in childhood diarrhea and the role of contaminated food products in disease transmission in Colombia are largely unknown. The aim of this study is to identify E. coli pathotypes, including E. coli O157:H7, from 108 stool samples from children with acute diarrhea, 38 meat samples and 38 vegetable samples. Multiplex PCR and Bax Dupont systems were used for E. coli pathotype detection. Eighteen (9.8%) E. coli diarrheagenic pathotypes were detected among al...

  16. Dipstick for rapid diagnosis of Shigella flexneri 2a in stool.

    Directory of Open Access Journals (Sweden)

    Faridabano Nato

    2007-04-01

    Full Text Available Shigellosis or bacillary dysentery, an acute bloody diarrhoea, is a major public health burden in developing countries. In the absence of prompt and appropriate treatment, the infection is often fatal, particularly in young malnourished children. Here, we describe a new diagnostic test for rapid detection, in stool, at the bedside of patients, of Shigella flexneri 2a, the most predominant agent of the endemic form of the disease.The test is based on the detection of S.flexneri 2a lipopolysaccharide (LPS using serotype 2a-specific monoclonal antibodies coupled to gold particles and displayed on one-step immunochromatographic dipstick. A concentration as low as 20 ng/ml of LPS is detected in distilled water and in reconstituted stools in under 15 minutes. The threshold of detection corresponds to a concentration of 5x10(7 CFU/ml of S. flexneri 2a, which provides an unequivocal positive reaction in three minutes in distilled water and reconstituted stools. The specificity is 100% when tested with a battery of Shigella and unrelated strains, in culture. When tested in Vietnam, on clinical samples, the specificity and sensitivity were 99.2 and 91.5%, respectively. A decrease of the sensitivity during the evaluation on stool samples was observed after five weeks at room temperature and was due to moistening of the dipsticks caused by the humidity of the air during the fifth week of the evaluation. This drawback is now overcome by improving the packaging and providing dipsticks individually wrapped in waterproof bags.This simple dipstick-bases test represents a powerful tool for case management and epidemiological surveys.

  17. Novel Stool-Based Protein Biomarkers for Improved Colorectal Cancer Screening: A Case-Control Study.

    Science.gov (United States)

    Bosch, Linda J W; de Wit, Meike; Pham, Thang V; Coupé, Veerle M H; Hiemstra, Annemieke C; Piersma, Sander R; Oudgenoeg, Gideon; Scheffer, George L; Mongera, Sandra; Sive Droste, Jochim Terhaar; Oort, Frank A; van Turenhout, Sietze T; Larbi, Ilhame Ben; Louwagie, Joost; van Criekinge, Wim; van der Hulst, Rene W M; Mulder, Chris J J; Carvalho, Beatriz; Fijneman, Remond J A; Jimenez, Connie R; Meijer, Gerrit A

    2017-12-19

    The fecal immunochemical test (FIT) for detecting hemoglobin is used widely for noninvasive colorectal cancer (CRC) screening, but its sensitivity leaves room for improvement. To identify novel protein biomarkers in stool that outperform or complement hemoglobin in detecting CRC and advanced adenomas. Case-control study. Colonoscopy-controlled referral population from several centers. 315 stool samples from one series of 12 patients with CRC and 10 persons without colorectal neoplasia (control samples) and a second series of 81 patients with CRC, 40 with advanced adenomas, and 43 with nonadvanced adenomas, as well as 129 persons without colorectal neoplasia (control samples); 72 FIT samples from a third independent series of 14 patients with CRC, 16 with advanced adenomas, and 18 with nonadvanced adenomas, as well as 24 persons without colorectal neoplasia (control samples). Stool samples were analyzed by mass spectrometry. Classification and regression tree (CART) analysis and logistic regression analyses were performed to identify protein combinations that differentiated CRC or advanced adenoma from control samples. Antibody-based assays for 4 selected proteins were done on FIT samples. In total, 834 human proteins were identified, 29 of which were statistically significantly enriched in CRC versus control stool samples in both series. Combinations of 4 proteins reached sensitivities of 80% and 45% for detecting CRC and advanced adenomas, respectively, at 95% specificity, which was higher than that of hemoglobin alone (P control samples (P control samples. Proof of concept that such proteins can be detected with antibody-based assays in small sample volumes indicates the potential of these biomarkers to be applied in population screening. Center for Translational Molecular Medicine, International Translational Cancer Research Dream Team, Stand Up to Cancer (American Association for Cancer Research and the Dutch Cancer Society), Dutch Digestive Foundation, and VU

  18. Techniques for the recovery and identification of Cryptosporidium oocysts from stool specimens.

    Science.gov (United States)

    Garcia, L S; Bruckner, D A; Brewer, T C; Shimizu, R Y

    1983-07-01

    Due to increasing numbers of patients with documented infections with Cryptosporidium and other coccidia, it is important for the physician and clinical laboratory to be aware of the appropriate diagnostic techniques necessary for organism recovery and identification. Although Cryptosporidium is found in the gastrointestinal tract, tissue biopsies may be insufficient for organism recovery; the examination of stool specimens is a noninvasive procedure and will provide better overall opportunities for organism recovery. Human clinical specimens were examined from 45 patients with confirmed cryptosporidiosis or suspected of having the infection. Tissue biopsy sections, fecal wet preparations, and permanent stained smears were examined. Stool specimens were submitted in 10% Formalin, 2.5% potassium dichromate, and polyvinyl alcohol and were examined for oocysts by using 15 different methods: phase-contrast and light microscopy; Sheather's sugar flotation; Formalin concentration techniques; 10% potassium hydroxide; Giemsa; trichrome; periodic acid-Schiff; modified periodic acid-Schiff; silver methenamine; acridine orange; auramine-rhodamine; Kinyoun acid-fast; Ziehl-Neelsen carbolfuchsin; and a modified acid-fast procedure. Each technique or combination of techniques was assessed by organism quantitation, organism morphology, and ease of visual recognition. Based on these comparative studies, the modified Ziehl-Neelsen carbolfuchsin stain on 10% Formalin-preserved stool is recommended for the recovery and identification of Cryptosporidium.

  19. The complete genome of klassevirus – a novel picornavirus in pediatric stool

    Directory of Open Access Journals (Sweden)

    Ganem Donald

    2009-06-01

    Full Text Available Abstract Background Diarrhea kills 2 million children worldwide each year, yet an etiological agent is not found in approximately 30–50% of cases. Picornaviral genera such as enterovirus, kobuvirus, cosavirus, parechovirus, hepatovirus, teschovirus, and cardiovirus have all been found in human and animal diarrhea. Modern technologies, especially deep sequencing, allow rapid, high-throughput screening of clinical samples such as stool for new infectious agents associated with human disease. Results A pool of 141 pediatric gastroenteritis samples that were previously found to be negative for known diarrheal viruses was subjected to pyrosequencing. From a total of 937,935 sequence reads, a collection of 849 reads distantly related to Aichi virus were assembled and found to comprise 75% of a novel picornavirus genome. The complete genome was subsequently cloned and found to share 52.3% nucleotide pairwise identity and 38.9% amino acid identity to Aichi virus. The low level of sequence identity suggests a novel picornavirus genus which we have designated klassevirus. Blinded screening of 751 stool specimens from both symptomatic and asymptomatic individuals revealed a second positive case of klassevirus infection, which was subsequently found to be from the index case's 11-month old twin. Conclusion We report the discovery of human klassevirus 1, a member of a novel picornavirus genus, in stool from two infants from Northern California. Further characterization and epidemiological studies will be required to establish whether klasseviruses are significant causes of human infection.

  20. The effect of inclined step stool on the quality of chest compression during in-hospital cardiopulmonary resuscitation.

    Science.gov (United States)

    Yun, Seong-Woo; Lee, Byung Kook; Jeung, Kyung Woon; Park, Sang Wook; Choi, Sung Soo; Lee, Chang-Hee; Ryu, So-Yeon

    2014-08-01

    A step stool is an ordinary device to improve the quality of chest compression (CC) during in-hospital cardiopulmonary resuscitation (CPR). We investigated the effect of an inclined step stool on the quality of CC during CPR on a hospital bed. We conducted a randomized crossover study of simulation using a manikin. Two different methods of CC were performed and compared: CC using a flat stool and CC using an inclined (20°) stool. Each session of CC was performed for 2 minutes using a metronome at a rate of 110 beats per minute. The primary outcome was the depth of CC. The adequate CC rate, duty cycle, rate of incomplete recoil, and the angle between the arm of the participants and the bed were also measured. The median value of the mean depth of CC was 50.5 mm (45.0-57.0 mm) in the flat stool group and 54.5 mm (47.0-58.3 mm) in the inclined stool group (P = .014). The adequate CC rate was significantly higher in the inclined stool group (84.2% [37.6%-99.1%] vs 57.0% [15.2%-95.0%]; P = .016). The duty cycle and the rate of incomplete recoil were comparable between the 2 groups. The angles between the arm of the participants and the bed were more vertical in the inclined stool group (84.0° ± 5.2° vs 81.0° ± 4.8°; P = .014). Using an inclined stool resulted in an improvement in the depth of CC and the adequate CC rate without increasing the rate of incomplete chest recoil. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Stool patterns of Malaysian adults with functional constipation: association with diet and physical activity.

    Science.gov (United States)

    Mazlyn, Mena M; Nagarajah, Lee H L; Fatimah, A; Norimah, A K; Goh, K L

    2013-04-01

    Diet and lifestyle modification is commonly used in constipation management. As there is a dearth of studies on this topic in Malaysia, we aim to elucidate the relations between stool patterns, dietary intake and physical activity levels among adults with functional constipation. From a database collected via surveys at public events, a convenience sample of 100 adults diagnosed with Rome II-defined functional constipation was enrolled in this cross-sectional study. After severity assessment using the Chinese Constipation Questionnaire, subjects completed 2-week bowel movement diaries to determine stool frequency, consistency and output. Dietary intake and physical activity levels were assessed twice using three-day 24-hour diet recalls and International Physical Activity Questionnaire, respectively. Ninety subjects who completed the study were included in the analysis. Mean weekly stool frequency was 3.9 +/- 1.9 times, consistency score was 2.6 +/- 0.6 (range 1.0-4.0), output was 11.0 +/- 6.3 balls (40 mm diameter) and severity score was 10.3 +/- 3.3 (range 5.0-22.0). Mean daily dietary intakes were: energy 1,719 +/- 427kcal, dietary fibre 15.0 +/- 4.9g and fluid 2.5 +/- 0.8L. The majority of subjects were physically inactive. Stool frequency and output were positively associated with dietary fibre (r(s) = 0.278, P < 0.01; r(s) = 0.226, P < 0.05) and fluid intake (r(s) = 0.257, P < 0.05; OR = 3.571, 95% CI [1.202-10.609]). Constipation severity was associated with higher physical activity levels (OR = 2.467, 95% CI [1.054-5.777]). Insufficient intake of dietary fibre and fluid are associated with aggravated constipation symptoms. Further studies are necessary to confirm usefulness of dietary intervention in treatment of constipation as dietary factors alone may not influence overall severity and stool consistency, an integral element of constipation.

  2. Is PCR the Next Reference Standard for the Diagnosis of Schistosoma in Stool? A Comparison with Microscopy in Senegal and Kenya.

    NARCIS (Netherlands)

    Meurs, L.; Brienen, E.; Mbow, M.; Ochola, E.A,; Mboup, S.; Karanja, D.M.; Secor, W.E.; Polman, K.; Lieshout, L.

    2015-01-01

    Background The current reference test for the detection of S. mansoni in endemic areas is stool microscopy based on one or more Kato-Katz stool smears. However, stool microscopy has several shortcomings that greatly affect the efficacy of current schistosomiasis control programs. A highly specific

  3. Magnetic resonance colonography without bowel cleansing using oral and rectal stool softeners (fecal cracking) - a feasibility study

    International Nuclear Information System (INIS)

    Ajaj, Waleed; Lauenstein, Thomas C.; Kuehle, Christiane; Herborn, Christoph U.; Goehde, Susanne C.; Schneemann, Hubert; Ruehm, Stefan G.; Goyen, Mathias

    2005-01-01

    The aim of our study was to assess the effect of oral and rectal stool softeners on dark-lumen magnetic resonance (MR) colonography without bowel cleansing. Ten volunteers underwent MR colonography without colonic cleansing. A baseline examination was performed without oral or rectal administration of stool softeners. In a second set, volunteers ingested 60 ml of lactulose 24 h prior to MR examination. In a third examination, water as a rectal enema was replaced by a solution of 0.5%-docusate sodium (DS). A fourth MR examination was performed, in conjunction with both oral administration of lactulose and rectal application of DS. A T1-weighted data set was acquired at scanning times of 0, 5 and 10 min after colonic filling. A fourth data set was acquired 75 s after i.v. injection of contrast agent. Signal intensity of stool was calculated for all colonic segments. Without oral ingestion of lactulose or rectal enema with DS stool signal intensity was high and did not decrease over time. However, lactulose and DS caused a decrease in stool signal intensity. Both substances together led to a decreasing signal intensity of feces. Combination of lactulose and DS provided the lowest signal intensity of stool. Thus, feces could hardly be distinguished from dark rectal enema allowing for the assessment of the colonic wall. (orig.)

  4. Prevalence of Salmonella Excretion in Stool: A Community Survey in 2 Sites, Guinea-Bissau and Senegal.

    Science.gov (United States)

    Im, Justin; Nichols, Chelsea; Bjerregaard-Andersen, Morten; Sow, Amy Gassama; Løfberg, Sandra; Tall, Adama; Pak, Gi Deok; Aaby, Peter; Baker, Stephen; Clemens, John D; Espinoza, Ligia Maria Cruz; Konings, Frank; May, Jürgen; Monteiro, Mario; Niang, Aissatou; Panzner, Ursula; Park, Se Eun; Schütt-Gerowitt, Heidi; Wierzba, Thomas F; Marks, Florian; von Kalckreuth, Vera

    2016-03-15

    Chronic and convalescent carriers play an important role in the transmission and endemicity of many communicable diseases. A high incidence of Salmonella enterica serovar Typhi and invasive nontyphoidal Salmonella (NTS) infection has been reported in parts of sub-Saharan Africa, yet the prevalence of Salmonella excretion in the general population is unknown. Stool specimens were collected from a random sample of households in 2 populations in West Africa: Bissau, Guinea-Bissau, and Dakar, Senegal. Stool was cultured to detect presence of Salmonella, and antimicrobial susceptibility testing was performed on the isolated organisms. Stool was cultured from 1077 and 1359 individuals from Guinea-Bissau and Senegal, respectively. Salmonella Typhi was not isolated from stool samples at either site. Prevalence of NTS in stool samples was 24.1 (95% confidence interval [CI], 16.5-35.1; n = 26/1077) per 1000 population in Guinea-Bissau and 10.3 (95% CI, 6.1-17.2; n = 14/1359) per 1000 population in Senegal. Evidence of NTS excretion in stool in both study populations indicates a possible NTS transmission route in these settings. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  5. Albendazole Stimulates the Excretion of Strongyloides stercoralis Larvae in Stool Specimens and Enhances Sensitivity for Diagnosis of Strongyloidiasis▿

    Science.gov (United States)

    Anamnart, Witthaya; Pattanawongsa, Attarat; Intapan, Pewpan Maleewong; Maleewong, Wanchai

    2010-01-01

    We succeeded in stimulation of excretion of Strongyloides stercoralis larvae in stool by oral administration of a single dose of 400 mg albendazole to strongyloidiasis patients. This result overcame the false-negative results of stool examination due to low larval numbers. Stool samples were collected from 152 asymptomatic strongyloidiasis patients in the morning, prior to eating. After breakfast, they were given a dose of 400 mg albendazole, and stool samples were collected the following morning. Agar plate culture (APC), modified formalin-ether concentration technique (MFECT), and direct-smear (DS) methods were used to examine stool specimens within 3 h after defecation. The results before and after albendazole was taken were compared. All APCs that were positive became negative after albendazole administration, while MFECT showed a 1.4- to 18.0-fold increase in larval numbers in 97.4% (148/152) of the samples. The DSs were positive in 3 out of 3 smears at a larval number of ≥45 larvae per g (lpg) of stool, and in 1or 2 out of 3 smears at a larval number between 35 and 44 lpg. At a larval number of albendazole administration became positive by MFECT after the treatment. Thus, MFECT can be effectively used for diagnosis of strongyloidiasis with prior administration of albendazole to the subject. PMID:20844212

  6. First detection and genotyping of Giardia intestinalis in stool samples collected from children in Ghazni Province, eastern Afghanistan and evaluation of the PCR assay in formalin-fixed specimens.

    Science.gov (United States)

    Lass, Anna; Karanis, Panagiotis; Korzeniewski, Krzysztof

    2017-08-01

    It is estimated that faecal-orally transmitted diseases are common in Afghanistan, as a consequence of poor hygienic standards of life and widespread contamination of water and food with both human and animal faeces. However, there is little information in the literature concerning infections caused by intestinal parasites in the Afghan population. In this study, we report the occurrence of Giardia intestinalis assemblages (A and B) in formalin-fixed stool samples collected from 245 Afghan schoolchildren living in Ghazni Province in eastern Afghanistan. Detection of the parasite's DNA and genotyping was performed using real-time PCR, specific to the β-giardin gene of G. intestinalis. Positive results were recorded in 52 (21.2%) samples. Genotyping was successful in 39 faecal samples and showed the predominance of assemblage B of G. intestinalis in this population (15 assemblage A and 24 assemblage B). Co-infection with both genotypes A and B was detected in four samples. Additionally, we evaluated the effect of 10% buffered formalin fixative on the detection of G. intestinalis DNA using real-time PCR and nested PCR characterised by different lengths of PCR products (74 and 479 bp, respectively). The human faeces containing the Giardia cysts were tested for 16 weeks. Amplification of G. intestinalis DNA with real-time PCR was possible up to 6 weeks of preservation of stool sample in formalin, compared to only 2 weeks with nested PCR. This suggests that real-time PCR is a more suitable tool in cases where stool samples have to be kept in formalin for longer periods of time.

  7. Implementation of a Clinical Decision Support Tool for Stool Cultures and Parasitological Studies in Hospitalized Patients.

    Science.gov (United States)

    Nikolic, D; Richter, S S; Asamoto, K; Wyllie, R; Tuttle, R; Procop, G W

    2017-12-01

    There is substantial evidence that stool culture and parasitological examinations are of minimal to no value after 3 days of hospitalization. We implemented and studied the impact of a clinical decision support tool (CDST) to decrease the number of unnecessary stool cultures (STCUL), ova/parasite (O&P) examinations, and Giardia / Cryptosporidium enzyme immunoassay screens (GC-EIA) performed for patients hospitalized >3 days. We studied the frequency of stool studies ordered before or on day 3 and after day 3 of hospitalization (i.e., categorical orders/total number of orders) before and after this intervention and denoted the numbers and types of microorganisms detected within those time frames. This intervention, which corresponded to a custom-programmed hard-stop alert tool in the Epic hospital information system, allowed providers to override the intervention by calling the laboratory, if testing was deemed medically necessary. Comparative statistics were employed to determine significance, and cost savings were estimated based on our internal costs. Before the intervention, 129/670 (19.25%) O&P examinations, 47/204 (23.04%) GC-EIA, and 249/1,229 (20.26%) STCUL were ordered after 3 days of hospitalization. After the intervention, 46/521 (8.83%) O&P examinations, 27/157 (17.20%) GC-EIA, and 106/1,028 (10.31%) STCUL were ordered after 3 days of hospitalization. The proportions of reductions in the number of tests performed after 3 days and the associated P values were 54.1% for O&P examinations ( P < 0.0001), 22.58% for GC-EIA ( P = 0.2807), and 49.1% for STCUL ( P < 0.0001). This was estimated to have resulted in $8,108.84 of cost savings. The electronic CDST resulted in a substantial reduction in the number of evaluations of stool cultures and the number of parasitological examinations for patients hospitalized for more than 3 days and in a cost savings while retaining the ability of the clinician to obtain these tests if clinically indicated. Copyright © 2017

  8. Helicobacter pylori stool antigen test (HpSA) for the diagnosis of gastric infection

    International Nuclear Information System (INIS)

    Faruqui, A.N.; Majid, U.; Ahmed, L.; Khalil, M.; Hussan, M.U.

    2007-01-01

    To determine the accuracy of Helicobacter pylori Stool Antigen test (HpSA), compared with endoscopic histopathology for the diagnosis of gastric Helicobecter pylori infection. A total of 50 patients underwent endoscopy for gastric antral mucosal tissue biopsy for histopathology of H.pylori and advised for HpSA. Patient's information including age, gender, past history, presenting signs and symptoms, results of HpSA and histopathology were recorded. Sensitivity analysis was performed to calculate sensitivity, specificity, and predictive values of HpSA. Among 50 patients, 48% males and 52% females (M: F 1: 1.08), a total of 27 (54%) were true positive while 20 (40%) were true negative. Two patients were false negative and only one was false positive. Sensitivity of HpSA was, therefore, 93.1%, specificity 95% and positive and negative predictive values were 96.42% and 90.9% respectively. Helicobacter pylori stool antigen was an accurate and reliable test for the diagnosis of gastric H. pylori infection. (author)

  9. Diagnosis of human fascioliasis by stool and blood techniques: update for the present global scenario.

    Science.gov (United States)

    Mas-Coma, S; Bargues, M D; Valero, M A

    2014-12-01

    Before the 1990s, human fascioliasis diagnosis focused on individual patients in hospitals or health centres. Case reports were mainly from developed countries and usually concerned isolated human infection in animal endemic areas. From the mid-1990s onwards, due to the progressive description of human endemic areas and human infection reports in developing countries, but also new knowledge on clinical manifestations and pathology, new situations, hitherto neglected, entered in the global scenario. Human fascioliasis has proved to be pronouncedly more heterogeneous than previously thought, including different transmission patterns and epidemiological situations. Stool and blood techniques, the main tools for diagnosis in humans, have been improved for both patient and survey diagnosis. Present availabilities for human diagnosis are reviewed focusing on advantages and weaknesses, sample management, egg differentiation, qualitative and quantitative diagnosis, antibody and antigen detection, post-treatment monitoring and post-control surveillance. Main conclusions refer to the pronounced difficulties of diagnosing fascioliasis in humans given the different infection phases and parasite migration capacities, clinical heterogeneity, immunological complexity, different epidemiological situations and transmission patterns, the lack of a diagnostic technique covering all needs and situations, and the advisability for a combined use of different techniques, at least including a stool technique and a blood technique.

  10. Oral Supplementation with Bovine Colostrum Decreases Intestinal Permeability and Stool Concentrations of Zonulin in Athletes.

    Science.gov (United States)

    Hałasa, Maciej; Maciejewska, Dominika; Baśkiewicz-Hałasa, Magdalena; Machaliński, Bogusław; Safranow, Krzysztof; Stachowska, Ewa

    2017-04-08

    Increased intestinal permeability has been implicated in various pathologies, has various causes, and can develop during vigorous athletic training. Colostrum bovinum is a natural supplement with a wide range of supposed positive health effects, including reduction of intestine permeability. We assessed influence of colostrum supplementation on intestinal permeability related parameters in a group of 16 athletes during peak training for competition. This double-blind placebo-controlled study compared supplementation for 20 days with 500 mg of colostrum bovinum or placebo (whey). Gut permeability status was assayed by differential absorption of lactulose and mannitol (L/M test) and stool zonulin concentration. Baseline L/M tests found that six of the participants (75%) in the colostrum group had increased intestinal permeability. After supplementation, the test values were within the normal range and were significantly lower than at baseline. The colostrum group Δ values produced by comparing the post-intervention and baseline results were also significantly lower than the placebo group Δ values. The differences in stool zonulin concentration were smaller than those in the L/M test, but were significant when the Δ values due to intervention were compared between the colostrum group and the placebo group. Colostrum bovinum supplementation was safe and effective in decreasing of intestinal permeability in this series of athletes at increased risk of its elevation.

  11. Oral Supplementation with Bovine Colostrum Decreases Intestinal Permeability and Stool Concentrations of Zonulin in Athletes

    Directory of Open Access Journals (Sweden)

    Maciej Hałasa

    2017-04-01

    Full Text Available Increased intestinal permeability has been implicated in various pathologies, has various causes, and can develop during vigorous athletic training. Colostrum bovinum is a natural supplement with a wide range of supposed positive health effects, including reduction of intestine permeability. We assessed influence of colostrum supplementation on intestinal permeability related parameters in a group of 16 athletes during peak training for competition. This double-blind placebo-controlled study compared supplementation for 20 days with 500 mg of colostrum bovinum or placebo (whey. Gut permeability status was assayed by differential absorption of lactulose and mannitol (L/M test and stool zonulin concentration. Baseline L/M tests found that six of the participants (75% in the colostrum group had increased intestinal permeability. After supplementation, the test values were within the normal range and were significantly lower than at baseline. The colostrum group Δ values produced by comparing the post-intervention and baseline results were also significantly lower than the placebo group Δ values. The differences in stool zonulin concentration were smaller than those in the L/M test, but were significant when the Δ values due to intervention were compared between the colostrum group and the placebo group. Colostrum bovinum supplementation was safe and effective in decreasing of intestinal permeability in this series of athletes at increased risk of its elevation.

  12. The effect of step stool use and provider height on CPR quality during pediatric cardiac arrest: A simulation-based multicentre study.

    Science.gov (United States)

    Cheng, Adam; Lin, Yiqun; Nadkarni, Vinay; Wan, Brandi; Duff, Jonathan; Brown, Linda; Bhanji, Farhan; Kessler, David; Tofil, Nancy; Hecker, Kent; Hunt, Elizabeth A

    2018-01-01

    We aimed to explore whether a) step stool use is associated with improved cardiopulmonary resuscitation (CPR) quality; b) provider adjusted height is associated with improved CPR quality; and if associations exist, c) determine whether just-in-time (JIT) CPR training and/or CPR visual feedback attenuates the effect of height and/or step stool use on CPR quality. We analysed data from a trial of simulated cardiac arrests with three study arms: No intervention; CPR visual feedback; and JIT CPR training. Step stool use was voluntary. We explored the association between 1) step stool use and CPR quality, and 2) provider adjusted height and CPR quality. Adjusted height was defined as provider height + 23 cm (if step stool was used). Below-average height participants were ≤ gender-specific average height; the remainder were above average height. We assessed for interaction between study arm and both adjusted height and step stool use. One hundred twenty-four subjects participated; 1,230 30-second epochs of CPR were analysed. Step stool use was associated with improved compression depth in below-average (female, p=0.007; male, pstep stool use (pStep stool use is associated with improved compression depth regardless of height. Increased provider height is associated with improved compression depth, with visual feedback attenuating the effects of height and step stool use.

  13. Ancient DNA

    DEFF Research Database (Denmark)

    Willerslev, Eske; Cooper, Alan

    2004-01-01

    ancient DNA, palaeontology, palaeoecology, archaeology, population genetics, DNA damage and repair......ancient DNA, palaeontology, palaeoecology, archaeology, population genetics, DNA damage and repair...

  14. Dual-energy CT characteristics of colon and rectal cancer allows differentiation from stool by dual-source CT.

    Science.gov (United States)

    Özdeniz, İlknur; İdilman, İlkay S; Köklü, Seyfettin; Hamaloğlu, Erhan; Özmen, Mustafa; Akata, Deniz; Karçaaltıncaba, Muşturay

    2017-01-01

    We aimed to determine dual-energy computed tomography (DECT) characteristics of colorectal cancer and investigate effectiveness of DECT method in differentiating tumor from stool in patients with colorectal cancer. Fifty consecutive patients with colorectal tumors were enrolled. Staging was performed by DECT (80-140 kV) using dual-source CT after rectal air insufflation and without bowel preparation. Both visual and quantitative analyses were performed at 80 kV and 140 kV, on iodine map and virtual noncontrast (VNC) images. All colorectal tumors had homogeneous pattern on iodine map. Stools demonstrated heterogeneous pattern in 86% (43/50) and homogeneous pattern in 14% (7/50) on iodine maps and were less visible on VNC images. Median density of tumors was 54 HU (18-100 HU) on iodine map and 28 HU (11-56 HU) on VNC images. Median density of stool was 36.5 HU (8-165 HU) on iodine map and -135.5 HU (-438 HU to -13 HU) on VNC images. The density of stools was significantly lower than tumors on both iodine map and VNC images (P VNC images was -1 HU with area under the curve of 1 and a sensitivity and specificity of 100%. Density or visual analysis of iodine map and VNC DECT images allow accurate differentiation of tumor from stool.

  15. Parasites in stool samples in the environment of Ilha da Marambaia, Rio de Janeiro, Brazil: an approach in public health

    Directory of Open Access Journals (Sweden)

    Beatriz Coronato

    2012-04-01

    Full Text Available This research aimed to describe the frequency of parasites in stool samples in the environment of Ilha da Marambaia, Rio de Janeiro, Brazil. One hundred and five stool samples were collected and processed by the coproparasitological techniques ethyl acetate sedimentation and centrifuge-flotation using saturated sugar solution. Parasites were detected in 81.9% of the samples, hookworm being the most prevalent, followed by Trichuris vulpis. Ascaris sp. eggs were also found. A high level of evolutive forms of parasites with public health risk was found in stool samples of the environment studied. We propose that health education programs, allied to an improvement of human and animal health care, must be employed to reduce the environmental contamination.

  16. Preanalytical blood sample workup for cell-free DNA analysis using Droplet Digital PCR for future molecular cancer diagnostics.

    Science.gov (United States)

    van Ginkel, Joost H; van den Broek, Daan A; van Kuik, Joyce; Linders, Dorothé; de Weger, Roel; Willems, Stefan M; Huibers, Manon M H

    2017-10-01

    In current molecular cancer diagnostics, using blood samples of cancer patients for the detection of genetic alterations in plasma (cell-free) circulating tumor DNA (ctDNA) is an emerging practice. Since ctDNA levels in blood are low, highly sensitive Droplet Digital PCR (ddPCR) can be used for detecting rare mutational targets. In order to perform ddPCR on blood samples, a standardized procedure for processing and analyzing blood samples is necessary to facilitate implementation into clinical practice. Therefore, we assessed the technical sample workup procedure for ddPCR on blood plasma samples. Blood samples from healthy individuals, as well as lung cancer patients were analyzed. We compared different methods and protocols for sample collection, storage, centrifugation, isolation, and quantification. Cell-free DNA (cfDNA) concentrations of several wild-type targets and BRAF and EGFR-mutant ctDNA concentrations quantified by ddPCR were primary outcome measurements. Highest cfDNA concentrations were measured in blood collected in serum tubes. No significant differences in cfDNA concentrations were detected between various time points of up to 24 h until centrifugation. Highest cfDNA concentrations were detected after DNA isolation with the Quick cfDNA Serum & Plasma Kit, while plasma isolation using the QIAamp Circulating Nucleic Acid Kit yielded the most consistent results. DdPCR results on cfDNA are highly dependent on multiple factors during preanalytical sample workup, which need to be addressed during the development of this diagnostic tool for cancer diagnostics in the future. © 2017 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  17. Evaluation of methods to improve the extraction and recovery of DNA from cotton swabs for forensic analysis.

    Science.gov (United States)

    Adamowicz, Michael S; Stasulli, Dominique M; Sobestanovich, Emily M; Bille, Todd W

    2014-01-01

    Samples for forensic DNA analysis are often collected from a wide variety of objects using cotton or nylon tipped swabs. Testing has shown that significant quantities of DNA are retained on the swab, however, and subsequently lost. When processing evidentiary samples, the recovery of the maximum amount of available DNA is critical, potentially dictating whether a usable profile can be derived from a piece of evidence or not. The QIAamp DNA Investigator extraction kit was used with its recommended protocol for swabs (one hour incubation at 56°C) as a baseline. Results indicate that over 50% of the recoverable DNA may be retained on the cotton swab tip, or otherwise lost, for both blood and buccal cell samples when using this protocol. The protocol's incubation time and temperature were altered, as was incubating while shaking or stationary to test for increases in recovery efficiency. An additional step was then tested that included periodic re-suspension of the swab tip in the extraction buffer during incubation. Aliquots of liquid blood or a buccal cell suspension were deposited and dried on cotton swabs and compared with swab-less controls. The concentration of DNA in each extract was quantified and STR analysis was performed to assess the quality of the extracted DNA. Stationary incubations and those performed at 65°C did not result in significant gains in DNA yield. Samples incubated for 24 hours yielded less DNA. Increased yields were observed with three and 18 hour incubation periods. Increases in DNA yields were also observed using a swab re-suspension method for both cell types. The swab re-suspension method yielded an average two-fold increase in recovered DNA yield with buccal cells and an average three-fold increase with blood cells. These findings demonstrate that more of the DNA collected on swabs can be recovered with specific protocol alterations.

  18. Evaluation of methods to improve the extraction and recovery of DNA from cotton swabs for forensic analysis.

    Directory of Open Access Journals (Sweden)

    Michael S Adamowicz

    Full Text Available Samples for forensic DNA analysis are often collected from a wide variety of objects using cotton or nylon tipped swabs. Testing has shown that significant quantities of DNA are retained on the swab, however, and subsequently lost. When processing evidentiary samples, the recovery of the maximum amount of available DNA is critical, potentially dictating whether a usable profile can be derived from a piece of evidence or not. The QIAamp DNA Investigator extraction kit was used with its recommended protocol for swabs (one hour incubation at 56°C as a baseline. Results indicate that over 50% of the recoverable DNA may be retained on the cotton swab tip, or otherwise lost, for both blood and buccal cell samples when using this protocol. The protocol's incubation time and temperature were altered, as was incubating while shaking or stationary to test for increases in recovery efficiency. An additional step was then tested that included periodic re-suspension of the swab tip in the extraction buffer during incubation. Aliquots of liquid blood or a buccal cell suspension were deposited and dried on cotton swabs and compared with swab-less controls. The concentration of DNA in each extract was quantified and STR analysis was performed to assess the quality of the extracted DNA. Stationary incubations and those performed at 65°C did not result in significant gains in DNA yield. Samples incubated for 24 hours yielded less DNA. Increased yields were observed with three and 18 hour incubation periods. Increases in DNA yields were also observed using a swab re-suspension method for both cell types. The swab re-suspension method yielded an average two-fold increase in recovered DNA yield with buccal cells and an average three-fold increase with blood cells. These findings demonstrate that more of the DNA collected on swabs can be recovered with specific protocol alterations.

  19. Effect of room temperature transport vials on DNA quality and phylogenetic composition of faecal microbiota of elderly adults and infants.

    LENUS (Irish Health Repository)

    Hill, Cian J

    2016-05-01

    Alterations in intestinal microbiota have been correlated with a growing number of diseases. Investigating the faecal microbiota is widely used as a non-invasive and ethically simple proxy for intestinal biopsies. There is an urgent need for collection and transport media that would allow faecal sampling at distance from the processing laboratory, obviating the need for same-day DNA extraction recommended by previous studies of freezing and processing methods for stool. We compared the faecal bacterial DNA quality and apparent phylogenetic composition derived using a commercial kit for stool storage and transport (DNA Genotek OMNIgene GUT) with that of freshly extracted samples, 22 from infants and 20 from older adults.

  20. Validation of a simple stool diary used by caregivers to document diarrhea among young children in a low-income country

    DEFF Research Database (Denmark)

    Grenov, Benedikte; Namusoke, Hanifa; Nabukeera-Barungi, Nicolette

    2017-01-01

    OBJECTIVES: Development and validation of a simple stool diary for caretakers collecting data on stool frequency and consistency among young children in a low-income country. METHODS: Focus group studies evaluated how diarrhea was understood by caregivers (content validity). The sensitivity......, reliability, and correlations between dehydration and diary scores (construct validity) were tested in a clinical trial. RESULTS: Caregivers recognized and understood the concept and severity of diarrhea. Stool frequency and liquid consistency decreased in children admitted with diarrhea (p 

  1. Dietary shift and dysbiosis may trigger mucous stools in giant pandas (Ailuropoda melanoleuca

    Directory of Open Access Journals (Sweden)

    Candace L Williams

    2016-05-01

    Full Text Available Dietary shifts can result in dysbiosis between the host and its gastrointestinal tract (GIT microbiota, leading to negative outcomes including inflammation. Giant pandas (Ailuropoda melanoleuca are physiologically classified as carnivores; however, they consume a herbivorous diet with dramatic seasonal feeding shifts and episodes of chronic GIT distress with symptoms including abdominal pain, loss of appetite and the excretion of mucous stools (mucoids. These episodes adversely affect the overall nutritional and health status of giant pandas. Here, we examined the fecal microbiota of two giant pandas’ normal and mucoid stools and compared these microbiota to baseline samples from a season with historically few episodes. To identify the microbiota present, we isolated and sequenced 16S rRNA using next-generation sequencing. Mucoids occurred following a seasonal feeding switch from predominately bamboo culm (stalk to leaves. All fecal samples displayed low diversity and were dominated by bacterial in the phyla Firmicutes and to a lesser extent, the Proteobacteria. Fecal samples immediately prior to mucoid episodes had lower microbial diversity compared to baseline samples, followed by increased diversity in mucoids. Mucoids were mostly comprised of common mucosal-associated taxa including Streptococcus and Leuconostoc species, and exhibited increased abundance for bacteria in the family Pasteurellaceae. Taken together, these findings indicate that diet-induced intestinal dysbiosis in giant pandas likely results in an expulsion of the mucosal lining in the form of mucoids. We suggest that these occurrences serve to reset their GIT microbiota, as giant pandas have retained a carnivorous GIT anatomy while shifting to an herbivorous diet.

  2. Repeat Rifaximin for Irritable Bowel Syndrome: No Clinically Significant Changes in Stool Microbial Antibiotic Sensitivity.

    Science.gov (United States)

    Pimentel, M; Cash, B D; Lembo, A; Wolf, R A; Israel, R J; Schoenfeld, P

    2017-09-01

    Rifaximin has demonstrated efficacy and safety for diarrhea-predominant irritable bowel syndrome (IBS-D). To determine the rifaximin repeat treatment effect on fecal bacterial antibiotic susceptibility. Patients with IBS in Trial 3 (TARGET 3) study who responded to open-label rifaximin 550 mg three times daily for 2 weeks, with symptom recurrence within 18 weeks, were randomized to double-blind treatment: two 2-week repeat courses of rifaximin or placebo, separated by 10 weeks. Prospective stool sample collection occurred before and after open-label rifaximin, before and after the first repeat course, and at the end of the study. Susceptibility testing was performed with 11 antibiotics, including rifaximin and rifampin, using broth microdilution or agar dilution methods. Of 103 patients receiving open-label rifaximin, 73 received double-blind rifaximin (n = 37) or placebo (n = 36). A total of 1429 bacterial and yeast isolates were identified, of which Bacteroidaceae (36.7%) and Enterobacteriaceae (33.9%) were the most common. In the double-blind phase, Clostridium difficile was highly susceptible to rifaximin [minimum inhibitory concentration (MIC) range 0.008-1 µg/mL] and rifampin (MIC range 0.004-0.25 µg/mL). Following double-blind rifaximin treatment, Staphylococcus isolates remained susceptible to rifaximin at all visits (MIC 50 range ≤0.06-32 µg/mL). Rifaximin exposure was not associated with long-term cross-resistance of Bacteroidaceae, Enterobacteriaceae, and Enterococcaceae to rifampin or nonrifamycin antibiotics tested. In this study, short-term repeat treatment with rifaximin has no apparent long-term effect on stool microbial susceptibility to rifaximin, rifampin, and nonrifamycin antibiotics. CLINICALTRIALS. NCT01543178.

  3. Morphologic changes of the anal sphincter musculature during and after temporary stool deviation.

    Science.gov (United States)

    Sailer, M; Fein, M; Fuchs, K H; Bussen, D; Grun, C; Thiede, A

    2001-04-01

    Temporary stool deviation, using a stoma, is a well-known surgical principle to protect low colorectal or coloanal anastomoses. The purpose of this study was to evaluate any morphologic changes with regard to the anal sphincter muscles during and after temporary ileostomy. Forty-four patients with rectal carcinomas were studied prospectively. All patients underwent low anterior resection. Reconstruction was performed using either a coloanal pouch or a straight end-to-end anastomosis. A protective stoma was fashioned in all 44 patients (ileostomy n=41; colostomy n=3). Stoma closure was carried out after a median of 85 days (41-330 days). Using a standard protocol, anal-sphincter thickness [m. puborectalis, external anal sphincter (EAS) and internal anal (IAS) sphincter] was assessed by means of endoanal ultrasonography preoperatively, at the time of stoma closure, and every 3 months thereafter for 1 year. The diameter of the puborectal muscle decreased from a median preoperative value of 6.3 mm to 5.7 mm at the time of stoma closure (P=0.03). After 3 months, 6.2 mm was measured. This value remained stable for the complete follow-up period. Similar results were recorded for the EAS. The IAS thickness remained stable throughout the study period, measuring between 2.1 mm and 2.4 mm. Temporary stool deviation does lead to morphologic changes of the anal sphincter. While the smooth muscle remains unchanged, the striated counterpart undergoes atrophic transformation. However, after passage reconstruction, i.e., stoma closure, a rapid regeneration of the voluntary muscles is observed.

  4. Freezing fecal samples prior to DNA extraction affects the Firmicutes to Bacteroidetes ratio determined by downstream quantitative PCR analysis

    DEFF Research Database (Denmark)

    Bahl, Martin Iain; Bergström, Anders; Licht, Tine Rask

    2012-01-01

    Freezing stool samples prior to DNA extraction and downstream analysis is widely used in metagenomic studies of the human microbiota but may affect the inferred community composition. In this study, DNA was extracted either directly or following freeze storage of three homogenized human fecal...

  5. Freezing fecal samples prior to DNA extraction affects the Firmicutes to Bacteroidetes ratio determined by downstream quantitative PCR analysis

    DEFF Research Database (Denmark)

    Bahl, Martin Iain; Bergström, Anders; Licht, Tine Rask

    Freezing stool samples prior to DNA extraction and downstream analysis is widely used in metagenomic studies of the human microbiota but may affect the inferred community composition. In this study DNA was extracted either directly or following freeze storage of three homogenized human fecal...

  6. Comparison of two commercial DNA extraction kits for the analysis of nasopharyngeal bacterial communities

    Directory of Open Access Journals (Sweden)

    Keith A. Crandall

    2016-04-01

    Full Text Available Characterization of microbial communities via next-generation sequencing (NGS requires an extraction ofmicrobial DNA. Methodological differences in DNA extraction protocols may bias results and complicate inter-study comparisons. Here we compare the effect of two commonly used commercial kits (Norgen and Qiagenfor the extraction of total DNA on estimatingnasopharyngeal microbiome diversity. The nasopharynxis a reservoir for pathogens associated with respiratory illnesses and a key player in understandingairway microbial dynamics. Total DNA from nasal washes corresponding to 30 asthmatic children was extracted using theQiagenQIAamp DNA and NorgenRNA/DNA Purification kits and analyzed via IlluminaMiSeq16S rRNA V4 ampliconsequencing. The Norgen samples included more sequence reads and OTUs per sample than the Qiagen samples, but OTU counts per sample varied proportionallybetween groups (r = 0.732.Microbial profiles varied slightly between sample pairs, but alpha- and beta-diversity indices (PCoAand clustering showed highsimilarity between Norgen and Qiagenmicrobiomes. Moreover, no significant differences in community structure (PERMANOVA and adonis tests and taxa proportions (Kruskal-Wallis test were observed betweenkits. Finally, aProcrustes analysis also showed low dissimilarity (M2 = 0.173; P< 0.001 between the PCoAs of the two DNA extraction kits. Contrary to what has been observed in previous studies comparing DNA extraction methods, our 16S NGS analysis of nasopharyngeal washes did not reveal significant differences in community composition or structure between kits. Our findingssuggest congruence between column-based chromatography kits and supportthe comparison of microbiomeprofilesacross nasopharyngeal metataxonomic studies.

  7. Effective testing for pulmonary tuberculosis using Xpert MTB/RIF assay for stool specimens in immunocompetent Pakistani children

    Directory of Open Access Journals (Sweden)

    Zahra Hasan

    2016-01-01

    Conclusion: Use of Xpert MTB/RIF assay for stool-based diagnosis of pulmonary TB in immunocompetent children is useful in a resource poor setting. This is a valuable and noninvasive diagnostic alternative for the diagnosis of childhood TB and can be adapted by pediatric arms of national TB programs.

  8. Evaluation of stool microbiota signatures in two cohorts of Asian (Singapore and Indonesia newborns at risk of atopy

    Directory of Open Access Journals (Sweden)

    Chua Kaw Yan

    2011-08-01

    Full Text Available Abstract Background Studies have suggested that demographic and lifestyle factors could shape the composition of fecal microbiota in early life. This study evaluated infant stool microbiota signatures in two Asian populations, Singapore (n = 42 and Indonesia (n = 32 with contrasting socioeconomic development, and examined the putative influences of demographic factors on these human fecal associated bacterial signatures. Results Longitudinal analysis showed associations of geographical origin with Clostridium leptum, Atopobium and Bifidobacterium groups. Mode of delivery had the largest effect on stool microbiota signatures influencing the abundance of four bacterial groups. Significantly higher abundance of bacterial members belonging to the Bacteroides-Prevotella, Bifidobacterium and Atopobium groups, but lower abundance of Lactobacilli-Enterococci group members, were observed in vaginal delivered compared to caesarean delivered infants. Demographic factors influencing the structure of infants stool microbiota during the first year of life included breastfeeding, age of weaning, sibship size and exposure to antibiotics. Conclusions Differences in stool microbiota signatures were observed in relation to various demographic factors. These features may confound studies relating to the association of the structure of fecal microbiota and the predisposition to human modern disease.

  9. The accuracy of the Helicobacter pylori stool antigen test in diagnosing H-pylori in treated and untreated patients

    NARCIS (Netherlands)

    Arents, NL; van Zwet, AA; Thijs, JC; de Jong, A; Pool, MO; Kleibeuker, JH

    Objective and design To evaluate the performance of the Helicobacter pylori stool antigen test (HpSA test) in detecting H. pylori infection and monitoring the effect of treatment. This was done in two separate studies using either a biopsy or the C-13-urea breath test based 'gold standard' (in

  10. Helicobacter pylori Stool Antigen test: a reliable non-invasive test for the diagnosis of Helicobacter pylori infection in children

    NARCIS (Netherlands)

    van Doorn, O. J.; Bosman, D. K.; van't Hoff, B. W.; Taminiau, J. A.; ten Kate, F. J.; van der Ende, A.

    2001-01-01

    OBJECTIVE: To evaluate the Helicobacter pylori Stool Antigen (HpSA) test for the diagnosis of H. pylori infection in children. DESIGN AND SETTING: Prospective cohort study in an academic medical centre. PATIENTS AND METHODS: A total of 106 consecutive children who underwent gastroscopy were

  11. Evaluation of BBL™ Sensi-Discs™ and FTA® cards as sampling devices for detection of rotavirus in stool samples.

    Science.gov (United States)

    Tam, Ka Ian; Esona, Mathew D; Williams, Alice; Ndze, Valantine N; Boula, Angeline; Bowen, Michael D

    2015-09-15

    Rotavirus is the most important cause of severe childhood gastroenteritis worldwide. Rotavirus vaccines are available and rotavirus surveillance is carried out to assess vaccination impact. In surveillance studies, stool samples are stored typically at 4°C or frozen to maintain sample quality. Uninterrupted cold storage is a problem in developing countries because of power interruptions. Cold-chain transportation of samples from collection sites to testing laboratories is costly. In this study, we evaluated the use of BBL™ Sensi-Discs™ and FTA(®) cards for storage and transportation of samples for virus isolation, EIA, and RT-PCR testing. Infectious rotavirus was recovered after 30 days of storage on Sensi-Discs™ at room temperature. We were able to genotype 98-99% of samples stored on Sensi-Discs™ and FTA(®) cards at temperatures ranging from -80°C to 37°C up to 180 days. A field sampling test using samples prepared and shipped from Cameroon, showed that both matrices yielded 100% genotyping success compared with whole stool and Sensi-Discs™ demonstrated 95% concordance with whole stool in EIA testing. The utilization of BBL™ Sensi-Discs™ and FTA(®) cards for stool sample storage and shipment has the potential to have great impact on global public health by facilitating surveillance and epidemiological investigations of rotavirus strains worldwide at a reduced cost. Published by Elsevier B.V.

  12. Evaluation of BBL™ Sensi-Discs™ and FTA® cards as sampling devices for detection of rotavirus in stool samples

    Science.gov (United States)

    Tam, Ka Ian; Esona, Mathew D.; Williams, Alice; Ndze, Valentine N.; Boula, Angeline; Bowen, Michael D.

    2015-01-01

    Rotavirus is the most important cause of severe childhood gastroenteritis worldwide. Rotavirus vaccines are available and rotavirus surveillance is carried out to assess vaccination impact. In surveillance studies, stool samples are stored typically at 4°C or frozen to maintain sample quality. Uninterrupted cold storage is a problem in developing countries because of power interruptions. Cold-chain transportation of samples from collection sites to testing laboratories is costly. In this study, we evaluated the use of BBL™ Sensi-Discs™ and FTA® cards for storage and transportation of samples for virus isolation, EIA, and RT-PCR testing. Infectious rotavirus was recovered after 30 days of storage on Sensi-Discs™ at room temperature. We were able to genotype 98–99% of samples stored on Sensi-Discs™ and FTA® cards at temperatures ranging from −80°C to 37°C up to 180 days. A field sampling test using samples prepared and shipped from Cameroon, showed that both matrices yielded 100% genotyping success compared with whole stool and Sensi-Discs™ demonstrated 95% concordance with whole stool in EIA testing. The utilization of BBL™ Sensi-Discs™ and FTA® cards for stool sample storage and shipment has the potential to have great impact on global public health by facilitating surveillance and epidemiological investigations of rotavirus strains worldwide at a reduced cost. PMID:26022083

  13. Performance Characteristics and Comparison of Abbott and artus Real-Time Systems for Hepatitis B Virus DNA Quantification ▿

    Science.gov (United States)

    Ismail, Ashrafali M.; Sivakumar, Jayashree; Anantharam, Raghavendran; Dayalan, Sujitha; Samuel, Prasanna; Fletcher, Gnanadurai J.; Gnanamony, Manu; Abraham, Priya

    2011-01-01

    Virological monitoring of hepatitis B virus (HBV) DNA is critical to the management of HBV infection. With several HBV DNA quantification assays available, it is important to use the most efficient testing system for virological monitoring. In this study, we evaluated the performance characteristics and comparability of three HBV DNA quantification systems: Abbott HBV real-time PCR (Abbott PCR), artus HBV real-time PCR with QIAamp DNA blood kit purification (artus-DB), and artus HBV real-time PCR with the QIAamp DSP virus kit purification (artus-DSP). The lower limits of detection of these systems were established against the WHO international standards for HBV DNA and were found to be 1.43, 82, and 9 IU/ml, respectively. The intra-assay and interassay coefficients of variation of plasma samples (1 to 6 log10 IU/ml) ranged between 0.05 to 8.34% and 0.16 to 3.48% for the Abbott PCR, 1.53 to 26.85% and 0.50 to 12.89% for artus-DB, and 0.29 to 7.42% and 0.94 to 3.01% for artus-DSP, respectively. Ninety HBV clinical samples were used for comparison of assays, and paired quantitative results showed strong correlation by linear regression analysis (artus-DB with Abbott PCR, r = 0.95; Abbott PCR with artus-DSP, r = 0.97; and artus-DSP with artus-DB, r = 0.94). Bland-Altman analysis showed a good level of agreement for Abbott PCR and artus-DSP, with a mean difference of 0.10 log10 IU/ml and limits of agreement of −0.91 to 1.11 log10 IU/ml. No genotype-specific bias was seen in all three systems for HBV genotypes A, C, and D, which are predominant in this region. This finding illustrates that the Abbott real-time HBV and artus-DSP systems show more comparable performance than the artus-DB system, meeting the current guidelines for assays to be used in the management of hepatitis B. PMID:21795507

  14. Performance characteristics and comparison of Abbott and artus real-time systems for hepatitis B virus DNA quantification.

    Science.gov (United States)

    Ismail, Ashrafali M; Sivakumar, Jayashree; Anantharam, Raghavendran; Dayalan, Sujitha; Samuel, Prasanna; Fletcher, Gnanadurai J; Gnanamony, Manu; Abraham, Priya

    2011-09-01

    Virological monitoring of hepatitis B virus (HBV) DNA is critical to the management of HBV infection. With several HBV DNA quantification assays available, it is important to use the most efficient testing system for virological monitoring. In this study, we evaluated the performance characteristics and comparability of three HBV DNA quantification systems: Abbott HBV real-time PCR (Abbott PCR), artus HBV real-time PCR with QIAamp DNA blood kit purification (artus-DB), and artus HBV real-time PCR with the QIAamp DSP virus kit purification (artus-DSP). The lower limits of detection of these systems were established against the WHO international standards for HBV DNA and were found to be 1.43, 82, and 9 IU/ml, respectively. The intra-assay and interassay coefficients of variation of plasma samples (1 to 6 log(10) IU/ml) ranged between 0.05 to 8.34% and 0.16 to 3.48% for the Abbott PCR, 1.53 to 26.85% and 0.50 to 12.89% for artus-DB, and 0.29 to 7.42% and 0.94 to 3.01% for artus-DSP, respectively. Ninety HBV clinical samples were used for comparison of assays, and paired quantitative results showed strong correlation by linear regression analysis (artus-DB with Abbott PCR, r = 0.95; Abbott PCR with artus-DSP, r = 0.97; and artus-DSP with artus-DB, r = 0.94). Bland-Altman analysis showed a good level of agreement for Abbott PCR and artus-DSP, with a mean difference of 0.10 log(10) IU/ml and limits of agreement of -0.91 to 1.11 log(10) IU/ml. No genotype-specific bias was seen in all three systems for HBV genotypes A, C, and D, which are predominant in this region. This finding illustrates that the Abbott real-time HBV and artus-DSP systems show more comparable performance than the artus-DB system, meeting the current guidelines for assays to be used in the management of hepatitis B.

  15. Detecting an elusive invasive species: a diagnostic PCR to detect Burmese python in Florida waters and an assessment of persistence of environmental DNA.

    Science.gov (United States)

    Piaggio, Antoinette J; Engeman, Richard M; Hopken, Matthew W; Humphrey, John S; Keacher, Kandy L; Bruce, William E; Avery, Michael L

    2014-03-01

    Recent studies have demonstrated that detection of environmental DNA (eDNA) from aquatic vertebrates in water bodies is possible. The Burmese python, Python bivittatus, is a semi-aquatic, invasive species in Florida where its elusive nature and cryptic coloration make its detection difficult. Our goal was to develop a diagnostic PCR to detect P. bivittatus from water-borne eDNA, which could assist managers in monitoring this invasive species. First, we used captive P. bivittatus to determine whether reptilian DNA could be isolated and amplified from water samples. We also evaluated the efficacy of two DNA isolation methods and two DNA extraction kits commonly used in eDNA preparation. A fragment of the mitochondrial cytochrome b gene from P. bivittatus was detected in all water samples isolated with the sodium acetate precipitate and the QIAamp DNA Micro Kit. Next, we designed P. bivittatus-specific primers and assessed the degradation rate of eDNA in water. Our primers did not amplify DNA from closely related species, and we found that P. bivittatus DNA was consistently detectable up to 96 h. Finally, we sampled water from six field sites in south Florida. Samples from five sites, where P. bivittatus has been observed, tested positive for eDNA. The final site was negative and had no prior documented evidence of P. bivittatus. This study shows P. bivittatus eDNA can be isolated from water samples; thus, this method is a new and promising technique for the management of invasive reptiles. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.

  16. Ancient DNA analyses of museum specimens from selected Presbytis (primate: Colobinae) based on partial Cyt b sequences

    Science.gov (United States)

    Aifat, N. R.; Yaakop, S.; Md-Zain, B. M.

    2016-11-01

    The IUCN Red List of Threatened Species has categorized Malaysian primates from being data deficient to critically endanger. Thus, ancient DNA analyses hold great potential to understand phylogeny, phylogeography and population history of extinct and extant species. Museum samples are one of the alternatives to provide important sources of biological materials for a large proportion of ancient DNA studies. In this study, a total of six museum skin samples from species Presbytis hosei (4 samples) and Presbytis frontata (2 samples), aged between 43 and 124 years old were extracted to obtain the DNA. Extraction was done by using QIAGEN QIAamp DNA Investigator Kit and the ability of this kit to extract museum skin samples was tested by amplification of partial Cyt b sequence using species-specific designed primer. Two primer pairs were designed specifically for P. hosei and P. frontata, respectively. These primer pairs proved to be efficient in amplifying 200bp of the targeted species in the optimized PCR conditions. The performance of the sequences were tested to determine genetic distance of genus Presbytis in Malaysia. From the analyses, P. hosei is closely related to P. chrysomelas and P. frontata with the value of 0.095 and 0.106, respectively. Cyt b gave a clear data in determining relationships among Bornean species. Thus, with the optimized condition, museum specimens can be used for molecular systematic studies of the Malaysian primates.

  17. gbpA as a Novel qPCR Target for the Species-Specific Detection of Vibrio cholerae O1, O139, Non-O1/Non-O139 in Environmental, Stool, and Historical Continuous Plankton Recorder Samples.

    Directory of Open Access Journals (Sweden)

    Luigi Vezzulli

    Full Text Available The Vibrio cholerae N-acetyl glucosamine-binding protein A (GbpA is a chitin-binding protein involved in V. cholerae attachment to environmental chitin surfaces and human intestinal cells. We previously investigated the distribution and genetic variations of gbpA in a large collection of V. cholerae strains and found that the gene is consistently present and highly conserved in this species. Primers and probe were designed from the gbpA sequence of V. cholerae and a new Taq-based qPCR protocol was developed for diagnostic detection and quantification of the bacterium in environmental and stool samples. In addition, the positions of primers targeting the gbpA gene region were selected to obtain a short amplified fragment of 206 bp and the protocol was optimized for the analysis of formalin-fixed samples, such as historical Continuous Plankton Recorder (CPR samples. Overall, the method is sensitive (50 gene copies, highly specific for V. cholerae and failed to amplify strains of the closely-related species Vibrio mimicus. The sensitivity of the assay applied to environmental and stool samples spiked with V. cholerae ATCC 39315 was comparable to that of pure cultures and was of 102 genomic units/l for drinking and seawater samples, 101 genomic units/g for sediment and 102 genomic units/g for bivalve and stool samples. The method also performs well when tested on artificially formalin-fixed and degraded genomic samples and was able to amplify V. cholerae DNA in historical CPR samples, the earliest of which date back to August 1966. The detection of V. cholerae in CPR samples collected in cholera endemic areas such as the Benguela Current Large Marine Ecosystem (BCLME is of particular significance and represents a proof of concept for the possible use of the CPR technology and the developed qPCR assay in cholera studies.

  18. Are bile acid malabsorption and bile acid diarrhoea important causes of loose stool complicating cancer therapy?

    Science.gov (United States)

    Phillips, F; Muls, A C G; Lalji, A; Andreyev, H J N

    2015-08-01

    Gastrointestinal (GI) symptoms during and after cancer therapy can significantly affect quality of life and interfere with treatment. This study assessed whether bile acid malabsorption (BAM) or bile acid diarrhoea (BAD) are important causes of diarrhoea associated with cancer treatment. A retrospective analysis was carried out of consecutive patients assessed for BAM using ((75) Se) Selenium homocholic acid taurocholate (SeHCAT) scanning, after reporting any episodes of loose stool, attending a gastroenterology clinic in a cancer centre. Between 2009 and 2013, 506 consecutive patients (54.5% male; age range: 20-91 years), were scanned. BAM/BAD was diagnosed in 215 (42.5%). It was mild in 25.6%, moderate in 29.3% and severe in 45.1%. Pelvic chemoradiation had induced BAM in > 50% of patients. BAM was also frequent after treatment for conditions not previously associated with BAM, such as anal and colorectal cancer, and was present in > 75% of patients referred after pancreatic surgery. It was also unexpectedly frequent in patients who were treated for malignancy outside the GI tract, such as breast cancer and haematological malignancy. BAM/BAD are very common and under-appreciated causes of GI symptoms after cancer treatment. Health professionals should have a low threshold in suspecting this condition, as diagnosis and treatment can significantly improve quality of life. Colorectal Disease © 2015 The Association of Coloproctology of Great Britain and Ireland.

  19. Performances of Four Helicobacter pylori Serological Detection Kits Using Stool Antigen Test as Gold Standard.

    Science.gov (United States)

    Biranjia-Hurdoyal, Susheela D; Seetulsingh-Goorah, Sharmila P

    2016-01-01

    The aim was to determine the performances of four Helicobacter pylori serological detection kits in different target groups, using Amplified IDEIA™ Hp StAR™ as gold standard. Kits studied were Rapid Immunochromatoghraphic Hexagon, Helicoblot 2.1, an EIA IgG kit and EIA IgA kit. Stool and blood samples were collected from 162 apparently healthy participants (control) and 60 Type 2 diabetes mellitus (T2DM) patients. The performances of the four serological detection kits were found to be affected by gender, age, health status and ethnicity of the participants. In the control group, the Helicoblot 2.1 kit had the best performance (AUC = 0.85; ppoor performances. In the T2DM subgroup, the kits H2.1 and EIA IgG had best performances, with accuracies of 96.5% and 93.1% respectively. The performance of EIA IgG improved with adjustment of its cut-off value. The performances of the detection kits were affected by various factors which should be taken into consideration.

  20. Performances of Four Helicobacter pylori Serological Detection Kits Using Stool Antigen Test as Gold Standard.

    Directory of Open Access Journals (Sweden)

    Susheela D Biranjia-Hurdoyal

    Full Text Available The aim was to determine the performances of four Helicobacter pylori serological detection kits in different target groups, using Amplified IDEIA™ Hp StAR™ as gold standard. Kits studied were Rapid Immunochromatoghraphic Hexagon, Helicoblot 2.1, an EIA IgG kit and EIA IgA kit.Stool and blood samples were collected from 162 apparently healthy participants (control and 60 Type 2 diabetes mellitus (T2DM patients.The performances of the four serological detection kits were found to be affected by gender, age, health status and ethnicity of the participants. In the control group, the Helicoblot 2.1 kit had the best performance (AUC = 0.85; p<0.05, accuracy = 86.4%, followed by EIA IgG (AUC = 0.75; p<0.05, accuracy = 75.2%. The Rapid Hexagon and EIA IgA kits had relatively poor performances. In the T2DM subgroup, the kits H2.1 and EIA IgG had best performances, with accuracies of 96.5% and 93.1% respectively. The performance of EIA IgG improved with adjustment of its cut-off value.The performances of the detection kits were affected by various factors which should be taken into consideration.

  1. Shotgun metagenomic data on the human stool samples to characterize shifts of the gut microbial profile after the Helicobacter pylori eradication therapy

    Directory of Open Access Journals (Sweden)

    Eugenia A. Boulygina

    2017-10-01

    Full Text Available The shotgun sequencing data presented in this report are related to the research article named “Gut microbiome shotgun sequencing in assessment of microbial community changes associated with H. pylori eradication therapy” (Khusnutdinova et al., 2016 [1]. Typically, the H. pylori eradication protocol includes a prolonged two-week use of the broad-spectrum antibiotics. The presented data on the whole-genome sequencing of the total DNA from stool samples of patients before the start of the eradication, immediately after eradication and several weeks after the end of treatment could help to profile the gut microbiota both taxonomically and functionally. The presented data together with those described in Glushchenko et al. (2017 [2] allow researchers to characterize the metagenomic profiles in which the use of antibiotics could result in dramatic changes in the intestinal microbiota composition. We perform 15 gut metagenomes from 5 patients with H. pylori infection, obtained through the shotgun sequencing on the SOLiD 5500 W platform. Raw reads are deposited in the ENA under project ID PRJEB21338.

  2. Evaluation of two methods DNA extraction from formalin-fixed, paraffin-embedded tissues on non-optimal conditions

    International Nuclear Information System (INIS)

    Bustamante, Javier Andres; Astudillo, Miryam; Pazos, Alvaro Jairo; Bravo, Luis Eduardo

    2011-01-01

    Paraffin wax embedded tissues are an invaluable material for retrospective studies requiring the application of molecular analysis. Multiple methods are available to extract DNA from these kinds of samples. However, the most common methods are slow and the reagents often contribute to the fragmentation of genetic material. In order to optimize the procedure, two methods for DNA extraction from paraffin embedded tissue non-optimal conditions were used. 47 blocks containing paraffin-embedded biopsies of pleura, lung and pericardium from 24 patients (66.6% males) older than 18 years, with biopsy proven chronic granulomatous inflammation referred to the department of pathology at University Hospital of Valle between 2002 and 2007 were selected. Each sample was subjected to 10 cuts and was to two methods of DNA extraction: 1. conventional and 2. QIAamp - DNA mini kit. The efficiency of the extracted DNA was assessed by spectrophotometry and PCR amplification of a fragment of the housekeeping gene GAPDH. The concentration of DNA samples extracted by the conventional method was of 65.52 ng/Mu l ± 11.47 (mean ± SE) and the 260/280 absorbance ratio ranged between 0.52 and 2.30 the average concentration of DNA of the samples extracted by the commercial method was 60.89 ng/Mu l ± 6.02 (mean ± SE), with an absorbance that fluctuated between 0 and 2.64. The DNA obtained was amplified by PCR, of 47 samples extracted by methods, 25 and 23 respectively the GAPDH gene amplified successfully. The methods used to obtain DNA showed similar performance, highlighting the potential utility of both extraction methods for the retrospective studies from paraffin embedded tissues in unsuitable conditions.

  3. Improved method for extraction and detection of Helicobacter pylori DNA in formalin-fixed paraffin embedded gastric biopsies using laser micro-dissection.

    Science.gov (United States)

    Loayza, María Fernanda; Villavicencio, Fernando Xavier; Santander, Stephanie Carolina; Baldeón, Manuel; Ponce, Lourdes Karina; Salvador, Iván; Vivar Díaz, Nicolás

    2015-01-01

    To assess the molecular events exerted by Helicobacter pylori interacting directly with gastric epithelial cells, an improved procedure for microbial DNA isolation from stained hematoxilin-eosin gastric biopsies was developed based on laser micro-dissection (LM) [1]. Few articles have described the use of LM to select and detect H. pylori genome from formalin-fixed paraffin embedded gastric tissue [2]. To improve the yield and quality of DNA isolated from H. pylori contacting intestinal epithelial cells, the following conditions were established after modification of the QIAamp DNA Micro kit. •Use of at least 25 cut sections of 10-20 μm of diameter and 3 μm thick with more than 10 bacteria in each cut.•Lysis with 30 μL of tissue lysis buffer and 20 μL of proteinase K (PK) with the tube in an upside-down position.•The use of thin purification columns with 35 μL of elution buffer. The mean of DNA concentration obtained from 25 LM cut sections was 1.94± 0 .16 ng/μL, and it was efficiently amplified with qPCR in a Bio Rad iCycler instrument. The LM can improve the sample selection and DNA extraction for molecular analysis of H. pylori associated with human gastric epithelium.

  4. Molecular detection of eukaryotes in a single human stool sample from Senegal.

    Directory of Open Access Journals (Sweden)

    Ibrahim Hamad

    Full Text Available BACKGROUND: Microbial eukaryotes represent an important component of the human gut microbiome, with different beneficial or harmful roles; some species are commensal or mutualistic, whereas others are opportunistic or parasitic. The diversity of eukaryotes inhabiting humans remains relatively unexplored because of either the low abundance of these organisms in human gut or because they have received limited attention from a whole-community perspective. METHODOLOGY/PRINCIPAL FINDING: In this study, a single fecal sample from a healthy African male was studied using both culture-dependent methods and extended molecular methods targeting the 18S rRNA and ITS sequences. Our results revealed that very few fungi, including Candida spp., Galactomyces spp., and Trichosporon asahii, could be isolated using culture-based methods. In contrast, a relatively a high number of eukaryotic species could be identified in this fecal sample when culture-independent methods based on various primer sets were used. A total of 27 species from one sample were found among the 977 analyzed clones. The clone libraries were dominated by fungi (716 clones/977, 73.3%, corresponding to 16 different species. In addition, 187 sequences out of 977 (19.2% corresponded to 9 different species of plants; 59 sequences (6% belonged to other micro-eukaryotes in the gut, including Entamoeba hartmanni and Blastocystis sp; and only 15 clones/977 (1.5% were related to human 18S rRNA sequences. CONCLUSION: Our results revealed a complex eukaryotic community in the volunteer's gut, with fungi being the most abundant species in the stool sample. Larger investigations are needed to assess the generality of these results and to understand their roles in human health and disease.

  5. Dipstick test for rapid diagnosis of Shigella dysenteriae 1 in bacterial cultures and its potential use on stool samples.

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    Neelam Taneja

    Full Text Available BACKGROUND: We describe a test for rapid detection of S. dysenteriae 1 in bacterial cultures and in stools, at the bedside of patients. METHODOLOGY/PRINCIPAL FINDINGS: The test is based on the detection of S. dysenteriae 1 lipopolysaccharide (LPS using serotype 1-specific monoclonal antibodies coupled to gold particles and displayed on a one-step immunochromatographic dipstick. A concentration as low as 15 ng/ml of LPS was detected in distilled water and in reconstituted stools in 10 minutes. In distilled water and in reconstituted stools, an unequivocal positive reaction was obtained with 1.6×10⁶ CFU/ml and 4.9×10⁶ CFU/ml of S. dysenteriae 1, respectively. Optimal conditions to read the test have been determined to limit the risk of ambiguous results due to appearance of a faint yellow test band in some negative samples. The specificity was 100% when tested with a battery of Shigella and unrelated strains in culture. When tested on 328 clinical samples in India, Vietnam, Senegal and France by laboratory technicians and in Democratic Republic of Congo by a field technician, the specificity (312/316 was 98.7% (95% CI:96.6-99.6% and the sensitivity (11/12 was 91.7% (95% CI:59.8-99.6%. Stool cultures and the immunochromatographic test showed concordant results in 98.4 % of cases (323/328 in comparative studies. Positive and negative predictive values were 73.3% (95% CI:44.8-91.1% and 99.7% (95% CI:98-100%. CONCLUSION: The initial findings presented here for a simple dipstick-based test to diagnose S. dysenteriae 1 demonstrates its promising potential to become a powerful tool for case management and epidemiological surveys.

  6. Impact of changing from staining to culture techniques on detection rates of Campylobacter spp. in routine stool samples in Chile.

    Science.gov (United States)

    Porte, Lorena; Varela, Carmen; Haecker, Thomas; Morales, Sara; Weitzel, Thomas

    2016-05-13

    Campylobacter is a leading cause of bacterial gastroenteritis, but sensitive diagnostic methods such as culture are expensive and often not available in resource limited settings. Therefore, direct staining techniques have been developed as a practical and economical alternative. We analyzed the impact of replacing Campylobacter staining with culture for routine stool examinations in a private hospital in Chile. From January to April 2014, a total of 750 consecutive stool samples were examined in parallel by Hucker stain and Campylobacter culture. Isolation rates of Campylobacter were determined and the performance of staining was evaluated against culture as the gold standard. Besides, isolation rates of Campylobacter and other enteric pathogens were compared to those of past years. Campylobacter was isolated by culture in 46 of 750 (6.1 %) stool samples. Direct staining only identified three samples as Campylobacter positive and reached sensitivity and specificity values of 6.5 and 100 %, respectively. In comparison to staining-based detection rates of previous years, we observed a significant increase of Campylobacter cases in our patients. Direct staining technique for Campylobacter had a very low sensitivity compared to culture. Staining methods might lead to a high rate of false negative results and an underestimation of the importance of campylobacteriosis. With the inclusion of Campylobacter culture, this pathogen became a leading cause of intestinal infection in our patient population.

  7. Comparison of four DNA extraction methods for the detection of Mycobacterium leprae from Ziehl-Neelsen-stained microscopic slides.

    Science.gov (United States)

    Ruiz-Fuentes, Jenny Laura; Díaz, Alexis; Entenza, Anayma Elena; Frión, Yahima; Suárez, Odelaisy; Torres, Pedro; de Armas, Yaxsier; Acosta, Lucrecia

    2015-12-01

    The diagnosis of leprosy has been a challenge due to the low sensibility of the conventional methods and the impossibility of culturing the causative organism. In this study, four methods for Mycobacterium leprae nucleic-acid extraction from Ziehl-Neelsen-stained slides (ZNS slides) were compared: Phenol/chloroform, Chelex 100 resin, and two commercial kits (Wizard Genomic DNA Purification Kit and QIAamp DNA Mini Kit). DNA was extracted from four groups of slides: a high-codification-slide group (bacteriological index [BI]⩾4), a low-codification-slide group (BI=1), a negative-slide group (BI=0), and a negative-control-slide group (BI=0). Quality DNA was evidenced by the amplification of specific repetitive element present in M. leprae genomic DNA (RLEP) using a nested polymerase chain reaction. This is the first report comparing four different extraction methods for obtaining M. leprae DNA from ZNS slides in Cuban patients, and applied in molecular diagnosis. Good-quality DNA and positive amplification were detected in the high-codification-slide group with the four methods, while from the low-codification-slide group only the QIAGEN and phenol-chloroform methods obtained amplification of M. leprae. In the negative-slide group, only the QIAGEN method was able to obtain DNA with sufficient quality for positive amplification of the RLEP region. No amplification was observed in the negative-control-slide group by any method. Patients with ZNS negative slides can still transmit the infection, and molecular methods can help identify and treat them, interrupting the chain of transmission and preventing the onset of disabilities. The ZNS slides can be sent easily to reference laboratories for later molecular analysis that can be useful not only to improve the diagnosis, but also for the application of other molecular techniques. Copyright © 2015 Asian-African Society for Mycobacteriology. Published by Elsevier Ltd. All rights reserved.

  8. Comparação entre dois métodos de detecção de DNA de papilomavírus humano em carcinoma epidermoide de lábio Comparison between two methods for human papilomavírus DNA detection in lip squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Adriana Demathe

    2010-04-01

    Full Text Available Introdução: Recentemente o papilomavírus humano (HPV tem sido associado à carcinogênese oral. A metodologia empregada na detecção do vírus é uma das maiores causas observadas da grande variabilidade nas taxas de detecção do HPV. Objetivo: Este estudo comparou a sensibilidade de detecção do DNA do HPV em casos de carcinoma epidermoide de lábio utilizando a amplificação do DNA viral por reação em cadeia da polimerase (PCR ou nPCR. Material e método: Foram utilizadas 33 amostras provenientes de casos de carcinoma epidermoide de lábio. Para as extrações do DNA utilizou-se o sistema QIAamp DNA Mini Kit. Como controle interno utilizou-se o gene da b-globina. Das 33 amostras iniciais, 30 foram positivas para o gene b-globina, sendo utilizadas para detectar o DNA viral. Comparou-se a amplificação do DNA viral pelos métodos da PCR com os oligonucleotídeos MY09/MY11 e nPCR, empregando-se os pares de oligonucleotídeos iniciadores MY09/MY11 e, na segunda etapa, o par GP5+/GP6+. O controle positivo para a presença do DNA do HPV utilizado foi a linhagem de células HeLa e, como controle negativo, a mistura de amplificação sem DNA. A análise dos produtos de PCR e nPCR para HPV foi realizada por eletroforese em gel de poliacrilamida a 8%. Resultados: Utilizando-se o método da PCR, a amplificação do DNA do HPV foi constatada em dois casos. Com a nPCR foi verificada presença de DNA viral em 13 das 30 amostras. Conclusão: Com a utilização da nPCR, a detecção do HPV nos casos estudados aumentou mais de seis vezes.Introduction: Human papillomavirus (HPV has been currently associated with oral carcinogesis. The methodology applied in virus detection is one of the main reasons for the great variability observed in HPV detection. Objectives: This study compared HPV DNA detection efficiency in lip squamous cell carcinoma samples (SCC using viral DNA amplification by PCR or nPCR. Methods: Thirty three samples of lip squamous cell

  9. Improved amplification efficiency on stool samples by addition of spermidine and its use for non-invasive detection of colorectal cancer

    KAUST Repository

    Roperch, Jean-Pierre; Benzekri, Karim; Mansour, Hicham; Incitti, Roberto

    2015-01-01

    Using quantitative methylation-specific PCR (QM-MSP) is a promising method for colorectal cancer (CRC) diagnosis from stool samples. Difficulty in eliminating PCR inhibitors of this body fluid has been extensively reported. Here

  10. and improvement of bowel function by increasing stool frequency pursuant to Article 13(5) of Regulation (EC) No 1924/2006

    DEFF Research Database (Denmark)

    Tetens, Inge

    2014-01-01

    to a combination of Bifidobacterium longum LA 101, Lactobacillus helveticus LA 102, Lactococcus lactis LA 103 and Streptococcus thermophilus LA 104 and improvement of bowel function by increasing stool frequency. The food that is the subject of the health claim is a combination of four bacterial strains—B. longum...... proposed by the applicant is "improves stool frequency". The Panel considers that improvement of bowel function by increasing stool frequency, provided that it does not result in diarrhoea, is a beneficial physiological effect. The Panel considers that the human study provided for the substantiation...... of the claim did not find an increase in stool frequency following consumption of a combination of the bacterial strains which is the subject of the claim. The Panel concludes that a cause and effect relationship has not been established between the consumption of a combination of B. longum LA 101, L...

  11. Recovery and identification of bacterial DNA from illicit drugs.

    Science.gov (United States)

    Cho, Kaymann T; Richardson, Michelle M; Kirkbride, K Paul; McNevin, Dennis; Nelson, Michelle; Pianca, Dennis; Roffey, Paul; Gahan, Michelle E

    2014-02-01

    Bacterial infections, including Bacillus anthracis (anthrax), are a common risk associated with illicit drug use, particularly among injecting drug users. There is, therefore, an urgent need to survey illicit drugs used for injection for the presence of bacteria and provide valuable information to health and forensic authorities. The objectives of this study were to develop a method for the extraction of bacterial DNA from illicit drugs and conduct a metagenomic survey of heroin and methamphetamine seized in the Australian Capital Territory during 2002-2011 for the presence of pathogens. Trends or patterns in drug contamination and their health implications for injecting drug users were also investigated. Methods based on the ChargeSwitch(®)gDNA mini kit (Invitrogen), QIAamp DNA extraction mini kit (QIAGEN) with and without bead-beating, and an organic phenol/chloroform extraction with ethanol precipitation were assessed for the recovery efficiency of both free and cellular bacterial DNA. Bacteria were identified using polymerase chain reaction and electrospray ionization-mass spectrometry (PCR/ESI-MS). An isopropanol pre-wash to remove traces of the drug and diluents, followed by a modified ChargeSwitch(®) method, was found to efficiently lyse cells and extract free and cellular DNA from Gram-positive and Gram-negative bacteria in heroin and methamphetamine which could then be identified by PCR/ESI-MS. Analysis of 12 heroin samples revealed the presence of DNA from species of Comamonas, Weissella, Bacillus, Streptococcus and Arthrobacter. No organisms were detected in the nine methamphetamine samples analysed. This study develops a method to extract and identify Gram-positive and Gram-negative bacteria from illicit drugs and demonstrates the presence of a range of bacterial pathogens in seized drug samples. These results will prove valuable for future work investigating trends or patterns in drug contamination and their health implications for injecting drug

  12. Comprehensive and Rapid Real-Time PCR Analysis of 21 Foodborne Outbreaks

    Directory of Open Access Journals (Sweden)

    Hiroshi Fukushima

    2009-01-01

    Full Text Available A set of four duplex SYBR Green I PCR (SG-PCR assay combined with DNA extraction using QIAamp DNA Stool Mini kit was evaluated for the detection of foodborne bacteria from 21 foodborne outbreaks. The causative pathogens were detected in almost all cases in 2 hours or less. The first run was for the detection of 8 main foodborne pathogens in 5 stool specimens within 2 hours and the second run was for the detection of other unusual suspect pathogens within a further 45 minutes. After 2 to 4 days, the causative agents were isolated and identified. The results proved that for comprehensive and rapid molecular diagnosis in foodborne outbreaks, Duplex SG-PCR assay is not only very useful, but is also economically viable for one-step differentiation of causative pathogens in fecal specimens obtained from symptomatic patients. This then allows for effective diagnosis and management of foodborne outbreaks.

  13. First identification of eggs of the Asian fish tapeworm Bothriocephalus acheilognathi (Cestoda: Bothriocephalidea) in human stool

    Czech Academy of Sciences Publication Activity Database

    Yera, H.; Kuchta, Roman; Brabec, Jan; Peyron, F.; Dupouy-Camet, J.

    2013-01-01

    Roč. 62, č. 3 (2013), s. 268-271 ISSN 1383-5769 R&D Projects: GA ČR GBP505/12/G112; GA ČR GAP506/12/1632 Institutional support: RVO:60077344 Keywords : COI * Diphyllobothrium * French Guiana * Man * Molecular identification * rDNA Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.111, year: 2013

  14. Probiotic properties of lactic acid bacteria isolated from stool samples of longevous people in regions of Hotan, Xinjiang and Bama, Guangxi, China.

    Science.gov (United States)

    Gu, Rui-Xia; Yang, Zhen-Quan; Li, Zheng-Hua; Chen, Shun-Li; Luo, Zhen-Lan

    2008-12-01

    A total of 567 lactic acid bacteria (LAB) strains were isolated from the stool samples of longevous people in regions of Hotan, Xinjiang and Bama, Guangxi, China. In order to reduce the number of strains for further examinations, 36 isolates were screened out for further examination whilst the other strains, which had lower probiotic properties, were not suitable for yogurt production due to the absence of growth in pH 3.5 MRS medium and no curding during fermentation, and so were excluded. The result of identification by API, sequence analysis of 16S rRNA and random amplified polymorphic DNA (RAPD) analysis showed that there were three strains of Lactobacillus acidophilus, 10 strains of Lactobacillus rhamnosus, three strains of Lactobacillus casein, three strains of Lactobacillus brevis, two strains of Enterococcus faecium, two strains of Enterococcus faecalis, four strains of Bifdibacterium infantis, three strains of Bifdibacterium brevise, three strains of Bifdibacterium bifidium, two strains of Bifdibacterium adolecentis and one strain of Bifdibacterium longam among the 36 isolates. These strains were evaluated by in vitro methods including survival upon exposure to pH 2.0, 3.0 and/or 0.3% oxgall and adhesion to the human colon adenocarcinoma cell line Caco-2 as well as antimicrobial activity against potential pathogens. The results presented here show that L. rhamnosus LV108, L. rhamnosus F, B. brevise R39 and B. infantis R42 are acid and bile tolerant, adhere to the cultured human intestinal Caco-2 cell line, antagonistic activity against potential pathogenic bacteria infection in vitro, and so are potential strains for probiotic use.

  15. Incidence of Giardia lamblia Subspecies by PCR-RFLP in Stool Specimens of Hospitalized Children at Urmia Mutahhari Hospital, West Azerbaijan Province, Iran.

    Directory of Open Access Journals (Sweden)

    Khosro Hazrati Tappeh

    2014-12-01

    Full Text Available Giardia lamblia is one of the most prevalent intestinal flagellate protozoa that infects a wide range of vertebrate hosts causing severe intestinal disorder in children.This study was performed to determine subspecies of G.lamblia by the PCR-RFLP method, targeting the glutamate dehydrogenase(gdhlocus, in hospitalized children at Urmia Mutahhari Hospital, West Azerbaijan Province,Iran and determining the infection transformational storages in this area.Overall, 720 stool specimens were collected from the hospitalized children, 34 samples were positive and Giardia cysts were detected under the microscope. Cysts were partially purified by the sucrose density gradient method and then washed with sterile distilled water to remove effectively the PCR inhibitors. Genomic DNA of G. lamblia isolates was extracted by freeze-thaw cycles followed by phenol/ chloroform/isoamyl alcohol method. The single step PCR-RFLP assay was used to differentiate the assemblages between A and B, which were found in humans. In this method, 432 bp expected size was amplified, and then for detection of subspecies, specific restriction RsaI and BspLI enzymes were used.Totally 34 samples were positive in terms of Giardia cyst out of 720 examined samples microscopically, so the parasite spread rate is reported 4.72%. Analysis PCR-RFLP on these samples revealed that 28 samples (93.3% have the genotype BIII and 2 samples (6.7% belong to the subgroup BIV.PCR-RFLP is a proper analytical method for determining the genotype among parasite types, using the glutamate dehydrogenizes zone's genes. Based on the results, an animal origin of infection cycle is suggested.

  16. Comparison of culture based methods for the isolation of Clostridium difficile from stool samples in a research setting.

    Science.gov (United States)

    Lister, Michelle; Stevenson, Emma; Heeg, Daniela; Minton, Nigel P; Kuehne, Sarah A

    2014-08-01

    Effective isolation of Clostridium difficile from stool samples is important in the research setting, especially where low numbers of spores/vegetative cells may be present within a sample. In this study, three protocols for stool culture were investigated to find a sensitive, cost effective and timely method of C. difficile isolation. For the initial enrichment step, the effectiveness of two different rich media, cycloserine-cefoxitin fructose broth (CCFB) and cycloserine-cefoxitin mannitol broth with taurocholate and lysozyme (CCMB-TAL) were compared. For the comparison of four different, selective solid media; Cycloserine-cefoxitin fructose agar (CCFA), Cycloserine-cefoxitin egg yolk agar (CCEY), ChromID C. difficile and tryptone soy agar (TSA) with 5% sheep's blood with and without preceding broth enrichment were used. As a means to enable differentiation between C. difficile and other fecal flora, the effectiveness of the inclusion of a pH indictor (1% Neutral Red), was also evaluated. The data derived indicated that CCFB is more sensitive than CCMB-TAL, however, the latter had an improved recovery rate. A broth enrichment step had a reduced sensitivity over direct plating. ChromID C. difficile showed the best recovery rate whereas CCEY egg yolk agar was the most sensitive of the four. The addition of 1% Neutral Red did not show sufficient colour change when added to CCEY egg yolk agar to be used as a differential medium. For a low cost, timely and sensitive method of isolating C. difficile from stool samples we recommend direct plating onto CCEY egg yolk agar after heat shock. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  17. Detection of Campylobacter in Stool and Determination of Significance by Culture, Enzyme Immunoassay, and PCR in Developing Countries

    Science.gov (United States)

    Platts-Mills, James A.; Liu, Jie; Gratz, Jean; Mduma, Esto; Amour, Caroline; Swai, Ndealilia; Taniuchi, Mami; Begum, Sharmin; Peñataro Yori, Pablo; Tilley, Drake H.; Lee, Gwenyth; Shen, Zeli; Whary, Mark T.; Fox, James G.; McGrath, Monica; Kosek, Margaret; Haque, Rashidul

    2014-01-01

    Campylobacter is a common bacterial enteropathogen that can be detected in stool by culture, enzyme immunoassay (EIA), or PCR. We compared culture for C. jejuni/C. coli, EIA (ProSpecT), and duplex PCR to distinguish Campylobacter jejuni/C. coli and non-jejuni/coli Campylobacter on 432 diarrheal and matched control stool samples from infants in a multisite longitudinal study of enteric infections in Tanzania, Bangladesh, and Peru. The sensitivity and specificity of culture were 8.5% and 97.6%, respectively, compared with the results of EIA and 8.7% and 98.0%, respectively, compared with the results of PCR for C. jejuni/C. coli. Most (71.6%) EIA-positive samples were positive by PCR for C. jejuni/C. coli, but 27.6% were positive for non-jejuni/coli Campylobacter species. Sequencing of 16S rRNA from 53 of these non-jejuni/coli Campylobacter samples showed that it most closely matched the 16S rRNA of C. hyointestinalis subsp. lawsonii (56%), C. troglodytis (33%), C. upsaliensis (7.7%), and C. jejuni/C. coli (2.6%). Campylobacter-negative stool spiked with each of the above-mentioned Campylobacter species revealed reactivity with EIA. PCR detection of Campylobacter species was strongly associated with diarrhea in Peru (odds ratio [OR] = 3.66, P < 0.001) but not in Tanzania (OR = 1.56, P = 0.24) or Bangladesh (OR = 1.13, P = 0.75). According to PCR, Campylobacter jejuni/C. coli infections represented less than half of all infections with Campylobacter species. In sum, in infants in developing country settings, the ProSpecT EIA and PCR for Campylobacter reveal extremely high rates of positivity. We propose the use of PCR because it retains high sensitivity, can ascertain burden, and can distinguish between Campylobacter infections at the species level. PMID:24452175

  18. Occurrence of Thermotolerant Campylobacter in Raw Poultry Meat, Environmental and Pigeon Stools Collected in Open-Air Markets.

    Science.gov (United States)

    Bellio, Alberto; Traversa, Amaranta; Adriano, Daniela; Bianchi, Daniela Manila; Colzani, Alberto; Gili, Stefano; Dondo, Alessandro; Gallina, Silvia; Grattarola, Carla; Maurella, Cristiana; Zoppi, Simona; Zuccon, Fabio; Decastelli, Lucia

    2014-08-28

    Campylobacteriosis was the most commonly reported zoonosis for confirmed human cases in European Union during 2011. Poultry meat was very often implicated in Campylobacter infections in humans. In Italy commerce of raw poultry meat is common in open-air markets: these areas can be considered at high risk of bacterial contamination due to the high presence birds like pigeons. The aim of this study was to collect data about the contamination by thermotolerant Campylobacter of raw poultry meat commercialised in open-air markets, of work-surfaces in contact with poultry meat and of pigeon stools sampled in the market areas in Turin, Northern Italy. Between September 2011 and December 2012, 86 raw poultry meat samples, 86 environmental swabs and 108 animal samples were collected in 38 open-air markets. Analysis were carried out according to ISO10272-1:2006 standard. C.coli was detected in 2.3% (2/86) of raw poultry meat samples, whereas no swab (0/86) resulted positive. Of pigeon stool 28% (30/107) was positive for C.jejuni (83.3% C.jejuni subsp . jejuni and 16.7% C.jejuni subsp . doylei ). C.jejuni subsp. jejuni was isolated from 1 dead pigeon . Our results showed lower rates of contamination than those reported at retail in Europe. Although samples were collected in areas at high risk of contamination, raw poultry meat and work surfaces reported a low level of presence of thermotolerant Campylobacter . The high percentage of C.jejuni isolated from pigeon stools showed the importance of a continuous application of preventive measures by the food business operators and the surveillance activity by the Competent Authority.

  19. Evaluation of a Single Procedure Allowing the Isolation of Enteropathogenic Yersinia along with Other Bacterial Enteropathogens from Human Stools

    Science.gov (United States)

    Savin, Cyril; Leclercq, Alexandre; Carniel, Elisabeth

    2012-01-01

    Enteropathogenic Yersinia are among the most frequent agents of human diarrhea in temperate and cold countries. However, the incidence of yersiniosis is largely underestimated because of the peculiar growth characteristics of pathogenic Yersinia, which make their isolation from poly-contaminated samples difficult. The use of specific procedures for Yersinia isolation is required, but is expensive and time consuming, and therefore is not systematically performed in clinical pathology laboratories. A means to circumvent this problem would be to use a single procedure for the isolation of all bacterial enteropathogens. Since the Statens Serum Institut enteric medium (SSI) has been reported to allow the growth at 37°C of most Gram-negative bacteria, including Yersinia, our study aimed at evaluating its performances for Yersinia isolation, as compared to the commonly used Yersinia-specific semi-selective Cefsulodin-Irgasan-Novobiocin medium (CIN) incubated at 28°C. Our results show that Yersinia pseudotuberculosis growth was strongly inhibited on SSI at 37°C, and therefore that this medium is not suitable for the isolation of this species. All Yersinia enterocolitica strains tested grew on SSI, while some non-pathogenic Yersinia species were inhibited. The morphology of Y. enterocolitica colonies on SSI allowed their differentiation from various other Gram-negative bacteria commonly isolated from stool samples. However, in artificially contaminated human stools, the recovery of Y. enterocolitica colonies on SSI at 37°C was difficult and was 3 logs less sensitive than on CIN at 28°C. Therefore, despite its limitations, the use of a specific procedure (CIN incubated at 28°C) is still required for an efficient isolation of enteropathogenic Yersinia from stools. PMID:22911756

  20. Effect of an ?-Lactalbumin-Enriched Infant Formula Supplemented With Oligofructose on Fecal Microbiota, Stool Characteristics, and Hydration Status

    OpenAIRE

    Wernimont, Susan; Northington, Robert; Kullen, Martin J.; Yao, Manjiang; Bettler, Jodi

    2014-01-01

    Aims. To evaluate the impact of oligofructose (OF)-supplemented infant formula on fecal microbiota, stool characteristics, and hydration. Methods. Ninety-five formula-fed infants were randomized to ?-lactalbumin-enriched control formula (CF) or identical formula with 3.0 g/L OF (EF) for 8 weeks; 50 infants fed human milk (HM) were included. Results. Eighty-four infants completed the study, 70 met per-protocol criteria. Over 8 weeks, bifidobacteria increased more in EF than CF group (0.70 vs 0...

  1. Stool screening of Syrian refugees and asylum seekers in Germany, 2013/2014: Identification of Sabin like polioviruses.

    Science.gov (United States)

    Böttcher, Sindy; Neubauer, Katrin; Baillot, Armin; Rieder, Gabriele; Adam, Maja; Diedrich, Sabine

    2015-10-01

    Germany is a partner of the Global Polio Eradication Initiative. Assurance of polio free status is based on enterovirus surveillance, which focuses on patients with signs of acute flaccid paralysis or aseptic meningitis/encephalitis, representing the key symptoms of poliovirus infection. In response to the wild poliovirus outbreak in Syria 2013 and high number of refugees coming from Syria to Germany, stool samples from 629 Syrian refugees/asylum seekers aged Syrian refugees and asylum seekers at that time. Copyright © 2015 Elsevier GmbH. All rights reserved.

  2. Identification of serotypes and virulence markers of Escherichia coli isolated from human stool and urine samples in Egypt

    Directory of Open Access Journals (Sweden)

    K M Osman

    2012-01-01

    Full Text Available Purpose: Haemorrhagic colitis and haemolytic-uremic syndrome are associated with Shiga-toxin producing Escherichia coli (STEC. There are others DEC (Diarrhoeagenic E. coli pathotypes responsible for outbreaks and others toxins associated to these. Most clinical signs of disease arise as a consequence of the production of Shiga toxin 1 (Stx1, Stx2 or combinations of these toxins. Other major virulence factors include E. coli haemolysin (hlyA, and intimin, the product of the eaeA gene that is involved in the attaching and effacing adherence phenotype. Materials and Methods: In this study, the PCR assay was used to detect 12 E. coli genes associated with virulence (stx1, stx2, hylA, Flic h7 , stb, F41, K99, sta, F17, LT-I, LT-II and eaeA. Results: A total of 108 E. coli strains were serotyped into 64 typable strains. The investigated strains from the stool, 8/80 (10% strains were O 164:K, while the 56/110 strains isolated from the urine were O126:K71 (44/110, 40% and O 86:K 61 (12/110, 11%. The distribution pattern of the detected virulence genes was observed to be in the following order: F17 (10% from the stool and 44% from the urine, Sta (10% from the stool, hylA (10% from the stool and 44% from the urine, Stb (44% from the urine and stx1 (27% from the urine. The 8 faecal strains encoded a combination of the F17, Sta and hylA genes, while the 56 urine strains encoded a combination of the F17 0+ Stb + hylA (44/110, 40% and Stx1 only (12/60, 20%. Conclusion: This is the first report on the molecular characterization of E. coli diarrhoeagenic strains in Egypt and the first report on the potential role of E. coli in diarrhoea and urinary tract infections in a localized geographic area where the people engage in various occupational activities.

  3. Limitations in the use of 14C-glycocholate breath and stool bile acid determinations in patients with chronic diarrhea

    International Nuclear Information System (INIS)

    Ferguson, J.; Walker, K.; Thomson, A.B.

    1986-01-01

    Analysis of a modified 14 C-glycocholate breath test on 165 consecutive in-patients being investigated for chronic diarrhea showed that the measurement of 14 CO 2 between 3 and 6 h after oral dosing of 5 microCi of 14 C-glycocholic acid was of only limited use to distinguish between patients with Crohn's disease (CD), idiopathic bile salt wastage (IBW), or ileal resection (IR) from those with the irritable bowel syndrome (IBS). Continuing 14 CO 2 collections for up to 24 h was of little more help in establishing the presence of bacterial overgrowth syndrome (BOS) and in distinguishing between BOS and CD. Stool bile acid measurements were of use in differentiating between IBW and IBS, but did not distinguish between CD and BOS or between CD and IR. Since the range of normal values was defined by measurements in the IBS group, a positive test was specific for an organic cause of chronic diarrhea. Even so, the sensitivity of the test was relatively low: CD, 53%; IR, 23%; IBW, 14 %; and BOS, 10%. We believe that the 24-h 14 C-glycocholic breath test combined with the measurement of stool bile acids represents a screening test of only limited use for the identification of organic causes of chronic diarrhea

  4. c-DNA of HIV-1 detection on spot of Buffy-Coat of leukocytes (DBCS

    Directory of Open Access Journals (Sweden)

    Marco Rossi de Gasperis

    2010-03-01

    Full Text Available Introduction:The elective way for the diagnosis of HIV-1-infection in the window period and in children under the age of 16-18 months is to search virus integrated in leukocytes. Aim of the study was to assess the sensitivity and specificity of extraction from Buffy-Dried Coat Spot (DBCS in leukocyte to detect c-DNA with nested-PCR in HIV-1-infected individuals compared to Dried Blood Spot (DBS both extracted by automated instrument EZ1 (QIAGEN, Hilden, Germany. Both DBCS and both DBS were compared with those tests from whole blood by conventional DNA-extraction Methods: Five ml of whole blood from 50 HIV-infected individuals were collected. 40 μl of each sample were spotted on “FTA ELUTE Micro Card” (Whatman, Inc., Clifton, NJ, 200 μl were extracted according to the manual procedure (QIAGEN “QIAamp DNA minikit and the remaining sample was incubated at 37 °C for 120 minutes. Plasma was centrifuged at 1000 rcf/1g for 10 minutes at room temperature. Forty μl of the obtained buffy-coat was spotted. Both DBCS and both DBS were dried at room temperature for 24 hours.Two of 5 punch from each spot were extracted with TISSUE DNA kit (Biorobot EZ1 DSP “Qiagen” and eluted in 50 μl of buffer.The recovery of genomic DNA was measured amplifying the ß-globin gene by Real-Time “SybrGreen I”.The DNA was amplified for the “pol” gene of HIV-1 by nested PCR and revealed in “SYBR-green I”. Eight HIV-antibody-negative samples were used as internal control. Results:The experimental protocol adopted for the DBCS showed high sensitivity and specificity.The extracted DNA from DBS and DBCS was characterized by excellent quality and without any inhibitory agents. The amount of proviral DNA extracted from DBCS is similar to that obtained by conventional extraction, while the DBS test was significantly less sensitive. Conclusion:These preliminary data suggest that the amount of c-DNA obtained with DBS technique is often not enough for the

  5. Acceptability of study procedures (self-collected introital swabs, blood draws and stool sample collection) by students 10-16 years for an HPV vaccine effectiveness study: a pilot study.

    Science.gov (United States)

    Nakalembe, Miriam; Mutyaba, Twaha; Mirembe, Florence

    2016-03-16

    A cohort study was planned to evaluate vaccine immunogenicity and effect of malaria and helminth co-infections on the bivalent Human papilloma virus (HPV) vaccine. The study would involve self collected introital swabs, blood draws and stool sample collection. We therefore conducted a pilot study to assess the acceptability of these procedures among the students and their parents. A cross-sectional study among forty four students from two purposively selected primary schools of Western Uganda. Exit interviews and two focus group discussions (FGD) (for parents) were conducted. Acceptability was measured by willingness to undergo the procedures again, recommending the procedures to others as well as proportion of introital swabs positive for β globulin. FGD determined acceptability of the parents and explored opinions and perceptions that would influence their decisions. HPV-16/18 and β globulin deoxyribonucleic acid (DNA) were analysed using a polymerase chain reaction (PCR) kit. All the students (100%) in the study were willing to provide a self- collected introital swab and a stool sample as well as recommending their friends while (86.3%) were willing for blood draws. There were 40/44 (90.1%) self collected introital swabs that had positive result for human β globulin though none of them was positive for HPV-16/18. In the FGD, it emerged that parents concerns were on the blood draws and introital swab collection which were addressed. The study procedures were highly acceptable among this study population of students and their parents. Follow-up to assess HPV vaccine effectiveness and factors that may influence the vaccine in this age group is feasible.

  6. Modeling DNA

    Science.gov (United States)

    Robertson, Carol

    2016-01-01

    Deoxyribonucleic acid (DNA) is life's most amazing molecule. It carries the genetic instructions that almost every organism needs to develop and reproduce. In the human genome alone, there are some three billion DNA base pairs. The most difficult part of teaching DNA structure, however, may be getting students to visualize something as small as a…

  7. Performance evaluation of direct saline stool microscopy, Formol ether concentration and Kato Katz diagnostic methods for intestinal parasitosis in the absence of gold standard methods.

    Science.gov (United States)

    Hailu, Tadesse; Abera, Bayeh

    2015-07-01

    The parasite load within the sample and the amount of sample taken during examination greatly compromise the sensitivity of direct saline stool microscopy. A cross-sectional study was conducted in March 2011 in Bahir Dar city among 778 fresh single stool samples to evaluate the performance of direct saline (DS), Kato Katz (KK) and Formol ether concentration (FEC) methods against the 'Gold' standard. Among 778 stool samples from school age children, the highest prevalence of intestinal parasites was recorded by FEC (55.1%). The sensitivity of DS, FEC and KK were 61.1%, 92.3% and 58.7%, respectively. FEC is more sensitive than DS and KK. Hence, use of the latter is preferred. © The Author(s) 2015.

  8. Cholera diagnosis in human stool and detection in water: protocol for a systematic review of available technologies.

    Science.gov (United States)

    Diaconu, Karin; Falconer, Jennifer; O'May, Fiona; Jimenez, Miguel; Matragrano, Joe; Njanpop-Lafourcade, Betty; Ager, Alastair

    2018-02-20

    Cholera is a highly infectious diarrheal disease spread via fecal contamination of water and food sources; it is endemic in parts of Africa and Asia and recent outbreaks have been reported in Haiti, the Zambia and Democratic Republic of the Congo. If left untreated, the disease can be fatal in less than 24 h and result in case fatality ratios of 30-50%. Cholera disproportionately affects those living in areas with poor access to water and sanitation: the long-term public health response is focused on improving water and hygiene facilities and access. Short-term measures for infection prevention and control, and disease characterization and surveillance, are impaired by diagnostic delays: culture methods are slow and rely on the availability of infrastructure and specialist equipment. Rapid diagnostic tests have shown promise under field conditions and further innovations in this area have been proposed. This paper is the protocol for a systematic review focused on identifying current technologies and methods used for cholera diagnosis in stool, and detection in water. We will synthesize and appraise information on product technical specifications, accuracy and design features in order to inform infection prevention and control and innovation development. Embase, MEDLINE, CINAHL, Proquest, IndMed and the WHO and Campbell libraries will be searched. We will include studies reporting on field evaluations, including within-study comparisons against a reference standard, and laboratory evaluations reporting on product validation against field stool or water samples. We will extract data according to protocol and attempt meta-analyses if appropriate given data availability and quality. The systematic review builds on a previous scoping review in this field and expands upon this by synthesising data on both product technical characteristics and design features. The review will be of particular value to stakeholders engaged in diagnostic procurement and manufacturers

  9. Cholera diagnosis in human stool and detection in water: protocol for a systematic review of available technologies

    Directory of Open Access Journals (Sweden)

    Karin Diaconu

    2018-02-01

    Full Text Available Abstract Background Cholera is a highly infectious diarrheal disease spread via fecal contamination of water and food sources; it is endemic in parts of Africa and Asia and recent outbreaks have been reported in Haiti, the Zambia and Democratic Republic of the Congo. If left untreated, the disease can be fatal in less than 24 h and result in case fatality ratios of 30–50%. Cholera disproportionately affects those living in areas with poor access to water and sanitation: the long-term public health response is focused on improving water and hygiene facilities and access. Short-term measures for infection prevention and control, and disease characterization and surveillance, are impaired by diagnostic delays: culture methods are slow and rely on the availability of infrastructure and specialist equipment. Rapid diagnostic tests have shown promise under field conditions and further innovations in this area have been proposed. Methods This paper is the protocol for a systematic review focused on identifying current technologies and methods used for cholera diagnosis in stool, and detection in water. We will synthesize and appraise information on product technical specifications, accuracy and design features in order to inform infection prevention and control and innovation development. Embase, MEDLINE, CINAHL, Proquest, IndMed and the WHO and Campbell libraries will be searched. We will include studies reporting on field evaluations, including within-study comparisons against a reference standard, and laboratory evaluations reporting on product validation against field stool or water samples. We will extract data according to protocol and attempt meta-analyses if appropriate given data availability and quality. Discussion The systematic review builds on a previous scoping review in this field and expands upon this by synthesising data on both product technical characteristics and design features. The review will be of particular value to

  10. Geographical Variation in Antibiotic-Resistant Escherichia coli Isolates from Stool, Cow-Dung and Drinking Water

    Science.gov (United States)

    Sahoo, Krushna Chandra; Tamhankar, Ashok J.; Sahoo, Soumyakanta; Sahu, Priyadarshi Soumyaranjan; Klintz, Senia Rosales; Lundborg, Cecilia Stålsby

    2012-01-01

    Little information is available on relationships between the biophysical environment and antibiotic resistance. This study was conducted to investigate the antibiotic resistance pattern of Escherichia coli isolated from child stool samples, cow-dung and drinking water from the non-coastal (230 households) and coastal (187 households) regions of Odisha, India. Susceptibility testing of E. coli isolates (n = 696) to the following antibiotics: tetracycline, ampicillin/sulbactam, cefuroxime, cefotaxime, cefixime, cotrimoxazole, amikacin, ciprofloxacin, norfloxacin and nalidixic acid was performed by the disk diffusion method. Ciprofloxacin minimum inhibitory concentration (MIC) values were determined for ciprofloxacin-resistant isolates (n = 83). Resistance to at least one antibiotic was detected in 90% or more of the E. coli isolates. Ciprofloxacin MIC values ranged from 8 to 32 µg/mL. The odds ratio (OR) of resistance in E. coli isolates from children’s stool (OR = 3.1, 95% CI 1.18–8.01), cow-dung (OR = 3.6, 95% CI 1.59–8.03, P = 0.002) and drinking water (OR = 3.8, 95% CI 1.00–14.44, P = 0.049) were higher in non-coastal compared to coastal region. Similarly, the co-resistance in cow-dung (OR = 2.5, 95% CI 1.39–4.37, P = 0.002) and drinking water (OR = 3.2, 95% CI 1.36–7.41, P = 0.008) as well as the multi-resistance in cow-dung (OR = 2.2, 95% CI 1.12–4.34, P = 0.022) and drinking water (OR = 2.7, 95% CI 1.06–7.07, P = 0.036) were also higher in the non-coastal compared to the coastal region. PMID:22690160

  11. Comparison of a Stool Antigen Detection Kit and PCR for Diagnosis of Entamoeba histolytica and Entamoeba dispar Infections in Asymptomatic Cyst Passers in Iran

    OpenAIRE

    Solaymani-Mohammadi, Shahram; Rezaian, Mostafa; Babaei, Zahra; Rajabpour, Azam; Meamar, Ahmad R.; Pourbabai, Ahmad A.; Petri, William A.

    2006-01-01

    The present study was conducted to compare stool antigen detection with PCR for the diagnosis of Entamoeba sp. infection in asymptomatic cyst passers from Iran. Entamoeba dispar and, in one case, E. moshkovskii were the Entamoeba spp. found in the amebic cyst passers. There was a 100% correlation between the results from the TechLab E. histolytica II stool antigen kit and those from nested PCR. We concluded that E. dispar is much more common in asymptomatic cyst passers in Iran and that antig...

  12. Comparison of a stool antigen detection kit and PCR for diagnosis of Entamoeba histolytica and Entamoeba dispar infections in asymptomatic cyst passers in Iran.

    Science.gov (United States)

    Solaymani-Mohammadi, Shahram; Rezaian, Mostafa; Babaei, Zahra; Rajabpour, Azam; Meamar, Ahmad R; Pourbabai, Ahmad A; Petri, William A

    2006-06-01

    The present study was conducted to compare stool antigen detection with PCR for the diagnosis of Entamoeba sp. infection in asymptomatic cyst passers from Iran. Entamoeba dispar and, in one case, E. moshkovskii were the Entamoeba spp. found in the amebic cyst passers. There was a 100% correlation between the results from the TechLab E. histolytica II stool antigen kit and those from nested PCR. We concluded that E. dispar is much more common in asymptomatic cyst passers in Iran and that antigen detection and PCR are comparable diagnostic modalities.

  13. Detection of Cryptosporidium sp infection by PCR and modified acid fast staining from potassium dichromate preserved stool

    OpenAIRE

    Agnes Kurniawan; Sri W. Dwintasari; Herbowo A. Soetomenggolo; Septelia I. Wanandi

    2009-01-01

    Aim To identify the frequency of Cryptosporidium infection in children below 3 years old by examining concentrated long term preserved stool using PCR detection of 18S rRNA gene and compared with modified acid fast staining technique.Methods Hundred eighty eight stools from children ≤ 3 years old were stored for 13 months in 2.5% K2Cr2O7 solution at 40C. Cryptosporidium oocysts were isolated by water-ether concentration technique. The concentrates were smeared onto object glass and stained wi...

  14. A novel method of selective removal of human DNA improves PCR sensitivity for detection of Salmonella Typhi in blood samples.

    Science.gov (United States)

    Zhou, Liqing; Pollard, Andrew J

    2012-07-27

    Enteric fever is a major public health problem, causing an estimated 21million new cases and 216,000 or more deaths every year. Current diagnosis of the disease is inadequate. Blood culture only identifies 45 to 70% of the cases and is time-consuming. Serological tests have very low sensitivity and specificity. Clinical samples obtained for diagnosis of enteric fever in the field generally have blood, so that even PCR-based methods, widely used for detection of other infectious diseases, are not a straightforward option in typhoid diagnosis. We developed a novel method to enrich target bacterial DNA by selective removal of human DNA from blood samples, enhancing the sensitivity of PCR tests. This method offers the possibility of improving PCR assays directly using clinical specimens for diagnosis of this globally important infectious disease. Blood samples were mixed with ox bile for selective lysis of human blood cells and the released human DNA was then digested with addition of bile resistant micrococcal nuclease. The intact Salmonella Typhi bacteria were collected from the specimen by centrifugation and the DNA extracted with QIAamp DNA mini kit. The presence of Salmonella Typhi bacteria in blood samples was detected by PCR with the fliC-d gene of Salmonella Typhi as the target. Micrococcal nuclease retained activity against human blood DNA in the presence of up to 9% ox bile. Background human DNA was dramatically removed from blood samples through the use of ox bile lysis and micrococcal nuclease for removal of mammalian DNA. Consequently target Salmonella Typhi DNA was enriched in DNA preparations and the PCR sensitivity for detection of Salmonella Typhi in spiked blood samples was enhanced by 1,000 fold. Use of a combination of selective ox-bile blood cell lysis and removal of human DNA with micrococcal nuclease significantly improves PCR sensitivity and offers a better option for improved typhoid PCR assays directly using clinical specimens in diagnosis of

  15. DNA Camouflage

    Science.gov (United States)

    2016-01-08

    1 DNA Camouflage Supplementary Information Bijan Zakeri1,2*, Timothy K. Lu1,2*, Peter A. Carr2,3* 1Department of Electrical Engineering and...ll.mit.edu). Distribution A: Public Release   2 Supplementary Figure 1 DNA camouflage with the 2-state device. (a) In the presence of Cre, DSD-2[α...10 1 + Cre 1 500 1,000 length (bp) chromatogram alignment template − Cre   4 Supplementary Figure 3 DNA camouflage with a switchable

  16. Relating Stool Microbial Metabolite Levels, Inflammatory Markers and Dietary Behaviors to Screening Colonoscopy Findings in a Racially/Ethnically Diverse Patient Population

    Directory of Open Access Journals (Sweden)

    Kristina M. Bridges

    2018-02-01

    Full Text Available Colorectal cancer (CRC is the third leading cause of cancer death for both men and women in the United States, yet it is treatable and preventable. African Americans have higher incidence of CRC than other racial/ethnic groups, however, it is unclear whether this disparity is primarily due to environmental or biological factors. Short chain fatty acids (SCFAs are metabolites produced by bacteria in the colon and are known to be inversely related to CRC progression. The aim of this study is to investigate how stool SCFA levels, markers of inflammation in stool and dietary intake relate to colonoscopy findings in a diverse patient population. Stool samples from forty-eight participants were analyzed for SCFA levels and inflammatory markers (lysozyme, secretory IgA, lactoferrin. Additionally, participants completed the National Cancer Institute’s Diet History Questionnaire II (DHQ II to report dietary intake over the past year. Subsequently, the majority of participants underwent screening colonoscopy. Our results showed that African Americans had higher total levels of SCFAs in stool than other racial/ethnic groups, significantly lower intake of non-starchy vegetables and similar inflammatory marker expression and colonoscopy outcomes, compared to others. This work is an initial exploration into the biological and clinical factors that may ultimately inform personalized screening approaches and clinical decision-making to improve colorectal cancer disparities for African Americans.

  17. Mathematical inference on helminth egg counts in stool and its applications in mass drug administration programmes to control soil-transmitted helminthiasis in public health.

    Science.gov (United States)

    Levecke, Bruno; Anderson, Roy M; Berkvens, Dirk; Charlier, Johannes; Devleesschauwer, Brecht; Speybroeck, Niko; Vercruysse, Jozef; Van Aelst, Stefan

    2015-03-01

    In the present study, we present a hierarchical model based on faecal egg counts (FECs; expressed in eggs per 1g of stool) in which we first describe the variation in FECs between individuals in a particular population, followed by describing the variance due to counting eggs under a microscope separately for each stool sample. From this general framework, we discuss how to calculate a sample size for assessing a population mean FEC and the impact of an intervention, measured as reduction in FECs, for any scenario of soil-transmitted helminth (STH) epidemiology (the intensity and aggregation of FECs within a population) and diagnostic strategy (amount of stool examined (∼sensitivity of the diagnostic technique) and examination of individual/pooled stool samples) and on how to estimate prevalence of STH in the absence of a gold standard. To give these applications the most wide relevance as possible, we illustrate each of them with hypothetical examples. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Relating Stool Microbial Metabolite Levels, Inflammatory Markers and Dietary Behaviors to Screening Colonoscopy Findings in a Racially/Ethnically Diverse Patient Population

    Science.gov (United States)

    Bridges, Kristina M.; Diaz, Francisco J.; Wang, Zhiwen; Ahmed, Ishfaq; Sullivan, Debra K.; Umar, Shahid; Buckles, Daniel C.; Greiner, K. Allen; Hester, Christina M.

    2018-01-01

    Colorectal cancer (CRC) is the third leading cause of cancer death for both men and women in the United States, yet it is treatable and preventable. African Americans have higher incidence of CRC than other racial/ethnic groups, however, it is unclear whether this disparity is primarily due to environmental or biological factors. Short chain fatty acids (SCFAs) are metabolites produced by bacteria in the colon and are known to be inversely related to CRC progression. The aim of this study is to investigate how stool SCFA levels, markers of inflammation in stool and dietary intake relate to colonoscopy findings in a diverse patient population. Stool samples from forty-eight participants were analyzed for SCFA levels and inflammatory markers (lysozyme, secretory IgA, lactoferrin). Additionally, participants completed the National Cancer Institute’s Diet History Questionnaire II (DHQ II) to report dietary intake over the past year. Subsequently, the majority of participants underwent screening colonoscopy. Our results showed that African Americans had higher total levels of SCFAs in stool than other racial/ethnic groups, significantly lower intake of non-starchy vegetables and similar inflammatory marker expression and colonoscopy outcomes, compared to others. This work is an initial exploration into the biological and clinical factors that may ultimately inform personalized screening approaches and clinical decision-making to improve colorectal cancer disparities for African Americans. PMID:29495356

  19. Cell-free mitochondrial DNA copy number variation in head and neck squamous cell carcinoma: A study of non-invasive biomarker from Northeast India.

    Science.gov (United States)

    Kumar, Manish; Srivastava, Shilpee; Singh, Seram Anil; Das, Anup Kumar; Das, Ganesh Chandra; Dhar, Bishal; Ghosh, Sankar Kumar; Mondal, Rosy

    2017-10-01

    Head and neck squamous cell carcinoma is the most commonly diagnosed cancer worldwide. The lifestyle, food habits, and customary practices manifest the Northeast Indian population toward higher susceptibility to develop head and neck squamous cell carcinoma. Here, we have investigated the association of smoke and smokeless tobacco, and alcohol with copy number variation of cell-free mitochondrial DNA and cell-free nuclear DNA in cases and controls. Cell-free DNA from plasma was isolated from 50 head and neck squamous cell carcinoma cases and 50 controls with informed written consent using QIAamp Circulating Nucleic Acid Kit. Real-time polymerase chain reaction was done for copy number variation in cell-free mitochondrial DNA and cell-free nuclear DNA. Receiver operating characteristic curve analysis was performed to evaluate the diagnostic application between the two study groups using clinicopathological parameters. The levels of cell-free nuclear DNA and cell-free mitochondrial DNA of cases in association with smoke and smokeless tobacco, alcohol with smoking (p squamous cell carcinoma cases and controls, we distinguished cell-free mitochondrial DNA (cutoff: 19.84 raw Ct; sensitivity: 84%; specificity: 100%; p < 0.001) and cell-free nuclear DNA (cutoff: 463,282 genomic equivalent/mL; sensitivity: 53%; specificity: 87%; p < 0.001). The copy number variation in cases (cell-free nuclear DNA: 5451.66 genomic equivalent/mL and cell-free mitochondrial DNA: 29,103,476.15 genomic equivalent/mL) and controls (cell-free nuclear DNA: 1650.9 genomic equivalent/mL and cell-free mitochondrial DNA: 9,189,312.54 genomic equivalent/mL), respectively. Our result indicates that the cell-free mitochondrial DNA content is highly associated with smoke and smokeless tobacco, betel quid chewing, and alcohol which shows greater promises, holding the key characteristics of diagnostic biomarkers, that is, minimal invasiveness, high specificity, and sensitivity.

  20. Diagnostic efficacy of monoclonal antibody based sandwich enzyme linked immunosorbent assay (ELISA for detection of Fasciola gigantica excretory/secretory antigens in both serum and stool

    Directory of Open Access Journals (Sweden)

    Zoheiry Mona K

    2011-09-01

    Full Text Available Abstract Background This research was carried out to develop a reliable monoclonal antibody (MoAb-based sandwich enzyme linked immunosorbent assay (ELISA for the diagnosis of active Fasciola gigantica infection in both serum and stool for comparative purposes. Methods From a panel of MoAbs raised against F. gigantica excretory/secretory antigens (ES Ags, a pair (12B/11D/3F and 10A/9D/10G was chosen due to its high reactivity and strict specificity to F. gigantica antigen by indirect ELISA. Results The two MoAbs were of the IgG1 and IgG2a subclasses, respectively. Using SDS-PAGE and EITB, the selected MoAbs recognized 83, 64, 45 and 26 kDa bands of ES Ags. The lower detection limit of ELISA assay was 3 ng/ml. In stool, the sensitivity, specificity and diagnostic efficacy of ELISA was 96%, 98.2 and 97.1%; while in serum they were 94%, 94.6% and 94.3%, respectively. Moreover, a positive correlation was found between ova count in stool of F. gigantica infected patients and the OD readings of ELISA in both stool and serum samples (r = 0.730, p Conclusions These data showed that the use of MoAb-based sandwich ELISA for the detection of F. gigantica coproantigens in stool specimens was superior to serum samples; it provides a highly efficient, non-invasive technique for the diagnosis of active F. gigantica infection.

  1. Difference in Ulex europaeus agglutinin I-binding activity of decay-accelerating factor detected in the stools of patients with colorectal cancer and ulcerative colitis.

    Science.gov (United States)

    Okazaki, Hiroaki; Mizuno, Motowo; Nasu, Junichirou; Makidono, Chiho; Hiraoka, Sakiko; Yamamoto, Kazuhide; Okada, Hiroyuki; Fujita, Teizo; Tsuji, Takao; Shiratori, Yasushi

    2004-03-01

    Expression of decay-accelerating factor (DAF, CD55), a complement-regulatory glycoprotein, is enhanced in colorectal-cancer (CC) cells and colonic epithelium in ulcerative colitis (UC), and stools from these patients contain increased amounts of DAF. Carbohydrate chains of glycoproteins are often altered during malignant transformation or inflammation. In this study, we investigated whether DAF molecules in patients with CC and those with UC differ with respect to oligosaccharide side chains. We analyzed DAF in stools and homogenates of colonic-tissue specimens obtained from patients with CC or UC using solid-phase enzyme-linked assay and Western blotting for reactivity with the lectins Ulex europaeus agglutinin I (UEA-I), wheat-germ agglutinin, peanut agglutinin, and concanavalin A. UEA-I bound to DAF in stools from patients with UC but not in that from the stools of CC patients, as demonstrated on the solid-phase enzyme-linked assay (P <.05, Mann-Whitney U test) and Western blotting. Binding of UEA-I was specifically inhibited by the addition of fucose. The difference in UEA-I reactivity with DAF was observed also in colonic-tissue homogenates from patients with UC and those with CC. DAF expressed in the mucosa and excreted into the stools of UC patients is different from that expressed in CC with regard to UEA-I reactivity. Future studies should be directed toward determining whether a qualitatively unique isoform of DAF is present, of which sugar chains are specific to CC in UC patients.

  2. Comparison of three stool antigen assays with the 13C- urea breath test for the primary diagnosis of Helicobacter pylori infection and monitoring treatment outcome.

    LENUS (Irish Health Repository)

    Hooton, Carmel

    2012-02-03

    BACKGROUND: The urea breath test (UBT) is the gold-standard non-invasive test for the detection of Helicobacter pylori infection, however, the lack of availability of the UBT due to the high cost of the test, and in particular the need for expensive analytical instrumentation, limits the usefulness of this method. Stool antigen assays may offer an alternative non-invasive method for the diagnosis of infection. OBJECTIVE: To compare the accuracy of three stool antigen assays (HpSA, IDEIA HpStAR, and ImmunoCard STAT) against the UBT for the primary diagnosis of H. pylori infection and for monitoring treatment outcome. METHODS: A total of 102 patients attending two gastroenterology day-case clinics for the investigation of dyspepsia were included. Each patient provided breath and stool samples for analysis. Patients who tested positive for H. pylori by the validated UBT were prescribed triple therapy and invited to return for repeat breath and stool sample analysis 6 weeks post-treatment. RESULTS: Of the 102 patients tested, 48 were diagnosed with H. pylori infection by the UBT. The HpSA assay interpreted 38 of these as positive (79% sensitive). Of the 54 UBT-negative patients the HpSA assay interpreted all 54 as negative (100% specific). The IDEIA HpStAR assay correctly identified 44 patients as positive (92% sensitive) and 50 as negative (92.5% specific). The ImmunoCard STAT assay interpreted 38 patients as positive (79% sensitive) and 52 as negative (96.3% specific). CONCLUSION: The findings indicate that the IDEIA HpStAR stool antigen kit is the most accurate assay of the three assays evaluated, and possibly represents a viable alternative to the UBT for the primary diagnosis of H. pylori infection and for monitoring treatment outcome.

  3. Efficiency of Direct Microscopy of Stool Samples Using an Antigen-Specific Adhesin Test for Entamoeba Histolytica

    Directory of Open Access Journals (Sweden)

    Arzu İrvem

    2016-10-01

    Full Text Available Background: E. histolytica is among the common causes of acute gastroenteritis. The pathogenic species E. histolytica and the nonpathogenic species E. dispar cannot be morphologically differentiated, although correct identification of these protozoans is important for treatment and public health. In many laboratories, the screening of leukocytes, erythrocytes, amoebic cysts, trophozoites and parasite eggs is performed using Native-Lugol’s iodine for pre-diagnosis. Aims: In this study, we aimed to investigate the frequency of E. histolytica in stool samples collected from 788 patients residing in the Anatolian region of İstanbul who presented with gastrointestinal complaints. We used the information obtained to evaluate the effectiveness of microscopic examinations when used in combination with the E. histolytica adhesin antigen test. Study Design: Retrospective cross-sectional study Methods: Preparations of stool samples stained with Native-Lugol’s iodine were evaluated using the E. histolytica adhesin test and examined using standard light microscopy at ×40 magnification. Pearson’s Chi-square and Fisher’s exact tests were used for statistical analysis. Logistic regression analysis was used for multivariate analysis. Results: Of 788 samples, 38 (4.8% were positive for E. histolytica adhesin antigens. When evaluated together with the presences of erythrocytes, leukocytes, cysts, and trophozoites, respectively, using logistic regression analysis, leukocyte positivity was significantly higher. The odds ratio of leukocyte positivity increased adhesin test-positivity by 2,530-fold (95% CI=1.01–6.330. Adhesin test-positivity was significant (p=0.047. Conclusion: In line with these findings, the consistency between the presence of cysts and erythrocytes and adhesin test-positivity was found to be highly significant, but that of higher levels of leukocytes was found to be discordant. It was concluded that leukocytes and trophozoites were

  4. DNA glue

    DEFF Research Database (Denmark)

    Filichev, Vyacheslav V; Astakhova, Irina V.; Malakhov, Andrei D.

    2008-01-01

    Significant alterations in thermal stability of parallel DNA triplexes and antiparallel duplexes were observed upon changing the attachment of ethynylpyrenes from para to ortho in the structure of phenylmethylglycerol inserted as a bulge into DNA (TINA). Insertions of two ortho-TINAs as a pseudo...

  5. Hyperstretching DNA

    NARCIS (Netherlands)

    Schakenraad, Koen; Biebricher, Andreas S.; Sebregts, Maarten; Ten Bensel, Brian; Peterman, Erwin J.G.; Wuite, Gijs J L; Heller, Iddo; Storm, Cornelis; Van Der Schoot, Paul

    2017-01-01

    The three-dimensional structure of DNA is highly susceptible to changes by mechanical and biochemical cues in vivo and in vitro. In particular, large increases in base pair spacing compared to regular B-DNA are effected by mechanical (over)stretching and by intercalation of compounds that are widely

  6. Evaluation of the BD Max Cdiff assay for the detection of toxigenic Clostridium difficile in human stool specimens.

    Science.gov (United States)

    Putsathit, Papanin; Morgan, Justin; Bradford, Damien; Engelhardt, Nelly; Riley, Thomas V

    2015-02-01

    The Becton Dickinson (BD) PCR-based GeneOhm Cdiff assay has demonstrated a high sensitivity and specificity for detecting Clostridium difficile. Recently, the BD Max platform, using the same principles as BD GeneOhm, has become available in Australia. This study aimed to investigate the sensitivity and specificity of BD Max Cdiff assay for the detection of toxigenic C. difficile in an Australian setting. Between December 2013 and January 2014, 406 stool specimens from 349 patients were analysed with the BD Max Cdiff assay. Direct and enrichment toxigenic culture were performed on bioMérieux ChromID C. difficile agar as a reference method. isolates from specimens with discrepant results were further analysed with an in-house PCR to detect the presence of toxin genes. The overall prevalence of toxigenic C. difficile was 7.2%. Concordance between the BD Max assay and enrichment culture was 98.5%. The sensitivity, specificity, positive predictive value and negative predictive value for the BD Max Cdiff assay were 95.5%, 99.0%, 87.5% and 99.7%, respectively, when compared to direct culture, and 91.7%, 99.0%, 88.0% and 99.4%, respectively, when compared to enrichment culture. The new BD Max Cdiff assay appeared to be an excellent platform for rapid and accurate detection of toxigenic C. difficile.

  7. Pilot Dietary Intervention with Heat-Stabilized Rice Bran Modulates Stool Microbiota and Metabolites in Healthy Adults

    Directory of Open Access Journals (Sweden)

    Amy M. Sheflin

    2015-02-01

    Full Text Available Heat-stabilized rice bran (SRB has been shown to regulate blood lipids and glucose, modulate gut mucosal immunity and inhibit colorectal cancer in animal and human studies. However, SRB’s effects on gut microbial composition and metabolism and the resulting implications for health remain largely unknown. A pilot, randomized-controlled trial was developed to investigate the effects of eating 30 g/day SRB on the stool microbiome and metabolome. Seven healthy participants consumed a study meal and snack daily for 28 days. The microbiome and metabolome were characterized using 454 pyrosequencing and gas chromatography-mass spectrometry (GC-MS at baseline, two and four weeks post-intervention. Increases in eight operational taxonomic units (OTUs, including three from Bifidobacterium and Ruminococcus genera, were observed after two and four weeks of SRB consumption (p < 0.01. Branched chain fatty acids, secondary bile acids and eleven other putative microbial metabolites were significantly elevated in the SRB group after four weeks. The largest metabolite change was a rice bran component, indole-2-carboxylic acid, which showed a mean 12% increase with SRB consumption. These data support the feasibility of dietary SRB intervention in adults and support that SRB consumption can affect gut microbial metabolism. These findings warrant future investigations of larger cohorts evaluating SRB’s effects on intestinal health.

  8. DNA probes

    International Nuclear Information System (INIS)

    Castelino, J.

    1992-01-01

    The creation of DNA probes for detection of specific nucleotide segments differs from ligand detection in that it is a chemical rather than an immunological reaction. Complementary DNA or RNA is used in place of the antibody and is labelled with 32 P. So far, DNA probes have been successfully employed in the diagnosis of inherited disorders, infectious diseases, and for identification of human oncogenes. The latest approach to the diagnosis of communicable and parasitic infections is based on the use of deoxyribonucleic acid (DNA) probes. The genetic information of all cells is encoded by DNA and DNA probe approach to identification of pathogens is unique because the focus of the method is the nucleic acid content of the organism rather than the products that the nucleic acid encodes. Since every properly classified species has some unique nucleotide sequences that distinguish it from every other species, each organism's genetic composition is in essence a finger print that can be used for its identification. In addition to this specificity, DNA probes offer other advantages in that pathogens may be identified directly in clinical specimens

  9. DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Castelino, J

    1993-12-31

    The creation of DNA probes for detection of specific nucleotide segments differs from ligand detection in that it is a chemical rather than an immunological reaction. Complementary DNA or RNA is used in place of the antibody and is labelled with {sup 32}P. So far, DNA probes have been successfully employed in the diagnosis of inherited disorders, infectious diseases, and for identification of human oncogenes. The latest approach to the diagnosis of communicable and parasitic infections is based on the use of deoxyribonucleic acid (DNA) probes. The genetic information of all cells is encoded by DNA and DNA probe approach to identification of pathogens is unique because the focus of the method is the nucleic acid content of the organism rather than the products that the nucleic acid encodes. Since every properly classified species has some unique nucleotide sequences that distinguish it from every other species, each organism`s genetic composition is in essence a finger print that can be used for its identification. In addition to this specificity, DNA probes offer other advantages in that pathogens may be identified directly in clinical specimens 10 figs, 2 tabs

  10. DNA methylation

    DEFF Research Database (Denmark)

    Williams, Kristine; Christensen, Jesper; Helin, Kristian

    2012-01-01

    DNA methylation is involved in key cellular processes, including X-chromosome inactivation, imprinting and transcriptional silencing of specific genes and repetitive elements. DNA methylation patterns are frequently perturbed in human diseases such as imprinting disorders and cancer. The recent...... discovery that the three members of the TET protein family can convert 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC) has provided a potential mechanism leading to DNA demethylation. Moreover, the demonstration that TET2 is frequently mutated in haematopoietic tumours suggests that the TET...... proteins are important regulators of cellular identity. Here, we review the current knowledge regarding the function of the TET proteins, and discuss various mechanisms by which they contribute to transcriptional control. We propose that the TET proteins have an important role in regulating DNA methylation...

  11. DNA data

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Raw DNA chromatogram data produced by the ABI 373, 377, 3130 and 3730 automated sequencing machines in ABI format. These are from fish (primarily Sebastes spp.,...

  12. DNA nanotechnology

    Science.gov (United States)

    Seeman, Nadrian C.; Sleiman, Hanadi F.

    2018-01-01

    DNA is the molecule that stores and transmits genetic information in biological systems. The field of DNA nanotechnology takes this molecule out of its biological context and uses its information to assemble structural motifs and then to connect them together. This field has had a remarkable impact on nanoscience and nanotechnology, and has been revolutionary in our ability to control molecular self-assembly. In this Review, we summarize the approaches used to assemble DNA nanostructures and examine their emerging applications in areas such as biophysics, diagnostics, nanoparticle and protein assembly, biomolecule structure determination, drug delivery and synthetic biology. The introduction of orthogonal interactions into DNA nanostructures is discussed, and finally, a perspective on the future directions of this field is presented.

  13. DNA expressions - A formal notation for DNA

    NARCIS (Netherlands)

    Vliet, Rudy van

    2015-01-01

    We describe a formal notation for DNA molecules that may contain nicks and gaps. The resulting DNA expressions denote formal DNA molecules. Different DNA expressions may denote the same molecule. Such DNA expressions are called equivalent. We examine which DNA expressions are minimal, which

  14. Genome sequence and description of Paenibacillus ihuae strain GD6 sp. nov., isolated from the stool of a 62-year-old Frenchman

    Directory of Open Access Journals (Sweden)

    C. Al-Bayssari

    2018-05-01

    Full Text Available Paenibacillus ihuae strain GD6 (=CSUR P892 = DSMZ 45751T is the new type strain collected from the stool of a 69-year-old Frenchman admitted to an intensive care unit and receiving a 10-day course of imipenem at the time of stool collection. This is a Gram-positive, facultative anaerobic, rod-shaped bacterium. We describe here the features of this organism, together with its complete genome sequence and annotation. The genome size is 6 719 043 bp with 49.6% G+C content and contains 6211 protein-coding and 65 sRNA genes, including four 5S rRNA genes, one 16S rRNA gene and one 23S rRNA gene. Keywords: Culturomics, Paenibacillus ihuae, taxonogenomics

  15. Isolation of human genomic DNA for genetic analysis from premature neonates: a comparison between newborn dried blood spots, whole blood and umbilical cord tissue

    Science.gov (United States)

    2013-01-01

    Background Genotyping requires biological sample collection that must be reliable, convenient and acceptable for patients and clinicians. Finding the most optimal procedure of sample collection for premature neonates who have a very limited blood volume is a particular challenge. The aim of the current study was to evaluate the use of umbilical cord (UC) tissue and newborn dried blood spot (DBS)-extracted genomic DNA (gDNA) as an alternative to venous blood-derived gDNA from premature neonates for molecular genetic analysis. All samples were obtained from premature newborn infants between 24-32 weeks of gestation. Paired blood and UC samples were collected from 31 study participants. gDNA was extracted from ethylenediaminetetraacetic acid (EDTA) anticoagulant-treated blood samples (~500 μl) and newborn DBSs (n = 723) using QIAamp DNA Micro kit (Qiagen Ltd., Crawley, UK); and from UC using Qiagen DNAeasy Blood and Tissue kit (Qiagen Ltd., Crawley, UK). gDNA was quantified and purity confirmed by measuring the A260:A280 ratio. PCR amplification and pyrosequencing was carried out to determine suitability of the gDNA for molecular genetic analysis. Minor allele frequency of two unrelated single nucleotide polymorphisms (SNPs) was calculated using the entire cohort. Results Both whole blood samples and UC tissue provided good quality and yield of gDNA, which was considerably less from newborn DBS. The gDNA purity was also reduced after 3 years of storage of the newborn DBS. PCR amplification of three unrelated genes resulted in clear products in all whole blood and UC samples and 86%-100% of newborn DBS. Genotyping using pyrosequencing showed 100% concordance in the paired UC and whole blood samples. Minor allele frequencies of the two SNPs indicated that no maternal gDNA contamination occurred in the genotyping of the UC samples. Conclusions gDNAs from all three sources are suitable for standard PCR and pyrosequencing assays. Given that UC provide good quality

  16. Patients with gastrointestinal complains due to enteric parasites, with reference to Entamoeba histolytica/dispar as dected by ELISA E. histolytica adhesion in stool.

    Science.gov (United States)

    El-Kadi, Mohammad A; Dorrah, Ahmad O; Shoukry, Nahla M

    2006-04-01

    A total of 210 patients with gastrointestinal troubles, of both sex and a mean age of 32 +/- 6.1 years, selected from the outpatient's clinics of Al-Azhar University Hospitals. 115 (54.76%) had dysentery, 95 (45.23%) did not have dysentery, 15 (14%) suffered flatulence, 20 (9.52%) had epi-gastric pain, 19 (9.05%) had vague abdominal pain, 5 vomiting (5.2%) and 10 (4.9%) had fever. Two symptoms were in 29 (13.81%) patients and three symptoms in 12 (5.71%). Of the 210 patients, 20 (9.9%) had helminthes infection, 121 (57.6%) had intestinal protozoa and 69 (32.9%) had no parasitic infection. Of these parasite-free patients, 16 had Shigella sp. and nine had Campylobacter sp. Of the patients with intestinal protozoa, 34 (16.2%) had E. histolytica/dispar by stool examination of stained smears. By using ELISA for detection of E. histolytica adhesion in stool samples of 115 with diarrhea only 18 had true E. histolytica infection and of 3 without diarrhea only one had E. histolytica infection. Mean-while, ELISA did not cross-reacted E. coli, Giardia lamblia, Cryptosporidium parvum, Endolimax nana or Blastocystis hominis. So, ELISA for detection of E. histolytica adhesion in stool samples was more specific than microscopy and safe direction to the E. histolytica treatment. Apart from intestinal protozoan and bacteria, helminthes were seen in stool analysis. These were Schistosoma mansoni (0.95%), Capillaria sp. (0.95%), Enterobius vermicularis (1.90%) macroscopically, Hymenolepis nana (4.3%) and Ascaris lumbricoides (1.43%).

  17. Comparison of the compositions of the stool microbiotas of infants fed goat milk formula, cow milk-based formula, or breast milk.

    Science.gov (United States)

    Tannock, Gerald W; Lawley, Blair; Munro, Karen; Gowri Pathmanathan, Siva; Zhou, Shao J; Makrides, Maria; Gibson, Robert A; Sullivan, Thomas; Prosser, Colin G; Lowry, Dianne; Hodgkinson, Alison J

    2013-05-01

    The aim of the study was to compare the compositions of the fecal microbiotas of infants fed goat milk formula to those of infants fed cow milk formula or breast milk as the gold standard. Pyrosequencing of 16S rRNA gene sequences was used in the analysis of the microbiotas in stool samples collected from 90 Australian babies (30 in each group) at 2 months of age. Beta-diversity analysis of total microbiota sequences and Lachnospiraceae sequences revealed that they were more similar in breast milk/goat milk comparisons than in breast milk/cow milk comparisons. The Lachnospiraceae were mostly restricted to a single species (Ruminococcus gnavus) in breast milk-fed and goat milk-fed babies compared to a more diverse collection in cow milk-fed babies. Bifidobacteriaceae were abundant in the microbiotas of infants in all three groups. Bifidobacterium longum, Bifidobacterium breve, and Bifidobacterium bifidum were the most commonly detected bifidobacterial species. A semiquantitative PCR method was devised to differentiate between B. longum subsp. longum and B. longum subsp. infantis and was used to test stool samples. B. longum subsp. infantis was seldom present in stools, even of breast milk-fed babies. The presence of B. bifidum in the stools of breast milk-fed infants at abundances greater than 10% of the total microbiota was associated with the highest total abundances of Bifidobacteriaceae. When Bifidobacteriaceae abundance was low, Lachnospiraceae abundances were greater. New information about the composition of the fecal microbiota when goat milk formula is used in infant nutrition was thus obtained.

  18. Comparison of 2 chromogenic media for the detection of extended-spectrum β-lactamase producing Enterobacteriaceae stool carriage in nursing home residents.

    Science.gov (United States)

    Blane, Beth; Brodrick, Hayley J; Gouliouris, Theodore; Ambridge, Kirsty E; Kidney, Angela D; Ludden, Catherine M; Limmathurotsakul, Direk; Török, M Estée; Peacock, Sharon J

    2016-03-01

    ChromID ESBL agar and Brilliance ESBL agar were compared for the isolation of extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae from 298 stools. These had comparable sensitivity and selectivity for the 116 positive samples. Pre-enrichment with cefpodoxime and extending incubation to 48 hours after direct plating both significantly increased sensitivity but reduced selectivity of both agars. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  19. Analysis of 13C-mixed triacylglycerol in stool by bulk (EA-IRMS) and compound specific (GC/MS) methods.

    Science.gov (United States)

    Slater, C; Ling, S C; Preston, T; Weaver, L T

    2002-06-01

    This paper was presented in poster form at the 17th International Congress of Nutrition, August 27-31, Vienna, Austria (Annals of Nutrition & Metabolism 2001; 45(Suppl.1):349). Some of the data were also presented in poster form at the British Society of Gastroenterology Meeting, March 18-21, Glasgow, UK (Gut 2001; 48(Suppl.1):A91). The 13C-mixed triacylglycerol (MTG) breath test is used to measure intraluminal fat digestion. In normal digestion, 20-40% of the ingested 13C label is recovered in breath CO2. We aimed to identify the proportions of ingested label excreted in stool, as well as breath following ingestion of 13C-MTG by children with impaired exocrine pancreatic function and healthy controls. 13C enrichment of breath samples was measured by continuous flow isotope ratio mass spectrometry (IRMS) and cumulative percent dose recovered (cPDR) in 10 h was calculated. Total 13C of a faecal fat extract from each stool was measured by elemental analyser-IRMS, and 13C enrichment and concentration of the TBDMS derivative of octanoic acid was measured by GC/MS after hydrolysis of the fat extract. Stool 5-day cPDR was calculated. Mean breath cPDR was 35%. Mean cPDR in stool by combustion-IRMS and GC/ MS, respectively, was 0.8% and 1.0%. Therefore, the remaining 64% of the 13C label must remain in the body and variability in breath cPDR is due to postabsorptive rather than predigestive factors.

  20. Detection of enteropathogens associated with travelers’ diarrhea using a multiplex Luminex-based assay performed on stool samples smeared on Whatman FTA Elute cards

    Science.gov (United States)

    Lalani, Tahaniyat; Tisdale, Michele D; Maguire, Jason D; Wongsrichanalai, Chansuda; Riddle, Mark S; Tribble, David R

    2015-01-01

    We evaluated the limits of detection (LoD) for an 11-plex PCR-Luminex assay performed on Whatman FTA Elute cards smeared with stool containing pathogens associated with travelers’ diarrhea. LoDs ranged between 102-105 CFU, PFU or cysts/g for most pathogens except Cryptosporidium. Campylobacter and norovirus LoD increased with prolonged storage of cards. PMID:26072151

  1. Detection of enteropathogens associated with travelers' diarrhea using a multiplex Luminex-based assay performed on stool samples smeared on Whatman FTA Elute cards.

    Science.gov (United States)

    Lalani, Tahaniyat; Tisdale, Michele D; Maguire, Jason D; Wongsrichanalai, Chansuda; Riddle, Mark S; Tribble, David R

    2015-09-01

    We evaluated the limits of detection (LoD) for an 11-plex PCR-Luminex assay performed on Whatman(™) FTA Elute cards smeared with stool containing pathogens associated with travelers' diarrhea. LoDs ranged from 10(2) to 10(5)CFU, PFU, or cysts/g for most pathogens except Cryptosporidium. Campylobacter and norovirus LoDs increased with prolonged storage of cards. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. What Is Mitochondrial DNA?

    Science.gov (United States)

    ... DNA What is mitochondrial DNA? What is mitochondrial DNA? Although most DNA is packaged in chromosomes within ... proteins. For more information about mitochondria and mitochondrial DNA: Molecular Expressions, a web site from the Florida ...

  3. Novel artificial stool material for external quality assurance (EQA) on a fecal immunochemical test for hemoglobin (FIT): The confirmed utility of stable hemoglobin and an internal standard material.

    Science.gov (United States)

    Yasui, Ryota; Yamada, Miyu; Takehara, Shizuka; Sakurabayashi, Ikunosuke; Watanabe, Katsunori

    2018-04-16

    The fecal immunochemical test for hemoglobin (FIT), which detects lower gastrointestinal bleeding, is widely accepted for population-based colorectal cancer (CRC) screening programs. However, the FIT screening process has not been standardized yet, and standardizing the pre-analytical phase and establishing an external quality assurance (EQA) program compliant with ISO requirements is urgently needed. Although there have been various attempts to establish EQA materials suitable for FIT, no materials have yet been reported to have sufficient uniformity and acceptable immunochemical stability of hemoglobin (Hb). The Health Care Technology Foundation (HECTEF; Tokyo Japan) is now developing a ready-to-use artificial stool containing Hb and an internal standard, glycerol. Accordingly, we verified the adaptability and efficacy of this material for the evaluation of the specimen collection phase of FIT. This material uniformly contained both Hb and glycerol. The glycerol allowed us to estimate the weight of the collected artificial stool and to correct the Hb concentration with the estimated weight. Furthermore, the stability of both Hb and glycerol were confirmed to be sufficient for an EQA material under appropriate storage, in-use, repeated freeze-thaw, and heated conditions. These in-house performance characteristics suggest that HECTEF artificial stool is acceptable as an EQA material for FIT. Copyright © 2018 Elsevier B.V. All rights reserved.

  4. Determination of 14CO2 in breath and 14C in stool after oral administration of cholyl-1-[14C]glycine: clinical application

    International Nuclear Information System (INIS)

    Roda, A.; Roda, E.; Aldini, R.; Mazzella, G.; Festi, D.; Sama, C.; Barbara, L.

    1977-01-01

    Twenty patients with intestinal bacterial overgrowth and 20 control subjects were investigated for bile acid deconjugation, by measuring 14 CO 2 in the breath after cholyl-1-[ 14 C]glycine administration. 14 CO 2 output/24 h was 11.0 +- 5.2% (mean +- SD) in controls and 54.2 +- 14.0% (mean +- SD) in bacterial-overgrowth patients (P 14 CO 2 excretion rate in 12 h, when normalized to 100% of the dose at the 12th hour, gave an even finer discrimination between the two groups (no false responses). 14 C in stool, analyzed in 20 malabsorption patients and 20 controls by two different techniques, was 6.6 +- 4% and 31.38 +- 20.7% (mean +- SD), respectively. Results by the two different techniques described here correlated well (r = 0.99). Bile acid malabsorption was in reasonable agreement (r = 0.67) with percentage of ''chenoid'' (chenodeoxycholic acid plus ursodeoxycholic acid) in the stool by gas-liquid chromatography; a poorer correlation was observed when ''choloid'' (cholic acid plus its epimers) were plotted vs. 14 C in stool

  5. Detection and differentiation of Cryptosporidium by real-time polymerase chain reaction in stool samples from patients in Rio de Janeiro, Brazil

    Directory of Open Access Journals (Sweden)

    Roberta Flávia Ribeiro Rolando

    2012-06-01

    Full Text Available This study reports the first genetic characterisation of Cryptosporidium isolates in Brazil using real-time polymerase chain reaction (RT-PCR. A total of 1,197 faecal specimens from children and 10 specimens from human immunodeficiency virus-infected patients were collected between 1999-2010 and screened using microscopy. Forty-eight Cryptosporidium oocyst-positive isolates were identified and analysed using a generic TaqMan assay targeting the 18S rRNA to detect Cryptosporidium species and two other TaqMan assays to identify Cryptosporidium hominis and Cryptosporidium parvum. The 18S rRNA assay detected Cryptosporidium species in all 48 of the stool specimens. The C. parvum TaqMan assay correctly identified five/48 stool samples, while 37/48 stool specimens were correctly amplified in the C. hominis TaqMan assay. The results obtained in this study support previous findings showing that C. hominis infections are more prevalent than C. parvum infections in Brazil and they demonstrate that the TaqMan RT-PCR procedure is a simple, fast and valuable tool for the detection and differentiation of Cryptosporidium species.

  6. Teaching retirement financial literacy in an undergraduate gerontology classroom: broadening the concept of the tripod or three-legged stool of retirement income utilizing active learning.

    Science.gov (United States)

    Baker, Hallie E; Brown, Pamela Pitman

    2015-01-01

    The three-legged stool concept is widely used in gerontological and geriatric education as an explanation on how one should fiscally approach his or her retirement. Financial managers, planners, retirees, business owners, even the Social Security Administration uses this metaphor of fiscal soundness in retirement planning. Gerontologists are moving away from the "tripod of retirement income" and "three-legged stool" term, as more often market work is needed for financial security. This activity focuses on the tripod or three-legged stool concepts of retirement planning using active learning, allowing the students to work collaboratively in a group, reflect upon the activity, and most importantly have fun. The game also allows for an expansion of the tripod concepts into the four pillars of economic security, broaching the use of personal assets and the possible need for longer employment. Game scenarios also emphasize macro- and microlevel forces, such as race, gender, health status, education, or marital status, which can influence timing of retirement or the level of retirement income available. The authors include instructions on how to set up the learning experience including worksheets, as well as reflection questions posed throughout the process.

  7. Does the amount of tagged stool and fluid significantly affect the radiation exposure in low-dose CT colonography performed with an automatic exposure control?

    Energy Technology Data Exchange (ETDEWEB)

    Lim, Hyun Kyong; Lee, Kyoung Ho; Kim, So Yeon; Kim, Young Hoon [Seoul National University Bundang Hospital, Department of Radiology, Seongnam-si, Gyeonggi-do (Korea, Republic of); Seoul National University College of Medicine, Seoul National University Medical Research Center, Institute of Radiation Medicine, Bundang (Korea, Republic of); Kim, Kil Joong [Seoul National University College of Medicine, Department of Radiation Applied Life Science, Seoul (Korea, Republic of); Kim, Bohyoung; Lee, Hyunna [Seoul National University, School of Computer Science and Engineering, Seoul (Korea, Republic of); Park, Seong Ho [University of Ulsan College of Medicine, Department of Radiology and Research Institute of Radiology, Asan Medical Center, Seoul (Korea, Republic of); Yanof, Jeffrey H. [Philips Healthcare, CT Clinical Science, Cleveland, OH (United States); Hwang, Seung-sik [Inha University School of Medicine, Department of Social and Preventive Medicine, Incheon (Korea, Republic of)

    2011-02-15

    To determine whether the amount of tagged stool and fluid significantly affects the radiation exposure in low-dose screening CT colonography performed with an automatic tube-current modulation technique. The study included 311 patients. The tagging agent was barium (n = 271) or iodine (n = 40). Correlation was measured between mean volume CT dose index (CTDI{sub vol}) and the estimated x-ray attenuation of the tagged stool and fluid (ATT). Multiple linear regression analyses were performed to determine the effect of ATT on CTDI{sub vol} and the effect of ATT on image noise while adjusting for other variables including abdominal circumference. CTDI{sub vol} varied from 0.88 to 2.54 mGy. There was no significant correlation between CTDI{sub vol} and ATT (p = 0.61). ATT did not significantly affect CTDI{sub vol} (p = 0.93), while abdominal circumference was the only factor significantly affecting CTDI{sub vol} (p < 0.001). Image noise ranged from 59.5 to 64.1 HU. The p value for the regression model explaining the noise was 0.38. The amount of stool and fluid tagging does not significantly affect radiation exposure. (orig.)

  8. Utility of a stool antigen test to detect the incidence of helicobacter pylori infection and familial and community enviromental risk factors for this infection in pediatric age.

    Science.gov (United States)

    Sabbi, T; Dall'Oglio, L; De Angelis, P; Torroni, E; Colistro, F; Azzolina, M; Santoni, A; Di Ciommo, V; Benedetto, M

    2012-01-01

    Helicobacter pylori (Hp) infection is mainly acquired during childhood; it is recognised as a cause of gastritis and peptic ulcer and it has been classified as a group A carcinogen by World Health Organization. The exact mode of transmission is as yet, not known. Aim of our study has been to identify risk factors associated with Helicobacter pylori infection in a preschool and school population and to confirm if Hp antigen in faeces is useful as screening in epidemiological studies. We interviewed, with questionnaire, 400 children (203 male; age range 3-10 years; mean age 6 years) of 3 different schools and stool samples were collected of all children too. 35 of 400 (8%) children underwent to upper gastrointestinal endoscopy because of a suspect of upper gastrointestinal disease. stool were collected from 400 school children and 35 of them shown positivity of Hp antigen test. A questionnaire about presence of nausea, vomit, recurrent abdominal pain, family size, parent's occupations and education, use of antibiotics, country of birth of child and parents, personal hygiene, breast feeding, presence of the animals was completed. 35 children with positive Hp stool antigen test and a suspicious of upper gastrointestinal disease (recurrent abdominal pain, diurnal or nocturnal abdominal pain, nausea, vomiting, iron deficiency) underwent to esophagogastroduodenoscopy (EGDS) that demonstrated antral gastritis and positive histology and urease rapid test. the results of this study suggest that risk factors for Hp infection are low socioeconomics factors, hygiene and living conditions and that Hp antigen in faeces is useful as screening test.

  9. Does the amount of tagged stool and fluid significantly affect the radiation exposure in low-dose CT colonography performed with an automatic exposure control?

    International Nuclear Information System (INIS)

    Lim, Hyun Kyong; Lee, Kyoung Ho; Kim, So Yeon; Kim, Young Hoon; Kim, Kil Joong; Kim, Bohyoung; Lee, Hyunna; Park, Seong Ho; Yanof, Jeffrey H.; Hwang, Seung-sik

    2011-01-01

    To determine whether the amount of tagged stool and fluid significantly affects the radiation exposure in low-dose screening CT colonography performed with an automatic tube-current modulation technique. The study included 311 patients. The tagging agent was barium (n = 271) or iodine (n = 40). Correlation was measured between mean volume CT dose index (CTDI vol ) and the estimated x-ray attenuation of the tagged stool and fluid (ATT). Multiple linear regression analyses were performed to determine the effect of ATT on CTDI vol and the effect of ATT on image noise while adjusting for other variables including abdominal circumference. CTDI vol varied from 0.88 to 2.54 mGy. There was no significant correlation between CTDI vol and ATT (p = 0.61). ATT did not significantly affect CTDI vol (p = 0.93), while abdominal circumference was the only factor significantly affecting CTDI vol (p < 0.001). Image noise ranged from 59.5 to 64.1 HU. The p value for the regression model explaining the noise was 0.38. The amount of stool and fluid tagging does not significantly affect radiation exposure. (orig.)

  10. The Bristol Stool Form Scale: its translation to Portuguese, cultural adaptation and validation Traducción, adaptación cultural y validación de la "Bristol Stool Form Scale" Tradução, adaptação cultural e validação da Bristol Stool Form Scale para a população brasileira

    Directory of Open Access Journals (Sweden)

    Anna Paula Martinez

    2012-06-01

    Full Text Available The Bristol Stool Form Scale is used for describing feces. The objective of this research was its translation, cultural adaptation and validation for Brazil. The methodology was translation, back-translation and discussion. Validation involved 85 nurses, 80 doctors, and 80 patients, who correlated images of seven types of feces with the descriptions. Results: there was a difference in sex distribution, with males predominating among the doctors and females among nurses and patients. In relation to concordance between definitions and pictures, the highest percentage was in type 5 in all three groups and the lowest was in types 6 and 7 for the doctors, in type 3 for the nurses, and type 6 for the patients. The general Kappa index was 0.826. Conclusion: the scale demonstrated high reliability for all the groups studied.La "Bristol Stool Form Scale" es usada para describir las heces. Objetivo: traducción, adaptación cultural y la validación para ser utilizada en Brasil. Metodología: Fue realizada la traducción, la traducción inversa y la discusión final. Para validar, se incluyeron 85 enfermeros y 80 médicos y pacientes que correlacionaron diseños de siete tipos de heces con descripciones. Resultados - Hubo diferencia en cuanto a la distribución del sexo con predominio masculino entre los médicos y femenino para los enfermeros y pacientes. Con respecto a la concordancia entre los conceptos y las imágenes, la mayor concordancia fue del tipo 5 en cuanto que el de menor correspondencia para los médicos fueron los tipos 6 y 7, para los enfermeros el 3 y el 6 para los pacientes. El índice de Kappa general fue de 0,826. Conclusión: Los valores obtenidos demuestran la alta confiabilidad de este cuestionario con respecto a los grupos estudiados.A Escala de Bristol para Consistência de Fezes é usada na descrição de fezes. O objetivo deste estudo foi realizar a tradução, adaptação cultural e validação para o Brasil, dessa escala. Como

  11. Comparison of individual and pooled stool samples for the assessment of intensity of Schistosoma mansoni and soil-transmitted helminth infections using the Kato-Katz technique.

    Science.gov (United States)

    Kure, Ashenafi; Mekonnen, Zeleke; Dana, Daniel; Bajiro, Mitiku; Ayana, Mio; Vercruysse, Jozef; Levecke, Bruno

    2015-09-24

    Our group has recently provided a proof-of-principle for the examination of pooled stool samples using McMaster technique as a strategy for the rapid assessment of intensity of soil-transmitted helminth infections (STH, Ascaris lumbricoides, Trichuris trichiura and hookworm). In the present study we evaluated this pooling strategy for the assessment of intensity of both STH and Schistosoma mansoni infections using the Kato-Katz technique. A cross-sectional survey was conducted in 360 children aged 5-18 years from six schools in Jimma Zone (southwest Ethiopia). We performed faecal egg counts (FECs) in both individual and pooled samples (pools sizes of 5, 10 and 20) to estimate the number of eggs per gram of stool (EPG) using the Kato-Katz technique. We also assessed the time to screen both individual and pooled samples. Except for hookworms, there was a significant correlation (correlation coefficient = 0.53-0.95) between the mean of individual FECs and the FECs of pooled samples for A. lumbricoides, T. trichiura and S. mansoni, regardless of the pool size. Mean FEC were 2,596 EPG, 125 EPG, 47 EPG, and 41 EPG for A. lumbricoides, T. trichiura, S. mansoni and hookworm, respectively. There was no significant difference in FECs between the examination of individual and pooled stool samples, except for hookworms. For this STH, pools of 10 resulted in a significant underestimation of infection intensity. The total time to obtain individual FECs was 65 h 5 min. For pooled FECs, this was 19 h 12 min for pools of 5, 14 h 39 min for pools of 10 and 12 h 42 min for pools of 20. The results indicate that pooling of stool sample holds also promise as a rapid assessment of infections intensity for STH and S. mansoni using the Kato-Katz technique. In this setting, the time in the laboratory was reduced by 70 % when pools of 5 instead of individual stool samples were screened.

  12. Identification and characterization of unrecognized viruses in stool samples of non-polio acute flaccid paralysis children by simplified VIDISCA

    NARCIS (Netherlands)

    Shaukat, Shahzad; Angez, Mehar; Alam, Muhammad Masroor; Jebbink, Maarten F.; Deijs, Martin; Canuti, Marta; Sharif, Salmaan; de Vries, Michel; Khurshid, Adnan; Mahmood, Tariq; van der Hoek, Lia; Zaidi, Syed Sohail Zahoor

    2014-01-01

    The use of sequence independent methods combined with next generation sequencing for identification purposes in clinical samples appears promising and exciting results have been achieved to understand unexplained infections. One sequence independent method, Virus Discovery based on cDNA Amplified

  13. DNA Vaccines

    Indian Academy of Sciences (India)

    diseases. Keywords. DNA vaccine, immune response, antibodies, infectious diseases. GENERAL .... tein vaccines require expensive virus/protein purification tech- niques as ... sphere continue to remain major health hazards in developing nations. ... significance since it can be produced at a very low cost and can be stored ...

  14. DNA Investigations.

    Science.gov (United States)

    Mayo, Ellen S.; Bertino, Anthony J.

    1991-01-01

    Presents a simulation activity that allow students to work through the exercise of DNA profiling and to grapple with some analytical and ethical questions involving a couple arranging with a surrogate mother to have a baby. Can be used to teach the principles of restriction enzyme digestion, gel electrophoresis, and probe hybridization. (MDH)

  15. Comparação de seis métodos de extração de DNA genômico de Giardia duodenalis

    Directory of Open Access Journals (Sweden)

    Paulo Henrique Exterchoter Weiss

    2016-05-01

    Methods: Suitable DNA samples for genotypic amplification protocols were obtained by means of PCR, comparing six methods for DNA extraction from G. duodenalis cysts in purified and unpurified samples: QIAmp DNA Stool Mini kit, freeze-thaw procedure, sonication glass bead disruption, formamide denaturation, and conventional method (phenol/chloroform.Results: The methods of sonication and the use of glass beads were more effective in extracting DNA. There was no difference between the use of purified and unpurified samples for protozoan DNA extraction.Conclusions: In the present study we verified that both purified and unpurified samples can be used to G. duodenalis DNA extraction.Though various DNA extraction methods are recommended in the literature, the use of sonicator and glass beads were more effective in this study.

  16. Evaluation of RIDA®GENE norovirus GI/GII real time RT-PCR using stool specimens collected from children and adults with acute gastroenteritis.

    Science.gov (United States)

    Kanwar, N; Hassan, F; Barclay, L; Langley, C; Vinjé, J; Bryant, P W; George, K St; Mosher, L; Matthews-Greer, J M; Rocha, M A; Beenhouwer, D O; Harrison, C J; Moffatt, M; Shastri, N; Selvarangan, R

    2018-04-10

    Norovirus is the leading cause of epidemic and sporadic acute gastroenteritis (AGE) in the United States. Widespread prevalence necessitates implementation of accurate norovirus detection assays in clinical diagnostic laboratories. To evaluate RIDA ® GENE norovirus GI/GII real-time RT-PCR assay (RGN RT-PCR) using stool samples from patients with sporadic AGE. Patients between 14 days to 101 years of age with symptoms of AGE were enrolled prospectively at four sites across the United States during 2014-2015. Stool specimens were screened for the presence of norovirus RNA by the RGN RT-PCR assay. Results were compared with a reference method that included conventional RT-PCR and sequencing of a partial region of the 5'end of the norovirus ORF2 gene. A total of 259 (36.0%) of 719 specimens tested positive for norovirus by the reference method. The RGN RT-PCR assay detected norovirus in 244 (94%) of these 259 norovirus positive specimens. The sensitivity and specificity (95% confidence interval) of the RGN RT-PCR assay for detecting norovirus genogroup (G) I was 82.8% (63.5-93.5) and 99.1% (98.0-99.6) and for GII was 94.8% (90.8-97.2) and 98.6% (96.9-99.4), respectively. Seven specimens tested positive by the RGN-RT PCR that were negative by the reference method. The fifteen false negative samples were typed as GII.4 Sydney, GII.13, GI.3, GI.5, GI.2, GII.1, and GII.3 in the reference method. The RGN RT-PCR assay had a high sensitivity and specificity for the detection of norovirus in stool specimens from patients with sporadic AGE. Copyright © 2018. Published by Elsevier B.V.

  17. Variation in the limit-of-detection of the ProSpecT Campylobacter microplate enzyme immunoassay in stools spiked with emerging Campylobacter species.

    Science.gov (United States)

    Bojanić, Krunoslav; Midwinter, Anne Camilla; Marshall, Jonathan Craig; Rogers, Lynn Elizabeth; Biggs, Patrick Jon; Acke, Els

    2016-08-01

    Campylobacter enteritis in humans is primarily associated with C. jejuni/coli infection. The impact of other Campylobacter spp. is likely to be underestimated due to the bias of culture methods towards Campylobacter jejuni/coli diagnosis. Stool antigen tests are becoming increasingly popular and appear generally less species-specific. A review of independent studies of the ProSpecT® Campylobacter Microplate enzyme immunoassay (EIA) developed for C. jejuni/coli showed comparable diagnostic results to culture methods but the examination of non-jejuni/coli Campylobacter spp. was limited and the limit-of-detection (LOD), where reported, varied between studies. This study investigated LOD of EIA for Campylobacter upsaliensis, Campylobacter hyointestinalis and Campylobacter helveticus spiked in human stools. Multiple stools and Campylobacter isolates were used in three different concentrations (10(4)-10(9)CFU/ml) to reflect sample heterogeneity. All Campylobacter species evaluated were detectable by EIA. Multivariate analysis showed LOD varied between Campylobacter spp. and faecal consistency as fixed effects and individual faecal samples as random effects. EIA showed excellent performance in replicate testing for both within and between batches of reagents, in agreement between visual and spectrophotometric reading of results, and returned no discordance between the bacterial concentrations within independent dilution test runs (positive results with lower but not higher concentrations). This study shows how limitations in experimental procedures lead to an overestimation of consistency and uniformity of LOD for EIA that may not hold under routine use in diagnostic laboratories. Benefits and limitations for clinical practice and the influence on estimates of performance characteristics from detection of multiple Campylobacter spp. by EIA are discussed. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Utility of a stool antigen test to detect the incidence of helicobacter pylori infection and familial and community enviromental risk factors for this infection in pediatric age

    Directory of Open Access Journals (Sweden)

    T. Sabbi

    2012-04-01

    Full Text Available Background: helicobacter pylori (hp infection is mainly acquired during childhood; it is recognised as a cause of gastritis and peptic ulcer and it has been classified as a group A carcinogen by World health organization. the exact mode of transmission is as yet, not known. Aim of our study has been to identify risk factors associated with helicobacter pylori infection in a preschool and school population and to confirm if hp antigen in faeces is useful as screening in epidemiological studies. Methods: We interviewed, with questionnaire, 400 children (203 male; age range 3-10 years; mean age 6 years of 3 different schools and stool samples were collected of all children too. 35 of 400 (8% children underwent to upper gastrointestinal endoscopy because of a suspect of upper gastrointestinal disease. Results: stool were collected from 400 school children and 35 of them shown positivity of hp antigen test. A questionnaire about presence of nausea, vomit, recurrent abdominal pain, family size, parent’s occupations and education, use of antibiotics, country of birth of child and parents, personal hygiene, breast feeding, presence of the animals was completed. 35 children with positive hp stool antigen test and a suspicious of upper gastrointestinal disease (recurrent abdominal pain, diurnal or nocturnal abdominal pain, nausea, vomiting, iron deficiency underwent to esophagogastroduodenoscopy (egdS that demonstrated antral gastritis and positive histology and urease rapid test. Conclusions: the results of this study suggest that risk factors for hp infection are low socioeconomics factors, hygiene and living conditions and that hp antigen in faeces is useful as screening test.

  19. Pain symptoms and stooling patterns do not drive diagnostic costs for children with functional abdominal pain and irritable bowel syndrome in primary or tertiary care.

    Science.gov (United States)

    Lane, Mariella M; Weidler, Erica M; Czyzewski, Danita I; Shulman, Robert J

    2009-03-01

    The objectives of this study were to (1) compare the cost of medical evaluation for children with functional abdominal pain or irritable bowel syndrome brought to a pediatric gastroenterologist versus children who remained in the care of their pediatrician, (2) compare symptom characteristics for the children in primary versus tertiary care, and (3) examine if symptom characteristics predicted the cost of medical evaluation. Eighty-nine children aged 7 to 10 years with functional abdominal pain or irritable bowel syndrome seen by a gastroenterologist (n = 46) or seen only by a pediatrician (n = 43) completed daily pain and stool diaries for 2 weeks. Mothers provided retrospective reports of their children's symptoms in the previous year. Cost of medical evaluation was calculated via chart review of diagnostic tests and application of prices as if the patients were self-pay. Child-reported diary data reflected no significant group differences with respect to pain, interference with activities, or stool characteristics. In contrast, mothers of children evaluated by a gastroenterologist viewed their children as having higher maximum pain intensity in the previous year. Excluding endoscopy costs, cost of medical evaluation was fivefold higher for children evaluated by a gastroenterologist, with higher cost across blood work, stool studies, breath testing, and diagnostic imaging. Symptom characteristics did not predict cost of care for either group. Despite the lack of difference in symptom characteristics between children in primary and tertiary care, a notable differential in cost of evaluation exists in accordance with level of care. Symptom characteristics do not seem to drive diagnostic evaluation in either primary or tertiary care. Given the lack of differences in child-reported symptoms and the maternal perspective that children evaluated by a gastroenterologist had more severe pain, we speculate that parent perception of child symptoms may be a primary factor in

  20. DNA repair

    International Nuclear Information System (INIS)

    Setlow, R.

    1978-01-01

    Some topics discussed are as follows: difficulty in extrapolating data from E. coli to mammalian systems; mutations caused by UV-induced changes in DNA; mutants deficient in excision repair; other postreplication mechanisms; kinds of excision repair systems; detection of repair by biochemical or biophysical means; human mutants deficient in repair; mutagenic effects of UV on XP cells; and detection of UV-repair defects among XP individuals

  1. Faecal urobilinogen levels and pH of stools in population groups with different incidence of cancer of the colon, and their possible role in its aetiology.

    OpenAIRE

    Malhotra, S L

    1982-01-01

    Mean faecal urobilinogen levels and the pH of stools were both found to be higher in subjects from a population group at high risk of developing cancer of the colon than in subjects matched for age, sex and socioeconomic status from a low-risk population group. An alkaline reaction of the colon contents seems to have a tumorigenic effect by a direct action on the mucus of the mucous cells. An acidic reaction, on the other hand, appears to be protective. These differences are dependent on the ...

  2. Comparison between the 13C-urea breath test and stool antigen test for the diagnosis of childhood Helicobacter pylori infection

    International Nuclear Information System (INIS)

    Kato, Seiichi; Minoura, Takanori; Iinuma, Kazuie; Konno, Mutsuko; Tajiri, Hitoshi; Matsuhisa, Takeshi

    2004-01-01

    As noninvasive tests for Helicobacter pylori infection, the 13 C-urea breath test (UBT) and stool antigen test have been widely used. In children, however, there are few studies reporting which test shows superior performance. The purpose of this study was to compare the 13 C-UBT and stool antigen test for their accuracy in diagnosing. H. pylori infection in children. A total of 123 Japanese children, ages 2 to 17 years (mean, 12 years) who underwent gastric biopsies for H. pylori infection were studied. The diagnoses included gastritis (n=55), gastric ulcer (n=5), duodenal ulcer (n=20), iron-deficiency anemia (n=7), and other conditions (n=36). The cutoff value of the 13 C-UBT was defined to be 3.5 per mille. The stool antigen test was performed using the H. pylori stool antigen (HpSA) enzyme-linked immunosorbent assay (ELISA) (Premier Platinum HpSA). In 16 patients who received eradication therapy, the 13 C-UBT and HpSA were repeated 2 months after treatment. Based on biopsy tests, 60 children were infected with H. pylori and 63 children were not. For the 13 C-UBT, the sensitivity, specificity, and accuracy were 95.0% (95% confidence interval [CI], 86.1%-99.0%), 98.4% (95% CI, 91.5%-100%), and 96.4% (95% CI, 93.6%-99.9%), respectively. For the HpSA, the sensitivity, specificity, and accuracy were 983.% (95% CI, 90.8%-100%), 98.4% (95% CI, 91.2%-100%), and 98.3% (95% CI, 96.0%-100%), respectively. There were no significant differences between the performance of these two tests. In the assessment of H. pylori eradication, the results of 13 C-UBT and HpSA agreed with those of biopsy tests. The 13 C-UBT and the HpSA are equally accurate for the diagnosis of active H. pylori infection in Japanese children. (author)

  3. Evaluation of four techniques for diagnosis of Giardia lamblia in children's stool from Belém city, Para State, Brazil

    OpenAIRE

    Machado, Ricardo Luiz Dantas; Figueredo, Maria Cristina; Frade, Amanda Farage; Kudó, Mônica Eriko; Silva Filho, Manoel Gomes; Póvoa, Marinete Marins

    2001-01-01

    Relatamos a comparação de quatro metodologias para o diagnóstico da Giardia lamblia em material fecal de crianças, Belém/PA. A Hematoxilina Férrica e o método direto apresentaram menor positividade, enquanto que o Método de Faust continua uma boa escolha para o diagnóstico e o Ensaio imunoenzimático melhora a qualidade da detecção deste parasito.We report the evaluation of four techniques for Giardia lamblia diagnosis in children's stool. The Iron haematoxilin staining and direct examination ...

  4. Multiplex PCR detection of Cryptosporidium sp, Giardia lamblia and Entamoeba histolytica directly from dried stool samples from Guinea-Bissauan children with diarrhoea

    DEFF Research Database (Denmark)

    Mero, Sointu; Kirveskari, Juha; Antikainen, Jenni

    2017-01-01

    and specificity, while new molecular methods may provide more accurate diagnostics. In poor regions with sample storage hampered by uncertain electricity supply, research would benefit from a method capable of analysing dried stools. Methods: A real-time multiplex PCR method with internal inhibition control...... microscopy revealed a parasite in 15%, while PCR detected 62% positive for at least one parasite: 44% of the dried samples had Giardia, 23% Cryptosporidium and 0% E. histolytica. Conclusions: Our new multiplex real-time PCR for protozoa presents a sensitive method applicable to dried samples. As proof...

  5. DNA repair

    International Nuclear Information System (INIS)

    Van Zeeland, A.A.

    1984-01-01

    In this chapter a series of DNA repair pathways are discussed which are available to the cell to cope with the problem of DNA damaged by chemical or physical agents. In the case of microorganisms our knowledge about the precise mechanism of each DNA repair pathway and the regulation of it has been improved considerably when mutants deficient in these repair mechanisms became available. In the case of mammalian cells in culture, until recently there were very little repair deficient mutants available, because in almost all mammalian cells in culture at least the diploid number of chromosomes is present. Therefore the frequency of repair deficient mutants in such populations is very low. Nevertheless because replica plating techniques are improving some mutants from Chinese hamsters ovary cells and L5178Y mouse lymphoma cells are now available. In the case of human cells, cultures obtained from patients with certain genetic diseases are available. A number of cells appear to be sensitive to some chemical or physical mutagens. These include cells from patients suffering from xeroderma pigmentosum, Ataxia telangiectasia, Fanconi's anemia, Cockayne's syndrome. However, only in the case of xeroderma pigmentosum cells, has the sensitivity to ultraviolet light been clearly correlated with a deficiency in excision repair of pyrimidine dimers. Furthermore the work with strains obtained from biopsies from man is difficult because these cells generally have low cloning efficiencies and also have a limited lifespan in vitro. It is therefore very important that more repair deficient mutants will become available from established cell lines from human or animal origin

  6. Detection of Entamoeba histolytica/dispar in stool specimens by using enzyme-linked immunosorbent assay in the population of Jeddah City, Saudi Arabia.

    Science.gov (United States)

    Barnawi, Abdulaziz B M; Tonkal, Abulkader M; Fouad, Mahmoud A H; Al-Braiken, Faten A

    2007-04-01

    This study determined the prevalence of intestinal parasites, particularly pathogenic Entamoeba sp. (E. histolytica), in patients attending three hospitals in Jeddah City, King Abdulaziz University Hospital, King Abdulaziz Hospital and King Fahad Hospital for gastro-intestinal troubles. 186 stool specimens were examined microscopically for parasites and by ELISA kit (E. histolytica II) for true E. histolytica. 83 samples (44.6%) were positive by microscopy for at least one parasite. Of which, 23 (12.4%) showed two parasites and 15 (8.1%) three parasites. Eight different parasite species were identified. The most prevalent were E. histolytica/dispar (n = 26, 31.3%) and Giardia lamblia (n = 13, 15.7%). Others were Blastocytosis hominis (n = 12, 14.5%), Entamoeba coli (n = 11, 13.3%), Trichuris trichuria (n = 8, 9.6%), Endolymax nana (n = 6, 7.2%), Hymenolepes nana (n = 4, 4.8%) and Chilomastix mesnili (n = 3, 3.6%). Only five stool samples (19%) from those identified by microscopy to contain E. histolytica/dispar, were E. histolytica positive by E. histolytica II ELISA. For the first time to the authors' knowledge the true prevalence of E. histolytica in Saudi Arabia was 2.7%. E. histolytica II ELISA proved to be a highly useful technique to differentiate pathogenic E. histolytica from non pathogenic E. dispar.

  7. Distribution of Parasites Detected in Stool Samples of Patients Admitted to Our Parasitology Laboratory during a Three-Year Period between 2012 and 2014.

    Science.gov (United States)

    Selek, Mehmet Burak; Bektöre, Bayhan; Karagöz, Ergenekon; Baylan, Orhan; Özyurt, Mustafa

    2016-09-01

    Parasitic diseases are among the major public health issues worldwide. A number of tests are available for diagnosis, but the sentivity and specifity of these tests are assumed to be insufficient. Nevertheless, the most common diagnostic method is microscopic examination. In this study, we aimed to introduce the distribution of parasites detected in stool samples of patients admitted to our laboratory on the basis of parameters such as, age, and gender during a 3-year period between 2012 and 2014. In total, 6757 stool samples were included in the study. After macroscopic examination, wet mounts of all samples were examined under a light microscope using ×100 and ×400 magnification lenses. Wet mounts were prepared with physiological saline and Lugol's iodine. Parasites were detected in 3.7% (252) of the samples, while no parasites were detected in 96.3% (6505) of the samples. The distribution of intestinal parasites was as follows: Blastocystis hominis (63.5%), Giardia intestinalis (26.2%), Taenia sp. (4.8%), Enterobius vermicularis (2.4%), Entamoeba histolytica/dispar (1.6%), and Hymenolepis nana (1.6%). When the burden of intestinal parasites on public health is considered, they are still a major health issue in Turkey. The frequency of parasitic diseases can be reduced by the education of individuals and implementation of effective diagnostic methods, treatments, and preventive measures.

  8. Association Between Stool Enteropathogen Quantity and Disease in Tanzanian Children Using TaqMan Array Cards: A Nested Case-Control Study

    Science.gov (United States)

    Platts-Mills, James A.; Gratz, Jean; Mduma, Esto; Svensen, Erling; Amour, Caroline; Liu, Jie; Maro, Athanasia; Saidi, Queen; Swai, Ndealilia; Kumburu, Happiness; McCormick, Benjamin J. J.; Kibiki, Gibson; Houpt, Eric R.

    2014-01-01

    Etiologic studies of diarrhea are limited by uneven diagnostic methods and frequent asymptomatic detection of enteropathogens. Polymerase chain reaction-based stool pathogen quantification may help distinguish clinically significant infections. We performed a nested case-control study of diarrhea in infants from a community-based birth cohort in Tanzania. We tested 71 diarrheal samples and pre-diarrheal matched controls with a laboratory-developed TaqMan Array Card for 19 enteropathogens. With qualitative detection, no pathogens were significantly associated with diarrhea. When pathogen quantity was considered, rotavirus (odds ratio [OR] = 2.70 per log10 increase, P < 0.001), astrovirus (OR = 1.49, P = 0.01), and Shigella/enteroinvasive Escherichia coli (OR = 1.47, P = 0.04) were associated with diarrhea. Enterotoxigenic E. coli (0.15 SD decline in length-for-age z score after 3 months per log10 increase, P < 0.001) and Campylobacter jejuni/C. coli (0.11 SD decline, P = 0.003) in pre-diarrheal stools were associated with poor linear growth. Quantitative analysis can help refine the association between enteropathogens and disease in endemic settings. PMID:24189366

  9. sTools - a data reduction pipeline for the GREGOR Fabry-Pérot Interferometer and the High-resolution Fast Imager at the GREGOR solar telescope

    Science.gov (United States)

    Kuckein, C.; Denker, C.; Verma, M.; Balthasar, H.; González Manrique, S. J.; Louis, R. E.; Diercke, A.

    2017-10-01

    A huge amount of data has been acquired with the GREGOR Fabry-Pérot Interferometer (GFPI), large-format facility cameras, and since 2016 with the High-resolution Fast Imager (HiFI). These data are processed in standardized procedures with the aim of providing science-ready data for the solar physics community. For this purpose, we have developed a user-friendly data reduction pipeline called ``sTools'' based on the Interactive Data Language (IDL) and licensed under creative commons license. The pipeline delivers reduced and image-reconstructed data with a minimum of user interaction. Furthermore, quick-look data are generated as well as a webpage with an overview of the observations and their statistics. All the processed data are stored online at the GREGOR GFPI and HiFI data archive of the Leibniz Institute for Astrophysics Potsdam (AIP). The principles of the pipeline are presented together with selected high-resolution spectral scans and images processed with sTools.

  10. Agreement between prospective diary data and retrospective questionnaire report of abdominal pain and stooling symptoms in children with irritable bowel syndrome.

    Science.gov (United States)

    Self, M M; Williams, A E; Czyzewski, D I; Weidler, E M; Shulman, R J

    2015-08-01

    In functional gastrointestinal disorders, patient recall of symptoms drives diagnostic decisions and evaluation of treatment response, and research conclusions about potential treatments. In pediatrics, parent report also impacts assessment and care. Hence, identifying methods for accurately capturing patient and parent report of irritable bowel syndrome (IBS) symptoms is important. This study evaluated correspondence between retrospective questionnaire (parent and child report) and prospective diary data for children and adolescents with IBS. Participants included 50 children/adolescents with IBS per Rome III criteria. Children completed a 2-week pain and stool diary. Children and parents subsequently completed a 2-week recall questionnaire, reporting number of pain days, maximum pain, days without bowel movement, and days with diarrhea during the diary interval. Intraclass correlation coefficients and Bland-Altman plots assessed agreement. For pain and days without bowel movement, overall agreement between child recall questionnaire and child diary was strong, although under conditions likely to facilitate agreement and with individual variation observed. Parent recall and child diary were less concordant, and agreement about diarrhea was poor for parent and child. Age did not significantly correlate with agreement. Child questionnaire with short recall interval may be a reasonable approximation for diary data, although this varies by individual and replication/investigation of lengthier recall are needed. Relying on parent questionnaire does not appear a suitable proxy, and recall of stool form by both parent and child appears more problematic. These results combined with existing literature support use of diary data whenever possible. © 2015 John Wiley & Sons Ltd.

  11. Recombinase Polymerase Amplification Compared to Real-Time Polymerase Chain Reaction Test for the Detection of Fasciola hepatica in Human Stool

    Science.gov (United States)

    Cabada, Miguel M.; Malaga, Jose L.; Castellanos-Gonzalez, Alejandro; Bagwell, Kelli A.; Naeger, Patrick A.; Rogers, Hayley K.; Maharsi, Safa; Mbaka, Maryann; White, A. Clinton

    2017-01-01

    Fasciola hepatica is the most widely distributed trematode infection in the world. Control efforts may be hindered by the lack of diagnostic capacity especially in remote endemic areas. Polymerase chain reaction (PCR)–based methods offer high sensitivity and specificity but require expensive technology. However, the recombinase polymerase amplification (RPA) is an efficient isothermal method that eliminates the need for a thermal cycler and has a high deployment potential to resource-limited settings. We report on the characterization of RPA and PCR tests to detect Fasciola infection in clinical stool samples with low egg burdens. The sensitivity of the RPA and PCR were 87% and 66%, respectively. Both tests were 100% specific showing no cross-reactivity with trematode, cestode, or nematode parasites. In addition, RPA and PCR were able to detect 47% and 26% of infections not detected by microscopy, respectively. The RPA adapted to a lateral flow platform was more sensitive than gel-based detection of the reaction products. In conclusion, the Fasciola RPA is a highly sensitive and specific test to diagnose chronic infection using stool samples. The Fasciola RPA lateral flow has the potential for deployment to endemic areas after further characterization. PMID:27821691

  12. Further development of the radioimmunoassay for carcinoembryonic antigen and the use of the measurement of the carcinoembryonic antigen in stools. Weiterentwicklung des Radioimmunoassay fuer carcino-embryonales Antigen und Anwendung auf die Messung von carcino-embryonalem Antigen im Stuhl

    Energy Technology Data Exchange (ETDEWEB)

    Britsch, R F

    1982-01-01

    A specific, direct CEA-RTA is presented with sequential saturation and double antibody separation and using guinea pig CEA anti-serums, whose isolated CEA was marked with /sup 125/I for tracer production and used as was for a CEA standard. This RIA was tested on serum from patients with colorectal and stomach-esophagus carcinomas and from healthy persons, whereby it showed a good sensitivity in comparison to control serums on the market. CEA is also present in a high concentration in the stools, however very nonhomogeneously distributed, by patients as well as healthy persons. An increased stool CEA value does not necessarily indicate a carcinomatic disease or even a preliminary stage. During a screening a carcinomatic intestinal disorder should not be looked for intensively unless there are first several positive stool CEA values or additional positive blood evidence in the stool. Simultaneously carried out determinations of serum and stool CEA allows for the discovery of more patients as the use of only one. (TRV)

  13. Diffuse and enteroaggregative patterns of adherence of Escherichia coli isolated from stools of children in northeastern Brazil

    Directory of Open Access Journals (Sweden)

    Scaletsky Isabel Cristina Affonso

    2001-01-01

    Full Text Available Childhood diarrheal diseases remain highly endemic in northeastern Brazil. The attributable fraction of all diarrheal diseases among children less than 2 years of age due to Escherichia coli was examined in a 2-year prospective study in two large urban centers of Brazil. Between May 1997 and June 1999, fecal E. coli isolates from 237 children with diarrhea (217 acute and 20 persistent cases and 231 children without diarrhea (controls attending two hospitals in Northeast Brazil were tested for their pattern of adherence to HEp-2 cells and for colony hybridization with DNA probes specific for the six pathotypes of diarrheagenic E. coli. Enteroinvasive E. coli, enterotoxigenic E. coli and enterohemorrhagic E. coli were not isolated from any children. Diffusely adherent E. coli (DAEC and enteroaggregative E. coli (EAEC were the most frequent isolates with similar frequencies from children with or without diarrhea. Atypical EPEC (EAF-negative strains were isolated with similiar frequency from both cases (5.5% and controls (5.6%. Enteropathogenic E. coli (typical EPEC strains, characterized by localized adherence pattern of adherence, hybridization with the EAF probe, and belonging to the classical O serogroups, were significantly associated with diarrhea (P = 0.03. These E. coli strains associated with diarrhea accounted for 9% of all children with diarrhea. Collectively, in Northeast Brazil, E. coli strains comprise a small proportion of severe diarrhea prevalence in children.

  14. Scientific Opinion on the substantiation of a health claim related to “native chicory inulin” and maintenance of normal defecation by increasing stool frequency pursuant to Article 13.5 of Regulation (EC) No 1924/2006

    DEFF Research Database (Denmark)

    Tetens, Inge

    2015-01-01

    claim related to “native chicory inulin” and maintenance of normal defecation by increasing stool frequency. The food constituent that is a subject of a claim is “native chicory inulin”. The Panel considers that “native chicory inulin”, a non-fractionated mixture of monosaccharides (...%), disaccharides, inulin-type fructans and inulin extracted from chicory, with a mean DP ≥ 9, is sufficiently characterised in relation to the claimed effect. The Panel considers that maintenance of normal defecation by increasing stool frequency (provided that it does not result in diarrhoea) is a beneficial...... physiological effect. Six studies involving 86 subjects consistently showed that consumption of “native chicory inulin” at an amount of at least 12 g/day increases stool frequency. The Panel also notes the plausible mechanisms by which inulin and inulin-type fructans in “native chicory inulin” could exert...

  15. DNA Repair Systems

    Indian Academy of Sciences (India)

    DNA molecule which makes it ideal for storage and propagation of genetic information. ... of these errors are broadly referred to as DNA repair. DNA can ... changes occur in the human genome per day. ..... nails, frequent physical and mental.

  16. Synthesis of DNA

    Science.gov (United States)

    Mariella, Jr., Raymond P.

    2008-11-18

    A method of synthesizing a desired double-stranded DNA of a predetermined length and of a predetermined sequence. Preselected sequence segments that will complete the desired double-stranded DNA are determined. Preselected segment sequences of DNA that will be used to complete the desired double-stranded DNA are provided. The preselected segment sequences of DNA are assembled to produce the desired double-stranded DNA.

  17. The origin of endodontic Enterococcus faecalis explored by comparison of virulence factor patterns and antibiotic resistance to that of isolates from stool samples, blood cultures and food.

    Science.gov (United States)

    Vidana, R; Rashid, M U; Özenci, V; Weintraub, A; Lund, B

    2016-04-01

    To elucidate the origin of Enterococcus faecalis isolated from secondary root canal infections and the possibility for a foodborne transmission by comparing them to strains recovered from food, blood and stool regarding putative virulence factors and antibiotic susceptibility profiles, where strains from common origin were hypothesized to harbour similar characteristics. A total of 108 E. faecalis strains recovered in the county of Stockholm, Sweden, were screened using PCR for putative virulence factors esp, cylA, gelE/gelatinase-negative phenotype (ef1841/fsrC), efaA, ace and asa1. The minimum inhibitory concentration (MIC) for ampicillin, piperacillin-tazobactam, imipenem, gentamicin, vancomycin, ciprofloxacin and linezolid was determined using the agar dilution method. Next to strains from blood, the food isolates presented the highest average number of virulence determinants and were frequently enriched with asa1 coding for aggregation substance. None of the endodontic strains carried cylA, and the gelatinase-negative phenotype caused by a deletion dominated the group. Altogether, the most prevalent genes were gelE, efaA and ace, and a combination of them was equally present in approximately 80% of the strains from food, stool and root canals in comparison with 43.3% of the blood isolates. High-level resistance to ciprofloxacin and gentamicin was observed in 30% of the blood isolates, whereas the isolates from other origins, with single exceptions, were susceptible to all tested antibiotics. Evidence for a foodborne transmission, explaining the high reported prevalence of E. faecalis in root filled teeth, could not be determined based on the similarities in virulence factor patterns and antibiotic susceptibility. The only linkage between isolates from food and root canals consisted of a shared common combination of the genes gelE, efaA and ace. The high occurrence of putative virulence traits in food isolates questions the safety of E. faecalis in food

  18. Low cost extraction and isothermal amplification of DNA for infectious diarrhea diagnosis.

    Directory of Open Access Journals (Sweden)

    Shichu Huang

    Full Text Available In order to counter the common perception that molecular diagnostics are too complicated to work in low resource settings, we have performed a difficult sample preparation and DNA amplification protocol using instrumentation designed to be operated without wall or battery power. In this work we have combined a nearly electricity-free nucleic acid extraction process with an electricity-free isothermal amplification assay to detect the presence of Clostridium difficile (C. difficile DNA in the stool of infected patients. We used helicase-dependent isothermal amplification (HDA to amplify the DNA in a low-cost, thermoplastic reaction chip heated with a pair of commercially available toe warmers, while using a simple Styrofoam insulator. DNA was extracted from known positive and negative stool samples. The DNA extraction protocol utilized an air pressure driven solid phase extraction device run using a standard bicycle pump. The simple heater setup required no electricity or battery and was capable of maintaining the temperature at 65°C±2°C for 55 min, suitable for repeatable HDA amplification. Experiments were performed to explore the adaptability of the system for use in a range of ambient conditions. When compared to a traditional centrifuge extraction protocol and a laboratory thermocycler, this disposable, no power platform achieved approximately the same lower limit of detection (1.25×10(-2 pg of C. difficile DNA while requiring much less raw material and a fraction of the lab infrastructure and cost. This proof of concept study could greatly impact the accessibility of molecular assays for applications in global health.

  19. Low Cost Extraction and Isothermal Amplification of DNA for Infectious Diarrhea Diagnosis

    Science.gov (United States)

    Huang, Shichu; Do, Jaephil; Mahalanabis, Madhumita; Fan, Andy; Zhao, Lei; Jepeal, Lisa; Singh, Satish K.; Klapperich, Catherine M.

    2013-01-01

    In order to counter the common perception that molecular diagnostics are too complicated to work in low resource settings, we have performed a difficult sample preparation and DNA amplification protocol using instrumentation designed to be operated without wall or battery power. In this work we have combined a nearly electricity-free nucleic acid extraction process with an electricity-free isothermal amplification assay to detect the presence of Clostridium difficile (C. difficile) DNA in the stool of infected patients. We used helicase-dependent isothermal amplification (HDA) to amplify the DNA in a low-cost, thermoplastic reaction chip heated with a pair of commercially available toe warmers, while using a simple Styrofoam insulator. DNA was extracted from known positive and negative stool samples. The DNA extraction protocol utilized an air pressure driven solid phase extraction device run using a standard bicycle pump. The simple heater setup required no electricity or battery and was capable of maintaining the temperature at 65°C±2°C for 55 min, suitable for repeatable HDA amplification. Experiments were performed to explore the adaptability of the system for use in a range of ambient conditions. When compared to a traditional centrifuge extraction protocol and a laboratory thermocycler, this disposable, no power platform achieved approximately the same lower limit of detection (1.25×10−2 pg of C. difficile DNA) while requiring much less raw material and a fraction of the lab infrastructure and cost. This proof of concept study could greatly impact the accessibility of molecular assays for applications in global health. PMID:23555883

  20. Survey of gastrointestinal reactions to foods in adults in relation to atopy, presence of mucus in the stools, swelling of joints and arthralgia in patients with gastrointestinal reactions to foods.

    Science.gov (United States)

    Bengtsson, U; Hanson, L A; Ahlstedt, S

    1996-12-01

    Food intolerance in adults is mostly associated with vague symptoms and not clearly related to atopy and food allergy. A combination of different pathogenetic mechanisms may be responsible for the symptoms. The aim of this study was to describe patients with a history of food-related gastrointestinal symptoms in relation to the presence of mucus in the stools, joint swelling and arthralgia and to determine whether or not there is an association between the presence of these parameters, atopic disease and the presence of immune complexes in serum. Fifty-eight patients consecutively referred to our clinic with food-related gastrointestinal symptoms were investigated. Thirty-five patients (60%) had mucus in their stools, 24 patients (41%) complained about joint swelling and 41 patients (71%) had arthralgia. There were no correlations between these parameters and atopy according to Phadiatope test or skin prick test (SPT). No correlations were found between the occurrence of mucus in the stools, arthralgia and joint swelling. There were significantly higher levels of circulating immune complexes in patients with a history of arthralgia compared with patients with no such history (P mucus in the stools. However, there were significant positive correlations between food-related gastrointestinal symptoms in the following instances: chocolate-induced gastrointestinal symptoms and mucus in the stools (P = 0.006), vegetable-induced gastrointestinal symptoms and mucus in the stools (P = 0.002) and meat-induced gastrointestinal symptoms and mucus in the stools (P = 0.003). In a group of individuals without food-related symptoms investigated separately, a very low frequency of mucus in the stools, joint swelling and arthralgia was seen (none, two and three individuals of the 20 subjects, respectively). Of 41 patients with immediate onset of gastrointestinal symptoms, 20 were atopic according to Phadiatope and SPT. Of 11 patients with late onset of symptoms 10 were negative in

  1. An optimized work-flow to reduce time-to-detection of carbapenemase-producing Enterobacteriaceae (CPE) using direct testing from rectal swabs.

    Science.gov (United States)

    O'Connor, C; Kiernan, M G; Finnegan, C; O'Hara, M; Power, L; O'Connell, N H; Dunne, C P

    2017-05-04

    Rapid detection of patients with carbapenemase-producing Enterobacteriaceae (CPE) is essential for the prevention of nosocomial cross-transmission, allocation of isolation facilities and to protect patient safety. Here, we aimed to design a new laboratory work-flow, utilizing existing laboratory resources, in order to reduce time-to-diagnosis of CPE. A review of the current CPE testing processes and of the literature was performed to identify a real-time commercial polymerase chain reaction (PCR) assay that could facilitate batch testing of CPE clinical specimens, with adequate CPE gene coverage. Stool specimens (210) were collected; CPE-positive inpatients (n = 10) and anonymized community stool specimens (n = 200). Rectal swabs (eSwab™) were inoculated from collected stool specimens and a manual DNA extraction method (QIAamp® DNA Stool Mini Kit) was employed. Extracted DNA was then processed on the Check-Direct CPE® assay. The three step process of making the eSwab™, extracting DNA manually and running the Check-Direct CPE® assay, took method of CPE screening; average time-to-diagnosis of 48/72 h. Utilizing this CPE work-flow would allow a 'same-day' result. Antimicrobial susceptibility testing results, as is current practice, would remain a 'next-day' result. In conclusion, the Check-Direct CPE® assay was easily integrated into a local laboratory work-flow and could facilitate a large volume of CPE screening specimens in a single batch, making it cost-effective and convenient for daily CPE testing.

  2. Avaliação de dois métodos de extração de DNA de material parafinado para amplificação em PCR Evaluation of two methods of DNA extraction from paraffin-embedded material for PCR amplification

    Directory of Open Access Journals (Sweden)

    Luciana Estevam Simonato

    2007-04-01

    Full Text Available INTRODUÇÃO: Diversos métodos para extração de DNA a partir de tecidos biológicos inclusos em parafina encontram-se descritos na literatura, sendo o sucesso desse procedimento de grande importância para a realização de métodos moleculares de diagnóstico empregando a reação em cadeia da polimerase (PCR. OBJETIVO: Este estudo avaliou dois métodos de extração de DNA de material parafinado, visando à amplificação do DNA genômico pela técnica da PCR. MATERIAL E MÉTODO: Foram utilizadas 35 amostras de casos de carcinoma epidermóide de assoalho bucal diagnosticados e tratados no Centro de Oncologia Bucal da Universidade Estadual Paulista (Unesp. Os métodos de extração de DNA avaliados incluíram: 1. digestão com proteinase K seguida por purificação com Chelex 100® (BioRad; e 2. sistema QIAamp DNA minikit® (Qiagen. O DNA obtido foi quantificado por espectrofotometria e amplificado pela técnica da PCR, utilizando-se oligonucleotídeos iniciadores para betaglobina. RESULTADOS: A concentração de DNA obtido do material extraído com o primeiro método apresentou média de 120,62 ng/µl com razão entre as leituras das absorbâncias 260/280 variando de 0,8 a 1,41. Para as amostras extraídas com o segundo procedimento, o rendimento médio foi de 67,38 ng/µl, no entanto a razão 260/280 variou entre 1,11 e 2,53. O material foi submetido à PCR e, das 35 amostras extraídas com cada método, respectivamente, 29 e 30 apresentaram sinal positivo. CONCLUSÃO: Os dois métodos utilizados para obtenção de DNA de material parafinado apresentaram desempenho semelhante, revelando que ambos têm potencial para auxiliar na prática da biologia molecular diagnóstica, assim como no estudo diagnóstico retrospectivo em material parafinizado.BACKGROUND: There are several methods for DNA extraction from formalin-fixed paraffin-embedded tissues reported in the literature. High success rate on this procedure is important for the use of

  3. Study of the Prevalence of Causative Bacterial&Protozoal Agents of in Stool Samples of 470 Gastroenteritis Patients Referring to the Nikoopour Clinic in Yazd,Iran

    Directory of Open Access Journals (Sweden)

    MR Sharifi

    2004-04-01

    Full Text Available Interoduction: Gasteroenteritis is one of the problems worth consideration all over the world. It is one of the important causes of mortality, especially in children < 5 years of age, in developing countries including Iran. The aim of this descriptive study was to determine the demographic conditions influencing the presence of causative bacteria and protozoa, followed by antibiograms of isolated bacteria from stool samples of patients with gasteroenteritis referring to Nikoopour Clinic in the city of Yazd, Iran from 1998 – 2001. Materials and method: A total of 470 samples were microbiologically examined by direct method, culture and then antibiogramed. In order to isolate the possible bacteria, differential and selected media were used. Also, wet – mount technique was applied for detection of protozoa. Results: Results revealed that 272 samples (57.9% were infected by pathogenic bacteria or protozoa. 138 (50.8% pathogenic specimens were from male patients and the remaining 134(49.3% were from female patients. Isolated species were: Enteropathogenic E.coli 117(43%, Shigella 51(18.8%, Salmonella.interetidis 25(9.2%, C.jejuni 16(5.9%, Giardia lambdia 51(18.8% and Amoebae spp 12(4.4%. The most commonly detected shigella species was dysenteriae, (74.5% while boydii with 2% was the least common type observed in the specimens. Except shigella, all the other bacteria were more common in males than female, but insignificant statistically. In order to determine the sensitivity and/or resistance of pathogenic bacteria, antibiogram test was performed using selected antibiotic disks such as Ampicillin, Nalidixic Acid, Ciprofloxacin, Gentamycin and Sulfamethaxazole. Conclusion: Results revealed that some patients were probably infected by pathogenic factors other than bacteria or protozoa. Since all viruses and parasites are almost resistant to antibiotics and on the other hand, administration of antibiotics may lead to resistance of bacterial agents

  4. Tocopherol isomer pattern in serum and stool of human following consumption of black currant seed press residue administered in whole grain bread.

    Science.gov (United States)

    Helbig, Dorit; Wagner, Andreas; Schubert, Rainer; Jahreis, Gerhard

    2009-12-01

    Serum gamma-tocopherol is thought to be associated with human health. The dietary influence of tocopherol and fibre-rich black currant seed press residue on serum and stool tocopherol concentration was investigated in a controlled human intervention study. Thirty-six women consumed bread enriched with black currant press residue (4 weeks). The resultant faecal and serum tocopherol concentrations were compared with those after a period consuming control bread without press residue and a normal baseline diet. Fibre intake and excretion, antioxidant capacity (TEAC), and vitamin C concentrations in serum and urine were also determined. Samples were obtained with a 5-day standardised diet at the end of each period. The press residue bread lead to significantly increased beta-, gamma-, delta- and total tocopherol intake, serum alpha-, beta-, gamma- and total tocopherol concentration (with and without lipid adjustment), fibre intake and urinary vitamin C concentration compared to control bread (Pconsumption could be due to a presumed interruption of the enzymatic tocopherol degradation mechanism by bread constituents.

  5. Microscopic diagnosis of sodium acetate-acetic acid-formalin-fixed stool samples for helminths and intestinal protozoa: a comparison among European reference laboratories.

    Science.gov (United States)

    Utzinger, J; Botero-Kleiven, S; Castelli, F; Chiodini, P L; Edwards, H; Köhler, N; Gulletta, M; Lebbad, M; Manser, M; Matthys, B; N'Goran, E K; Tannich, E; Vounatsou, P; Marti, H

    2010-03-01

    The present study aimed to compare the diagnostic performance of different European reference laboratories in diagnosing helminths and intestinal protozoa, using an ether-concentration method applied to sodium acetate-acetic acid-formalin (SAF)-preserved faecal samples. In total, 102 stool specimens were analysed during a cross-sectional parasitological survey in urban farming communities in Côte d'Ivoire. Five SAF-preserved faecal samples were prepared from each specimen and forwarded to the participating reference laboratories, processed and examined under a microscope adhering to a standard operating procedure (SOP). Schistosoma mansoni (cumulative prevalence: 51.0%) and hookworm (cumulative prevalence: 39.2%) were the predominant helminths. There was excellent agreement (kappa > 0.8; p protozoa were Entamoeba coli (median prevalence: 67.6%), Blastocystis hominis (median prevalence: 55.9%) and Entamoeba histolytica/Entamoeba dispar (median prevalence: 47.1%). Substantial agreement among reference laboratories was found for E. coli (kappa = 0.69), but only fair or moderate agreement was found for other Entamoeba species, Giardia intestinalis and Chilomastix mesnili. There was only poor agreement for B. hominis, Isospora belli and Trichomonas intestinalis. In conclusion, although common helminths were reliably diagnosed by European reference laboratories, there was only moderate agreement between centres for pathogenic intestinal protozoa. Continued external quality assessment and the establishment of a formal network of reference laboratories is necessary to further enhance both accuracy and uniformity in parasite diagnosis.

  6. Epidemiological trends and patterns of antimicrobial resistance of Shigella spp. isolated from stool cultures in two different populations in Southern Israel.

    Science.gov (United States)

    Peleg, Itai; Givon-Lavi, Noga; Leibovitz, Eugene; Broides, Arnon

    2014-03-01

    Southern Israel is inhabited by Bedouins, living in conditions similar to developing countries and Jews, living in conditions similar to developed countries. We determined the epidemiology of Shigella spp. in these populations. We retrospectively reviewed Shigella spp. stool isolations between 2005-2009. Overall, 3295 isolates were analyzed. S. sonnei was isolated in 2057/3295 (62.4%) and S. flexneri in 1058 (32.1%). S. sonnei was isolated in 1567/1707 (91.8%) from Jewish patients and S. flexneri in 931/1542 (60.4%) from Bedouin patients. Ampicillin resistance increased linearly from 217/373 (58.2%) in 2005 to 186/256 (72.7%) in 2009, (P Shigella spp. to ampicilin and trimethoprim-sulfamethoxazole were found in Jewish patients: 1527/1706 (89.5%) versus 977/1542 (63.4%) (P Shigella spp. infections can differ in populations residing in the same geographical area. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. Combination therapy with biofeedback, loperamide, and stool-bulking agents is effective for the treatment of fecal incontinence in women - a randomized controlled trial.

    Science.gov (United States)

    Sjödahl, Jenny; Walter, Susanna A; Johansson, Elin; Ingemansson, Anna; Ryn, Ann-Katrine; Hallböök, Olof

    2015-08-01

    Biofeedback and medical treatments have been extensively used for moderate fecal incontinence (FI). There is limited data comparing and combining these two treatments. The objective of this study was to evaluate the effect of biofeedback and medical treatments, separately and in combination. Sixty-four consecutive female patients, referred to a tertiary centre for FI, were included. The patients were randomized to start with either biofeedback (4-6 months) or medical treatment with loperamide and stool-bulking agents (2 months). Both groups continued with a combination of treatments, i.e. medical treatment was added to biofeedback and vice versa. A two-week prospective bowel symptom diary and anorectal physiology were evaluated at baseline, after single- and combination treatments. Fifty-seven patients completed the study. Median number of leakage episodes during two weeks decreased from 6 to 3 (p biofeedback and medical treatment is effective for symptom relief in FI. The symptom improvement was associated with improved fecal consistency, reduced urgency, and increased rectal sensory thresholds.

  8. Sediminibacillus massiliensis sp. nov., a moderately halophilic, Gram-positive bacterium isolated from a stool sample of a young Senegalese man.

    Science.gov (United States)

    Senghor, Bruno; Bassène, Hubert; Khelaifia, Saber; Robert, Catherine; Fournier, Pierre-Edouard; Ruimy, Raymond; Sokhna, Cheikh; Raoult, Didier; Lagier, Jean-Christophe

    2018-07-01

    A Gram-positive, moderately halophilic bacterium, referred to as strain Marseille-P3518 T , was isolated from a stool sample with 2% NaCl concentration from a healthy 15-year-old male living in Dielmo, a village in Senegal. Cells are aerobic, rod-shaped and motile and display endospore formation. Strain Marseille-P3518 T can grow in a medium with 0-20% (w/v) sodium chloride (optimally at 5-7.5% w/v). The major fatty acids were 12-methyl-tetradecanoic acid (45.8%), 13-methyl-tetradecanoic acid (26.9%) and 12-methyl-tridecanoic acid (12.8%). The genome is 4,347,479 bp long with 42.1% G+C content. It contains 4282 protein-coding and 107 RNA genes. Phylogenetic analysis based on 16S rRNA gene sequence comparisons showed that strain Marseille-P3518 T is a member of the Bacillaceae family and is closely related to Sediminibacillus albus (97.4% gene sequence similarity). Strain Marseille-P3518 T was clearly differentiated from its phylogenetic neighbors on the basis of phenotypic and genotypic features. Strain Marseille-P3518 T is, therefore, considered to be a novel representative of the genus Sediminibacillus, for which the name Sediminibacillus massiliensis sp. nov. is proposed, and the type strain is Marseille-P3518 T (CSUR P3518T, DSM69894).

  9. DNA preservation in silk.

    Science.gov (United States)

    Liu, Yawen; Zheng, Zhaozhu; Gong, He; Liu, Meng; Guo, Shaozhe; Li, Gang; Wang, Xiaoqin; Kaplan, David L

    2017-06-27

    The structure of DNA is susceptible to alterations at high temperature and on changing pH, irradiation and exposure to DNase. Options to protect and preserve DNA during storage are important for applications in genetic diagnosis, identity authentication, drug development and bioresearch. In the present study, the stability of total DNA purified from human dermal fibroblast cells, as well as that of plasmid DNA, was studied in silk protein materials. The DNA/silk mixtures were stabilized on filter paper (silk/DNA + filter) or filter paper pre-coated with silk and treated with methanol (silk/DNA + PT-filter) as a route to practical utility. After air-drying and water extraction, 50-70% of the DNA and silk could be retrieved and showed a single band on electrophoretic gels. 6% silk/DNA + PT-filter samples provided improved stability in comparison with 3% silk/DNA + filter samples and DNA + filter samples for DNA preservation, with ∼40% of the band intensity remaining at 37 °C after 40 days and ∼10% after exposure to UV light for 10 hours. Quantitative analysis using the PicoGreen assay confirmed the results. The use of Tris/borate/EDTA (TBE) buffer enhanced the preservation and/or extraction of the DNA. The DNA extracted after storage maintained integrity and function based on serving as a functional template for PCR amplification of the gene for zinc finger protein 750 (ZNF750) and for transgene expression of red fluorescence protein (dsRed) in HEK293 cells. The high molecular weight and high content of a crystalline beta-sheet structure formed on the coated surfaces likely accounted for the preservation effects observed for the silk/DNA + PT-filter samples. Although similar preservation effects were also obtained for lyophilized silk/DNA samples, the rapid and simple processing available with the silk-DNA-filter membrane system makes it appealing for future applications.

  10. Force induced DNA melting

    International Nuclear Information System (INIS)

    Santosh, Mogurampelly; Maiti, Prabal K

    2009-01-01

    When pulled along the axis, double-strand DNA undergoes a large conformational change and elongates by roughly twice its initial contour length at a pulling force of about 70 pN. The transition to this highly overstretched form of DNA is very cooperative. Applying a force perpendicular to the DNA axis (unzipping), double-strand DNA can also be separated into two single-stranded DNA, this being a fundamental process in DNA replication. We study the DNA overstretching and unzipping transition using fully atomistic molecular dynamics (MD) simulations and argue that the conformational changes of double-strand DNA associated with either of the above mentioned processes can be viewed as force induced DNA melting. As the force at one end of the DNA is increased the DNA starts melting abruptly/smoothly above a critical force depending on the pulling direction. The critical force f m , at which DNA melts completely decreases as the temperature of the system is increased. The melting force in the case of unzipping is smaller compared to the melting force when the DNA is pulled along the helical axis. In the case of melting through unzipping, the double-strand separation has jumps which correspond to the different energy minima arising due to sequence of different base pairs. The fraction of Watson-Crick base pair hydrogen bond breaking as a function of force does not show smooth and continuous behavior and consists of plateaus followed by sharp jumps.

  11. DNA damage and autophagy

    International Nuclear Information System (INIS)

    Rodriguez-Rocha, Humberto; Garcia-Garcia, Aracely; Panayiotidis, Mihalis I.; Franco, Rodrigo

    2011-01-01

    Both exogenous and endogenous agents are a threat to DNA integrity. Exogenous environmental agents such as ultraviolet (UV) and ionizing radiation, genotoxic chemicals and endogenous byproducts of metabolism including reactive oxygen species can cause alterations in DNA structure (DNA damage). Unrepaired DNA damage has been linked to a variety of human disorders including cancer and neurodegenerative disease. Thus, efficient mechanisms to detect DNA lesions, signal their presence and promote their repair have been evolved in cells. If DNA is effectively repaired, DNA damage response is inactivated and normal cell functioning resumes. In contrast, when DNA lesions cannot be removed, chronic DNA damage triggers specific cell responses such as cell death and senescence. Recently, DNA damage has been shown to induce autophagy, a cellular catabolic process that maintains a balance between synthesis, degradation, and recycling of cellular components. But the exact mechanisms by which DNA damage triggers autophagy are unclear. More importantly, the role of autophagy in the DNA damage response and cellular fate is unknown. In this review we analyze evidence that supports a role for autophagy as an integral part of the DNA damage response.

  12. DNA Open states and DNA hydratation

    International Nuclear Information System (INIS)

    Lema-Larre, B. de; Martin-Landrove, M

    1995-01-01

    It is a very well-known fact that an protonic exchange exists among natural DNA filaments and synthetic polynucleotides with the solvent (1--2). The existence of DNA open states, that is to say states for which the interior of the DNA molecule is exposed to the external environment, it has been demonstrated by means of proton-deuterium exchange (3). This work has carried out experiments measuring the dispersion of the traverse relaxation rate (4), as a pulsation rate function in a Carr-Purcell-Meiboom-Gill (CPMG) pulses sequence rate, to determine changes in the moist layer of the DNA molecule. The experiments were carried out under different experimental conditions in order to vary the probability that open states occurs, such as temperature or the exposure to electromagnetic fields. Some theoretical models were supposed to adjust the experimental results including those related to DNA non linear dynamic [es

  13. Application of DNA hybridization techniques in the assessment of diarrheal disease among refugess in Thailand. [Shigella; Escherichia coli; Campylobacter; Cryptosporidium

    Energy Technology Data Exchange (ETDEWEB)

    Taylor, D.N.; Echeverria, P.; Pitarangsi, C.; Seriwatana, J.; Sethabutr, O.; Bodhidatta, L.; Brown, C.; Herrmann, J.E.; Blacklow, N.R.

    1988-01-01

    The epidemiology and etiology of acute diarrheal disease were determined in a Hmong refugee camp on the Thai-Laotian border from April 11 to May 14, 1985. DNA hybridization techniques were used to detect Shigella species, enteroinvasive Escherichia coli, and enterotoxigenic E. coli. A monoclonal enzyme-linked immunosorbent assay was used to detect rotavirus, and standard microbiology was used to detect other enteropathogens. The age-specific diarrheal disease rates were 47 episodes per month per 1000 children less than five years old and 113 episodes per month per 1000 children less than one year old. Rotavirus, enterotoxigenic E. coli, Campylobacter, and Cryptosporidium were the predominant pathogens in children less than two years old. The DNA probe hybridized with 94% of 31 specimens identified as enterotoxigenic E. coli by the standard assays and with none of the specimens in which the standard assays were negative. The probe for Shigella and enteroinvasive E. coli hybridized in eight of 10 stools that contained Shigella and four of 314 stools from which Shigella and enteroinvasive E. coli were not isolated. The use of DNA probes allows specimens to be collected in remote areas with a minimum amount of equipment and technical expertise so that they can be easily transported to a central laboratory for further processing.

  14. Immunoassay of DNA damage

    International Nuclear Information System (INIS)

    Gasparro, F.P.; Santella, R.M.

    1988-01-01

    The direct photomodification of DNA by ultraviolet light or the photo-induced addition of exogenous compounds to DNA components results in alterations of DNA structure ranging from subtle to profound. There are two consequences of these conformational changes. First, cells in which the DNA has been damaged are capable of executing repair steps. Second, the DNA which is usually of very low immunogenicity now becomes highly antigenic. This latter property has allowed the production of a series of monoclonal antibodies that recognize photo-induced DNA damage. Monoclonal antibodies have been generated that recognize the 4',5'-monoadduct and the crosslink of 8-methoxypsoralen in DNA. In addition, another antibody has been prepared which recognizes the furan-side monoadduct of 6,4,4'-trimethylangelicin in DNA. These monoclonal antibodies have been characterized as to sensitivity and specificity using non-competitive and competitive enzyme-linked-immunosorbent assays (ELISA). (author)

  15. Immunoassay of DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Gasparro, F P; Santella, R M

    1988-09-01

    The direct photomodification of DNA by ultraviolet light or the photo-induced addition of exogenous compounds to DNA components results in alterations of DNA structure ranging from subtle to profound. There are two consequences of these conformational changes. First, cells in which the DNA has been damaged are capable of executing repair steps. Second, the DNA which is usually of very low immunogenicity now becomes highly antigenic. This latter property has allowed the production of a series of monoclonal antibodies that recognize photo-induced DNA damage. Monoclonal antibodies have been generated that recognize the 4',5'-monoadduct and the crosslink of 8-methoxypsoralen in DNA. In addition, another antibody has been prepared which recognizes the furan-side monoadduct of 6,4,4'-trimethylangelicin in DNA. These monoclonal antibodies have been characterized as to sensitivity and specificity using non-competitive and competitive enzyme-linked-immunosorbent assays (ELISA).

  16. DNA computing models

    CERN Document Server

    Ignatova, Zoya; Zimmermann, Karl-Heinz

    2008-01-01

    In this excellent text, the reader is given a comprehensive introduction to the field of DNA computing. The book emphasizes computational methods to tackle central problems of DNA computing, such as controlling living cells, building patterns, and generating nanomachines.

  17. DNA tagged microparticles

    Science.gov (United States)

    Farquar, George Roy; Leif, Roald N; Wheeler, Elizabeth

    2015-05-05

    A simulant that includes a carrier and DNA encapsulated in the carrier. Also a method of making a simulant including the steps of providing a carrier and encapsulating DNA in the carrier to produce the simulant.

  18. Modeling DNA Replication.

    Science.gov (United States)

    Bennett, Joan

    1998-01-01

    Recommends the use of a model of DNA made out of Velcro to help students visualize the steps of DNA replication. Includes a materials list, construction directions, and details of the demonstration using the model parts. (DDR)

  19. DNA: Structure and function

    DEFF Research Database (Denmark)

    Sinden, Richard R.; E. Pearson, Christopher; N. Potaman, Vladimir

    1998-01-01

    This chapter discusses the structure and function of DNA. DNA occupies a critical role in cells, because it is the source of all intrinsic genetic information. Chemically, DNA is a very stable molecule, a characteristic important for a macromolecule that may have to persist in an intact form...

  20. Replicating animal mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    Emily A. McKinney

    2013-01-01

    Full Text Available The field of mitochondrial DNA (mtDNA replication has been experiencing incredible progress in recent years, and yet little is certain about the mechanism(s used by animal cells to replicate this plasmid-like genome. The long-standing strand-displacement model of mammalian mtDNA replication (for which single-stranded DNA intermediates are a hallmark has been intensively challenged by a new set of data, which suggests that replication proceeds via coupled leading-and lagging-strand synthesis (resembling bacterial genome replication and/or via long stretches of RNA intermediates laid on the mtDNA lagging-strand (the so called RITOLS. The set of proteins required for mtDNA replication is small and includes the catalytic and accessory subunits of DNA polymerase y, the mtDNA helicase Twinkle, the mitochondrial single-stranded DNA-binding protein, and the mitochondrial RNA polymerase (which most likely functions as the mtDNA primase. Mutations in the genes coding for the first three proteins are associated with human diseases and premature aging, justifying the research interest in the genetic, biochemical and structural properties of the mtDNA replication machinery. Here we summarize these properties and discuss the current models of mtDNA replication in animal cells.

  1. Fast phylogenetic DNA barcoding

    DEFF Research Database (Denmark)

    Terkelsen, Kasper Munch; Boomsma, Wouter Krogh; Willerslev, Eske

    2008-01-01

    We present a heuristic approach to the DNA assignment problem based on phylogenetic inferences using constrained neighbour joining and non-parametric bootstrapping. We show that this method performs as well as the more computationally intensive full Bayesian approach in an analysis of 500 insect...... DNA sequences obtained from GenBank. We also analyse a previously published dataset of environmental DNA sequences from soil from New Zealand and Siberia, and use these data to illustrate the fact that statistical approaches to the DNA assignment problem allow for more appropriate criteria...... for determining the taxonomic level at which a particular DNA sequence can be assigned....

  2. Radiation and DNA

    Energy Technology Data Exchange (ETDEWEB)

    Riabchenko, N I

    1979-01-01

    Consideration is given to the effects of ionizing radiation on the structure of DNA. Physical and chemical methods of determining radiation damage to the primary (polynucleotide chain and nitrogenous base) and secondary (helical) structure of DNA are discussed, and the effects of ionizing radiation on deoxyribonucleoprotein complexes are considered. The radiolysis of DNA in vitro and in bacterial and mammalian cells is examined and cellular mechanisms for the repair of radiation-damaged DNA are considered, taking into account single-strand and double-strand breaks, gamma-radiation damage and deoxyribonucleoprotein-membrane complex damage. Postradiation DNA degradation in bacteria and lymphatic cells is also discussed.

  3. DNA-Mediated Electrochemistry

    Science.gov (United States)

    Gorodetsky, Alon A.; Buzzeo, Marisa C.

    2009-01-01

    The base pair stack of DNA has been demonstrated as a medium for long range charge transport chemistry both in solution and at DNA-modified surfaces. This chemistry is exquisitely sensitive to structural perturbations in the base pair stack as occur with lesions, single base mismatches, and protein binding. We have exploited this sensitivity for the development of reliable electrochemical assays based on DNA charge transport at self-assembled DNA monolayers. Here we discuss the characteristic features, applications, and advantages of DNA-mediated electrochemistry. PMID:18980370

  4. Evaluation of multiplex tandem real-time PCR for detection of Cryptosporidium spp., Dientamoeba fragilis, Entamoeba histolytica, and Giardia intestinalis in clinical stool samples.

    Science.gov (United States)

    Stark, D; Al-Qassab, S E; Barratt, J L N; Stanley, K; Roberts, T; Marriott, D; Harkness, J; Ellis, J T

    2011-01-01

    The aim of this study was to describe the first development and evaluation of a multiplex tandem PCR (MT-PCR) assay for the detection and identification of 4 common pathogenic protozoan parasites, Cryptosporidium spp., Dientamoeba fragilis, Entamoeba histolytica, and Giardia intestinalis, from human clinical samples. A total of 472 fecal samples submitted to the Department of Microbiology at St. Vincent's Hospital were included in the study. The MT-PCR assay was compared to four real-time PCR (RT-PCR) assays and microscopy by a traditional modified iron hematoxylin stain. The MT-PCR detected 28 G. intestinalis, 26 D. fragilis, 11 E. histolytica, and 9 Cryptosporidium sp. isolates. Detection and identification of the fecal protozoa by MT-PCR demonstrated 100% correlation with the RT-PCR results, and compared to RT-PCR, MT-PCR exhibited 100% sensitivity and specificity, while traditional microscopy of stained fixed fecal smears exhibited sensitivities and specificities of 56% and 100% for Cryptosporidium spp., 38% and 99% for D. fragilis, 47% and 97% for E. histolytica, and 50% and 100% for G. intestinalis. No cross-reactivity was detected in 100 stool samples containing various other bacterial, viral, and protozoan species. The MT-PCR assay was able to provide rapid, sensitive, and specific simultaneous detection and identification of the four most important diarrhea-causing protozoan parasites that infect humans. This study also highlights the lack of sensitivity demonstrated by microscopy, and thus, molecular methods such as MT-PCR must be considered the diagnostic methods of choice for enteric protozoan parasites.

  5. Método rápido para la observación de Cryptosporidium en heces Rapid method for detection of cryptosporidium in stools

    Directory of Open Access Journals (Sweden)

    Graciela Barona

    1993-02-01

    Full Text Available Entre agosto de 1990 y diciembre de 1991 se examinaron 120 muestras de materia fecal de niños o adultos que consultaron por diarrea, sugestiva de ser causada por Cryptosporidium spp. En todos los casos se realizó la coloración con Lugol-Nigroslna, que proponemos, y se hizo la confirmación con la de Ziehl Neelsen modificada, pese a su limitación de teñir con el mismo patrón de coloración el Cryptosporldium y estructuras diferentes a él. En 20 (16.6% muestras (12 de niños y 8 de adultos se identificaron ooquistes de Cryptosporldlum spp y todas se confirmaron como positivas por la coloración de Ziehl Neelsen modificada. Dado que no siempre es fácil la observación de parásitos de poca prevalencia sugerimos esta coloración como ensayo de rutina porque ayuda a distinguir los ooquistes de Cryptosporidium y mejora la observación de todos los protozoarios.

    We examined 120 stool specimens from patients with diarrheal disease, suspected of being infected with Cryptosporidium. Preliminary observation was made with a Lugol-Nigrosine stain and confirmation with modified Ziehl-Neelsen. Twenty specimens (12 from children and 8 from adults (16.6% were positive for Cryptosporidium oocysts andevery one of them was confirmed with ZN stain. Since It may be difficult to detect low-prevalence parasites we suggest routine use of Lugol-Nigrosine which is useful for the detection of Cryptosporidium as well as of other protozoa.

  6. Providing Quantitative Information and a Nudge to Undergo Stool Testing in a Colorectal Cancer Screening Decision Aid: A Randomized Clinical Trial.

    Science.gov (United States)

    Schwartz, Peter H; Perkins, Susan M; Schmidt, Karen K; Muriello, Paul F; Althouse, Sandra; Rawl, Susan M

    2017-08-01

    Guidelines recommend that patient decision aids should provide quantitative information about probabilities of potential outcomes, but the impact of this information is unknown. Behavioral economics suggests that patients confused by quantitative information could benefit from a "nudge" towards one option. We conducted a pilot randomized trial to estimate the effect sizes of presenting quantitative information and a nudge. Primary care patients (n = 213) eligible for colorectal cancer screening viewed basic screening information and were randomized to view (a) quantitative information (quantitative module), (b) a nudge towards stool testing with the fecal immunochemical test (FIT) (nudge module), (c) neither a nor b, or (d) both a and b. Outcome measures were perceived colorectal cancer risk, screening intent, preferred test, and decision conflict, measured before and after viewing the decision aid, and screening behavior at 6 months. Patients viewing the quantitative module were more likely to be screened than those who did not ( P = 0.012). Patients viewing the nudge module had a greater increase in perceived colorectal cancer risk than those who did not ( P = 0.041). Those viewing the quantitative module had a smaller increase in perceived risk than those who did not ( P = 0.046), and the effect was moderated by numeracy. Among patients with high numeracy who did not view the nudge module, those who viewed the quantitative module had a greater increase in intent to undergo FIT ( P = 0.028) than did those who did not. The limitations of this study were the limited sample size and single healthcare system. Adding quantitative information to a decision aid increased uptake of colorectal cancer screening, while adding a nudge to undergo FIT did not increase uptake. Further research on quantitative information in decision aids is warranted.

  7. Whole genome sequencing analysis of Salmonella enterica serovar Weltevreden isolated from human stool and contaminated food samples collected from the Southern coastal area of China.

    Science.gov (United States)

    Li, Baisheng; Yang, Xingfen; Tan, Hailing; Ke, Bixia; He, Dongmei; Wang, Haiyan; Chen, Qiuxia; Ke, Changwen; Zhang, Yonghui

    2018-02-02

    Salmonella enterica serovar Weltevreden is the most common non-typhoid Salmonella found in South and Southeast Asia. It causes zoonoses worldwide through the consumption of contaminated foods and seafood, and is considered as an important food-borne pathogen in China, especially in the Southern coastal area. We compared the whole genomes of 44 S. Weltevreden strains isolated from human stool and contaminated food samples from Southern Coastal China, in order to investigate their phylogenetic relationships and establish their genetic relatedness to known international strains. ResFinder analysis of the draft genomes of isolated strains detected antimicrobial resistance (AMR) genes in only eight isolates, equivalent to minimum inhibitory concentration assay, and only a few isolates showed resistance to tetracycline, ciprofloxacin or ampicillin. In silico MLST analysis revealed that 43 out of 44 S. Weltevreden strains belonged to sequence type 365 (CC205), the most common sequence type of the serovars. Phylogenetic analysis of the 44 domestic and 26 international isolates suggested that the population of S. Weltevreden could be segregated into six phylogenetic clusters. Cluster I included two strains from food and strains of the "Island Cluster", indicating potential inter-transmission between different countries and regions through foods. The predominant S. Weltevreden isolates obtained from the samples from Southern coastal China were found to be phylogenetically related to strains from Southern East Asia, and formed clusters II-VI. The study has demonstrated that WGS-based analysis may be used to improve our understanding of the epidemiology of this bacterium as part of a food-borne disease surveillance program. The methods used are also more widely applicable to other geographical regions and areas and could therefore be useful for improving our understanding of the international spread of S. Weltevreden on a global scale. Copyright © 2017. Published by Elsevier

  8. DNA fragmentation in spermatozoa

    DEFF Research Database (Denmark)

    Rex, A S; Aagaard, J.; Fedder, J

    2017-01-01

    Sperm DNA Fragmentation has been extensively studied for more than a decade. In the 1940s the uniqueness of the spermatozoa protein complex which stabilizes the DNA was discovered. In the fifties and sixties, the association between unstable chromatin structure and subfertility was investigated....... In the seventies, the impact of induced DNA damage was investigated. In the 1980s the concept of sperm DNA fragmentation as related to infertility was introduced as well as the first DNA fragmentation test: the Sperm Chromatin Structure Assay (SCSA). The terminal deoxynucleotidyl transferase nick end labelling...... (TUNEL) test followed by others was introduced in the nineties. The association between DNA fragmentation in spermatozoa and pregnancy loss has been extensively investigated spurring the need for a therapeutic tool for these patients. This gave rise to an increased interest in the aetiology of DNA damage...

  9. Biophysics of DNA

    CERN Document Server

    Vologodskii, Alexander

    2015-01-01

    Surveying the last sixty years of research, this book describes the physical properties of DNA in the context of its biological functioning. It is designed to enable both students and researchers of molecular biology, biochemistry and physics to better understand the biophysics of DNA, addressing key questions and facilitating further research. The chapters integrate theoretical and experimental approaches, emphasising throughout the importance of a quantitative knowledge of physical properties in building and analysing models of DNA functioning. For example, the book shows how the relationship between DNA mechanical properties and the sequence specificity of DNA-protein binding can be analyzed quantitatively by using our current knowledge of the physical and structural properties of DNA. Theoretical models and experimental methods in the field are critically considered to enable the reader to engage effectively with the current scientific literature on the physical properties of DNA.

  10. Molecular typing and virulence analysis of serotype K1 Klebsiella pneumoniae strains isolated from liver abscess patients and stool samples from noninfectious subjects in Hong Kong, Singapore, and Taiwan.

    Science.gov (United States)

    Siu, L Kristopher; Fung, Chang-Phone; Chang, Feng-Yee; Lee, Nelson; Yeh, Kuo-Ming; Koh, Tse Hsien; Ip, Margaret

    2011-11-01

    Serotype K1 Klebsiella pneumoniae with multilocus sequence type 23 (ST23) has been strongly associated with liver abscess in Taiwan. Few data regarding the strain types and virulence of this serotype from other Asian countries are available. Serotype K1 K. pneumoniae strains isolated from liver abscess and stool samples from subjects hospitalized in Hong Kong, Singapore, and Taiwan hospitals were examined. Forty-seven serotype K1 isolates were identified: 26 from liver abscess samples and 21 from stool samples. MLST revealed 7 sequence types: 85.1% (40 of 47 isolates) belonged to ST23, 1 isolate belonged to ST163 (a single-locus variant of ST23), and 2 isolates were ST249 (a 3-locus variant of ST23). New STs, namely, ST367, ST425, and ST426, were allocated to 3 of 4 isolates from stool samples. The virulence of these strains was determined by neutrophil phagocytosis and mouse infection models. Except for two ST23 isolates, all Klebsiella pneumoniae isolates were resistant to phagocytosis. Resistance to serum killing varied in isolates of ST23, while all non-ST23 strains were susceptible to serum killing except one with ST249 from a liver abscess. All hypervirulent isolates with a 50% lethal dose of serum killing, and also carried both virulence-associated genes, rmpA and aerobactin. Multilocus sequence typing genotype 23 was the most prevalent sequence type among serotype K1 K. pneumoniae isolates from both liver abscess and stool samples in the Asia Pacific region. Serotype K1 K. pneumoniae isolates with capsule expression leading to phagocytic resistance and with the aerobactin gene were associated with hypervirulence.

  11. Comparison of the Triage Micro Parasite Panel and Microscopy for the Detection of Entamoeba histolytica/Entamoeba dispar, Giardia lamblia, and Cryptosporidium parvum in Stool Samples Collected in Kenya

    Directory of Open Access Journals (Sweden)

    Brett Swierczewski

    2012-01-01

    Full Text Available Entamoeba histolytica, Giardia lamblia, and Cryptosporidium parvum are three of the most important parasitic causes of acute diarrhea worldwide. Laboratory diagnosis of these parasites is usually done by ova and parasite examination (O&P examination via microscopy. The sensitivity and specificity of O&P examination varies among laboratories and can be labor intensive and time consuming. The Triage Micro Parasite Panel (BioSite, San Diego, California is an enzyme immunoassay kit that can detect E. histolytica/E. dispar, G. lamblia, and C. parvum simultaneously using fresh or frozen stool. The present study evaluated the Triage Micro Parasite Panel in detecting E. histolytica/E. dispar, G. lamblia, and C. parvum compared to O&P examination in 266 stool samples collected at medical facilities in Kenya. The sensitivity and specificity results for the Triage Micro Parasite Panel were: for E. histolytica/E. dispar: 100%, 100%, G. lamblia: 100%, 100% and C. parvum: 73%, 100%. There was no evidence of cross reactivity using the kit with other parasites identified in the stool specimens. These results indicate that the Triage Micro Parasite Panel is a highly sensitive kit that can be used for screening purposes in large scale studies or outbreak investigations or as a possible alternative to O&P examination.

  12. The effect of changing stool collection processes on compliance in nationwide organized screening using a fecal occult blood test (FOBT) in Korea: study protocol for a randomized controlled trial.

    Science.gov (United States)

    Shin, Hye Young; Suh, Mina; Baik, Hyung Won; Choi, Kui Son; Park, Boyoung; Jun, Jae Kwan; Lee, Chan Wha; Oh, Jae Hwan; Lee, You Kyoung; Han, Dong Soo; Lee, Do-Hoon

    2014-11-26

    Colorectal cancer (CRC) screening by fecal occult blood test (FOBT) significantly reduces CRC mortality, and compliance rates directly influence the efficacy of this screening method. The aim of this study is to investigate whether stool collection strategies affect compliance with the FOBT. In total, 3,596 study participants aged between 50 and 74 years will be recruited. The study will be conducted using a randomized controlled trial, with a 2 × 2 factorial design resulting in four groups. The first factor is the method of stool-collection device distribution (mailing vs. visiting the clinic) and the second is the type of stool-collection device (sampling kit vs. conventional container). Participants will be randomly assigned to one of four groups: (1) sampling kit received by mail; (2) conventional container received by mail; (3) sampling kit received at the clinic; (4) conventional container received at the clinic (control group). The primary outcome will be the FOBT compliance rate; satisfaction and intention to be rescreened in the next screening round will also be evaluated. The rates of positive FOBT results and detection of advanced adenomas or cancers through colonoscopies will also be compared between the two collection containers. Identifying a method of FOBT that yields high compliance rates will be a key determinant of the success of CRC screening. The findings of this study will provide reliable information for health policy makers to develop evidence-based strategies for a high compliance rate. KCT0000803 Date of registration in primary registry: 9 January, 2013.

  13. Using stool antigen to screen for Helicobacter pylori in immigrants and refugees from high prevalence countries is relatively cost effective in reducing the burden of gastric cancer and peptic ulceration.

    Directory of Open Access Journals (Sweden)

    Thomas R Schulz

    Full Text Available OBJECTIVES: Refugees and immigrants from developing countries settling in industrialised countries have a high prevalence of Helicobacter pylori (H. pylori. Screening these groups for H. pylori and use of eradication therapy to reduce the future burden of gastric cancer and peptic ulcer disease is not currently recommended in most countries. We investigated whether a screening and eradication approach would be cost effective in high prevalence populations. METHODS: Nine different screening and follow-up strategies for asymptomatic immigrants from high H. pylori prevalence areas were compared with the current approach of no screening. Cost effectiveness comparisons assumed population prevalence's of H. pylori of 25%, 50% or 75%. The main outcome measure was the net cost for each cancer prevented for each strategy. Total costs of each strategy and net costs including savings from reductions in ulcers and gastric cancer were also calculated. RESULTS: Stool antigen testing with repeat testing after treatment was the most cost effective approach relative to others, for each prevalence value. The net cost per cancer prevented with this strategy was US$111,800 (assuming 75% prevalence, $132,300 (50% and $193,900 (25%. A test and treat strategy using stool antigen remained relatively cost effective, even when the prevalence was 25%. CONCLUSIONS: H. pylori screening and eradication can be an effective strategy for reducing rates of gastric cancer and peptic ulcers in high prevalence populations and our data suggest that use of stool antigen testing is the most cost effective approach.

  14. Helicobacter pylori from Peptic Ulcer Patients in Uganda Is Highly Resistant to Clarithromycin and Fluoroquinolones: Results of the GenoType HelicoDR Test Directly Applied on Stool

    Directory of Open Access Journals (Sweden)

    Denish Calmax Angol

    2017-01-01

    Full Text Available Background. Around 70–90% of peptic ulcer disease (PUD is due to Helicobacter pylori and requires treatment with antimicrobials to which these bacteria are susceptible. Common H. pylori diagnostic tests do not provide drug susceptibility data. Using the GenoType HelicoDR PCR test designed for gastric biopsies for simultaneous detection of H. pylori and its resistance to clarithromycin (CLA/fluoroquinolones (FLQ, we present evidence for stool as an optional test specimen and also provide data on prevalence of H. pylori resistance to CLA and FLQ in Uganda. Methods. Stool from 142 symptomatic PUD patients at three hospitals in Kampala was screened for H. pylori using a rapid antigen test. The GenoType HelicoDR test was run on all H. pylori antigen positives to determine PCR positivity and resistance to CLA/FLQ. Results. Thirty-one samples (22% were H. pylori antigen positive, and 21 (68% of these were H. pylori PCR positive. Six of the 21 (29% were resistant to CLA and eight to FLQ (42%, while two gave invalid FLQ resistance results. Conclusion. Stool is a possible specimen for the GenoType HelicoDR test for rapid detection of H. pylori and drug resistance. In Uganda, Helicobacter pylori is highly resistant to CLA and FLQ.

  15. Beyond DNA repair: DNA-PK function in cancer

    OpenAIRE

    Goodwin, Jonathan F.; Knudsen, Karen E.

    2014-01-01

    The DNA-dependent protein kinase (DNA-PK) is a pivotal component of the DNA repair machinery that governs the response to DNA damage, serving to maintain genome integrity. However, the DNA-PK kinase component was initially isolated with transcriptional complexes, and recent findings have illuminated the impact of DNA-PK-mediated transcriptional regulation on tumor progression and therapeutic response. DNA-PK expression has also been correlated with poor outcome in selected tumor types, furthe...

  16. DNA topology and transcription

    Science.gov (United States)

    Kouzine, Fedor; Levens, David; Baranello, Laura

    2014-01-01

    Chromatin is a complex assembly that compacts DNA inside the nucleus while providing the necessary level of accessibility to regulatory factors conscripted by cellular signaling systems. In this superstructure, DNA is the subject of mechanical forces applied by variety of molecular motors. Rather than being a rigid stick, DNA possesses dynamic structural variability that could be harnessed during critical steps of genome functioning. The strong relationship between DNA structure and key genomic processes necessitates the study of physical constrains acting on the double helix. Here we provide insight into the source, dynamics, and biology of DNA topological domains in the eukaryotic cells and summarize their possible involvement in gene transcription. We emphasize recent studies that might inspire and impact future experiments on the involvement of DNA topology in cellular functions. PMID:24755522

  17. Eukaryotic DNA Replication Fork.

    Science.gov (United States)

    Burgers, Peter M J; Kunkel, Thomas A

    2017-06-20

    This review focuses on the biogenesis and composition of the eukaryotic DNA replication fork, with an emphasis on the enzymes that synthesize DNA and repair discontinuities on the lagging strand of the replication fork. Physical and genetic methodologies aimed at understanding these processes are discussed. The preponderance of evidence supports a model in which DNA polymerase ε (Pol ε) carries out the bulk of leading strand DNA synthesis at an undisturbed replication fork. DNA polymerases α and δ carry out the initiation of Okazaki fragment synthesis and its elongation and maturation, respectively. This review also discusses alternative proposals, including cellular processes during which alternative forks may be utilized, and new biochemical studies with purified proteins that are aimed at reconstituting leading and lagging strand DNA synthesis separately and as an integrated replication fork.

  18. DNA-based machines.

    Science.gov (United States)

    Wang, Fuan; Willner, Bilha; Willner, Itamar

    2014-01-01

    The base sequence in nucleic acids encodes substantial structural and functional information into the biopolymer. This encoded information provides the basis for the tailoring and assembly of DNA machines. A DNA machine is defined as a molecular device that exhibits the following fundamental features. (1) It performs a fuel-driven mechanical process that mimics macroscopic machines. (2) The mechanical process requires an energy input, "fuel." (3) The mechanical operation is accompanied by an energy consumption process that leads to "waste products." (4) The cyclic operation of the DNA devices, involves the use of "fuel" and "anti-fuel" ingredients. A variety of DNA-based machines are described, including the construction of "tweezers," "walkers," "robots," "cranes," "transporters," "springs," "gears," and interlocked cyclic DNA structures acting as reconfigurable catenanes, rotaxanes, and rotors. Different "fuels", such as nucleic acid strands, pH (H⁺/OH⁻), metal ions, and light, are used to trigger the mechanical functions of the DNA devices. The operation of the devices in solution and on surfaces is described, and a variety of optical, electrical, and photoelectrochemical methods to follow the operations of the DNA machines are presented. We further address the possible applications of DNA machines and the future perspectives of molecular DNA devices. These include the application of DNA machines as functional structures for the construction of logic gates and computing, for the programmed organization of metallic nanoparticle structures and the control of plasmonic properties, and for controlling chemical transformations by DNA machines. We further discuss the future applications of DNA machines for intracellular sensing, controlling intracellular metabolic pathways, and the use of the functional nanostructures for drug delivery and medical applications.

  19. DNA repair and cancer

    International Nuclear Information System (INIS)

    Rathore, Shakuntla; Joshi, Pankaj Kumar; Gaur, Sudha

    2012-01-01

    DNA repair refers to a collection of processes by which a cell identifies and corrects damage to the DNA molecule that encode it's genome. In human cells, both normal metabolic activities and environmental factors such as UV light and radiation can cause DNA damage, resulting in as many one million individual molecular lesions per day. Many of these lesions cause structural damage to the DNA molecule and can alter or eliminate the cell's ability to transcribe the gene that the affected DNA encodes. Other lesions include potentially harmful mutation in cell's genome which affect the survival of it's daughter cells after it undergoes mitosis. As a consequence, the DNA repair process is constantly active as it responds to damage in the DNA structure. Inherited mutation that affect DNA repair genes are strongly associated with high cancer risks in humans. Hereditary non polyposis colorectal cancer (HNPCC) is strongly associated with specific mutation in the DNA mismatch repair pathway. BRCA1, BRCA2 two famous mutation conferring a hugely increased risk of breast cancer on carrier, are both associated with a large number of DNA repair pathway, especially NHEJ and homologous recombination. Cancer therapy procedures such as chemotherapy and radiotherapy work by overwhelming the capacity of the cell to repair DNA damage, resulting in cell death. Cells that are most rapidly dividing most typically cancer cells are preferentially affected. The side effect is that other non-cancerous but rapidly dividing cells such as stem cells in the bone marrow are also affected. Modern cancer treatment attempt to localize the DNA damage to cells and tissue only associated with cancer, either by physical means (concentrating the therapeutic agent in the region of the tumor) or by biochemical means (exploiting a feature unique to cancer cells in the body). (author)

  20. Propagação da goiabeira serrana Feijoa sellowiana Berg, através da mergulhia de cepa Vegetative propagation of Feijoa sellowiana Berg through stool layring

    Directory of Open Access Journals (Sweden)

    J.C. Fachinello

    1992-01-01

    Full Text Available Este trabalho teve como objetivo verificar a influência de três tipos de mergulhia de cepa na obtenção de novas plantas de Feijoa sellowiana Berg. Para isso, foi conduzido experimento na EMBRAPA-CNPFT, Pelotas-RS, durante o período de agosto de 90 a julho de 91. Foram utilizados como tratamentos: T1 -corte total da planta a l0cm do nível do solo, seguido de amontoa; T2 - Sem corte da planta, seguido de amontoa a 20 cm de altura e T3 - corte total da planta a 10 cm do nível do solo e amontoa com terra a 20 cm de altura, 2 meses depois do corte. Os parâmetros avaliados foram: número de brotos emitidos por planta; número de brotos enraizados por plantas; número de raízes por broto e vigor dos brotos, representado pelas medidas do comprimento total e diâmetro da base. De acordo com o resultados obtidos o tratamento T3 foi o que proporcionou um maior número de brotos emitidos e enraizados. Para as demais variáveis não houve diferença significativa entre os tratamentos.An experiment to evaluate the effect of stool layring of feijoa was carried out in EMBR APA/CNPFT, Pelotas-RS, from August 90 to July 91. Three treatments of layring propagation were used: T1 - plant cutting 10 cm above the ground and covered with soil; T2 - No plant cutting and covered with soil up to 10 cm above the ground; and T3 - Plant cutting 10 cm above the ground and covered with soil two months later. The results obtained showed that the number of shoots and rooted shoots were more significant in T3. There was no diference between number of roots, diameter and length of shoots among treatments.

  1. Comparative Study of Wheatley’s Trichrome Stain and In-vitro Culture against PCR Assay for the Diagnosis of Blastocystis sp. in Stool Samples

    Directory of Open Access Journals (Sweden)

    Nabilah Amelia MOHAMMAD

    2018-03-01

    Full Text Available Background: This study evaluated the performance of routine permanent stain and cultivation method in comparison with polymerase chain reaction assay as the reference technique to detect Blastocystis sp.Methods: A cross-sectional study was conducted among aboriginal populations that reside in Pahang, Peninsular Malaysia in Feb to Mar 2015. A total of 359 stool samples were examined using Wheatley’s trichrome stain, in-vitro cultivation in Jones’ medium and PCR assay. Positive amplicons were subjected to sequencing and phylogenetic analysis.Results: Fifty-six (15.6% samples were detected positive with Blastocystis sp. by Wheatley’s trichrome stain and 73 (20.3% by in-vitro culture, while PCR assay detected 71 (19.8% positive samples. Detection rate of Blastocystis sp. was highest in combination of microscopic techniques (27.9%. The sensitivity and specificity of Wheatley’s trichrome staining and in-vitro culture techniques compared to PCR assay were 49.3% (95% CI: 37.2-61.4 and 92.7% (95% CI: 89.1-95.4 and 39.4% (95% CI: 28.0-51.8 and 84.4% (95% CI: 79.7-88.4, respectively. However, the sensitivity [60.6% (95% CI: 48.3-71.9] of the method increased when both microscopic techniques were performed together. False negative results produced by microscopic techniques were associated with subtype 3. The agreement between Wheatley’s trichrome stain, in-vitro culture and combination of microscopic techniques with PCR assay were statistically significant by Kappa statistics (Wheatley’s trichrome stain: K = 0.456, P<0.001; in-vitro culture: K = 0.236, P<0.001 and combination techniques: K = 0.353, P<0.001.Conclusion: The combination of microscopic technique is highly recommended to be used as a screening method for the diagnosis of Blastocystis infection either for clinical or epidemiological study to ensure better and accurate diagnosis.

  2. Nonisotopic DNA probe techniques

    National Research Council Canada - National Science Library

    Kricka, Larry J

    1992-01-01

    The objective of this book is to bring together descriptions of the principal nonisotopic methods for DNA hybridization assays, together with experimental details of the methods, including labelling...

  3. DNA replication and cancer

    DEFF Research Database (Denmark)

    Boyer, Anne-Sophie; Walter, David; Sørensen, Claus Storgaard

    2016-01-01

    A dividing cell has to duplicate its DNA precisely once during the cell cycle to preserve genome integrity avoiding the accumulation of genetic aberrations that promote diseases such as cancer. A large number of endogenous impacts can challenge DNA replication and cells harbor a battery of pathways...... causing DNA replication stress and genome instability. Further, we describe cellular and systemic responses to these insults with a focus on DNA replication restart pathways. Finally, we discuss the therapeutic potential of exploiting intrinsic replicative stress in cancer cells for targeted therapy....

  4. Forensic DNA testing.

    Science.gov (United States)

    Butler, John M

    2011-12-01

    Forensic DNA testing has a number of applications, including parentage testing, identifying human remains from natural or man-made disasters or terrorist attacks, and solving crimes. This article provides background information followed by an overview of the process of forensic DNA testing, including sample collection, DNA extraction, PCR amplification, short tandem repeat (STR) allele separation and sizing, typing and profile interpretation, statistical analysis, and quality assurance. The article concludes with discussions of possible problems with the data and other forensic DNA testing techniques.

  5. Structure of DNA toroids and electrostatic attraction of DNA duplexes

    International Nuclear Information System (INIS)

    Cherstvy, A G

    2005-01-01

    DNA-DNA electrostatic attraction is considered as the driving force for the formation of DNA toroids in the presence of DNA condensing cations. This attraction comes from the DNA helical charge distribution and favours hexagonal toroidal cross-sections. The latter is in agreement with recent cryo-electron microscopy studies on DNA condensed with cobalt hexammine. We treat the DNA-DNA interactions within the modern theory of electrostatic interaction between helical macromolecules. The size and thickness of the toroids is calculated within a simple model; other models of stability of DNA toroids are discussed and compared

  6. EFSA NDA Panel (EFSA Panel on Dietetic Products, Nutrition and Allergies), 2014. Scientific Opinion on the substantiation of a health claim related to beta-palmitate and contribution to softening of stools pursuant to Article 14 of Regulation (EC) No 1924/2006

    DEFF Research Database (Denmark)

    Tetens, Inge

    2014-01-01

    Following an application from Specialised Nutrition Europe (formerly IDACE), submitted for authorisation of a health claim pursuant to Article 14 of Regulation (EC) No 1924/2006 via the Competent Authority of France, the EFSA Panel on Dietetic Products, Nutrition and Allergies (NDA) was asked...... to deliver an opinion on the scientific substantiation of a health claim related to beta-palmitate and contribution to softening of stools. The food constituent, beta-palmitate, that is the subject of the health claim, is sufficiently characterised. Contribution to softening of stools is a beneficial...... physiological effect for infants. In weighing the evidence the Panel took into account that, out of two human intervention studies with important methodological limitations, one suggested a stool-softening effect of beta-palmitate whereas the second did not, that one animal study did not support a stool...

  7. Extended DNA Tile Actuators

    DEFF Research Database (Denmark)

    Kristiansen, Martin; Kryger, Mille; Zhang, Zhao

    2012-01-01

    A dynamic linear DNA tile actuator is expanded to three new structures of higher complexity. The original DNA actuator was constructed from a central roller strand which hybridizes with two piston strands by forming two half-crossover junctions. A linear expansion of the actuator is obtained...

  8. Dna fingerprinting - review paper

    OpenAIRE

    Blundell, Renald

    2006-01-01

    Before the Polymerase Chain Reaction (PCR) was established, DNA fingerprinting technology has relied for years on Restriction Fragment Length Polymorphism (RFLP) and Variable Number of Tandom Repeats (VNTR) analysis, a very efficient technique but quite laborious and not suitable for high throughput mapping. Since its, development, PCR has provided a new and powerful tool for DNA fingerprinting.

  9. DNA Repair Systems

    Indian Academy of Sciences (India)

    Thanks to the pioneering research work of Lindahl, Sancar, Modrich and their colleagues, we now have an holistic awareness of how DNA damage occurs and how the damage is rectified in bacteria as well as in higher organisms including human beings. A comprehensive understanding of DNA repair has proven crucial ...

  10. DNA repair genes

    International Nuclear Information System (INIS)

    Morimyo, Mitsuoki

    1995-01-01

    Fission yeast S. pombe is assumed to be a good model for cloning of human DNA repair genes, because human gene is normally expressed in S. pombe and has a very similar protein sequence to yeast protein. We have tried to elucidate the DNA repair mechanisms of S. pombe as a model system for those of mammals. (J.P.N.)

  11. DNA-cell conjugates

    Science.gov (United States)

    Hsiao, Shih-Chia; Francis, Matthew B.; Bertozzi, Carolyn; Mathies, Richard; Chandra, Ravi; Douglas, Erik; Twite, Amy; Toriello, Nicholas; Onoe, Hiroaki

    2018-05-15

    The present invention provides conjugates of DNA and cells by linking the DNA to a native functional group on the cell surface. The cells can be without cell walls or can have cell walls. The modified cells can be linked to a substrate surface and used in assay or bioreactors.

  12. Characterization of muntjac DNA

    International Nuclear Information System (INIS)

    Davis, R.C.

    1981-01-01

    Sister chromatid exchange (SCE) in muntjac chromosomes is generally proportional to the chromosomal DNA content, but the SCE frequency is reduced in the heterochromatic neck region of the X chromosome. The physical properties of muntjac DNA and the kinetics of repair of UV damage in muntjac heterochromatin and euchromatin were examined and compared with the distribution of sister chromatid exchange

  13. DNA-cell conjugates

    Science.gov (United States)

    Hsiao, Shih-Chia; Francis, Matthew B.; Bertozzi, Carolyn; Mathies, Richard; Chandra, Ravi; Douglas, Erik; Twite, Amy; Toriello, Nicholas; Onoe, Hiroaki

    2016-05-03

    The present invention provides conjugates of DNA and cells by linking the DNA to a native functional group on the cell surface. The cells can be without cell walls or can have cell walls. The modified cells can be linked to a substrate surface and used in assay or bioreactors.

  14. Characterization of muntjac DNA

    Energy Technology Data Exchange (ETDEWEB)

    Davis, R.C.

    1981-05-27

    Sister chromatid exchange (SCE) in muntjac chromosomes is generally proportional to the chromosomal DNA content, but the SCE frequency is reduced in the heterochromatic neck region of the X chromosome. The physical properties of muntjac DNA and the kinetics of repair of UV damage in muntjac heterochromatin and euchromatin were examined and compared with the distribution of sister chromatid exchange.

  15. Whose DNA is this?

    DEFF Research Database (Denmark)

    Taroni, Franco; Biedermann, Alex; Vuille, Joëlle

    2013-01-01

    This communication seeks to draw the attention of researchers and practitioners dealing with forensic DNA profiling analyses to the following question: is a scientist's report, offering support to a hypothesis according to which a particular individual is the source of DNA detected during...... evoked during the international conference "The hidden side of DNA profiles. Artifacts, errors and uncertain evidence" held in Rome (April 27th to 28th, 2012). Indeed, despite the fact that this conference brought together some of the world's leading forensic DNA specialists, it appeared clearly...... talk considerably different languages. It thus is fundamental to address this issue of communication about results of forensic DNA analyses, and open a dialogue with practicing non-scientists at large who need to make meaningful use of scientific results to approach and help solve judicial cases...

  16. DNA repair protocols

    DEFF Research Database (Denmark)

    Bjergbæk, Lotte

    In its 3rd edition, this Methods in Molecular Biology(TM) book covers the eukaryotic response to genomic insult including advanced protocols and standard techniques in the field of DNA repair. Offers expert guidance for DNA repair, recombination, and replication. Current knowledge of the mechanisms...... that regulate DNA repair has grown significantly over the past years with technology advances such as RNA interference, advanced proteomics and microscopy as well as high throughput screens. The third edition of DNA Repair Protocols covers various aspects of the eukaryotic response to genomic insult including...... recent advanced protocols as well as standard techniques used in the field of DNA repair. Both mammalian and non-mammalian model organisms are covered in the book, and many of the techniques can be applied with only minor modifications to other systems than the one described. Written in the highly...

  17. Racemic DNA crystallography.

    Science.gov (United States)

    Mandal, Pradeep K; Collie, Gavin W; Kauffmann, Brice; Huc, Ivan

    2014-12-22

    Racemates increase the chances of crystallization by allowing molecular contacts to be formed in a greater number of ways. With the advent of protein synthesis, the production of protein racemates and racemic-protein crystallography are now possible. Curiously, racemic DNA crystallography had not been investigated despite the commercial availability of L- and D-deoxyribo-oligonucleotides. Here, we report a study into racemic DNA crystallography showing the strong propensity of racemic DNA mixtures to form racemic crystals. We describe racemic crystal structures of various DNA sequences and folded conformations, including duplexes, quadruplexes, and a four-way junction, showing that the advantages of racemic crystallography should extend to DNA. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. A comparative analysis of preservation techniques for the optimal molecular detection of hookworm DNA in a human fecal specimen

    Science.gov (United States)

    Pilotte, Nils; Baumer, Ben; Grant, Jessica; Asbjornsdottir, Kristjana; Schaer, Fabian; Hu, Yan; Aroian, Raffi; Walson, Judd; Williams, Steven A.

    2018-01-01

    Background Proper collection and storage of fecal samples is necessary to guarantee the subsequent reliability of DNA-based soil-transmitted helminth diagnostic procedures. Previous research has examined various methods to preserve fecal samples for subsequent microscopic analysis or for subsequent determination of overall DNA yields obtained following DNA extraction. However, only limited research has focused on the preservation of soil-transmitted helminth DNA in stool samples stored at ambient temperature or maintained in a cold chain for extended periods of time. Methodology Quantitative real-time PCR was used in this study as a measure of the effectiveness of seven commercially available products to preserve hookworm DNA over time and at different temperatures. Results were compared against “no preservative” controls and the “gold standard” of rapidly freezing samples at -20°C. The preservation methods were compared at both 4°C and at simulated tropical ambient temperature (32°C) over a period of 60 days. Evaluation of the effectiveness of each preservative was based on quantitative real-time PCR detection of target hookworm DNA. Conclusions At 4°C there were no significant differences in DNA amplification efficiency (as measured by Cq values) regardless of the preservation method utilized over the 60-day period. At 32°C, preservation with FTA cards, potassium dichromate, and a silica bead two-step desiccation process proved most advantageous for minimizing Cq value increases, while RNA later, 95% ethanol and Paxgene also demonstrate some protective effect. These results suggest that fecal samples spiked with known concentrations of hookworm-derived egg material can remain at 4°C for 60 days in the absence of preservative, without significant degradation of the DNA target. Likewise, a variety of preservation methods can provide a measure of protection in the absence of a cold chain. As a result, other factors, such as preservative toxicity

  19. Carriage rates, circulating serotypes and antibiotic resistance ...

    African Journals Online (AJOL)

    volume SDS) and incubated at. 37oC for 1 hour. Genomic DNA (gDNA) was prepared from this lysate using the QiaAmp DNA kit (Qiagen,. UK). Pneumococcal capsular typing was performed on genomic DNA isolated from sub-cultured isolates by.

  20. Simultaneous identification of DNA and RNA viruses present in pig faeces using process-controlled deep sequencing.

    Directory of Open Access Journals (Sweden)

    Jana Sachsenröder

    Full Text Available BACKGROUND: Animal faeces comprise a community of many different microorganisms including bacteria and viruses. Only scarce information is available about the diversity of viruses present in the faeces of pigs. Here we describe a protocol, which was optimized for the purification of the total fraction of viral particles from pig faeces. The genomes of the purified DNA and RNA viruses were simultaneously amplified by PCR and subjected to deep sequencing followed by bioinformatic analyses. The efficiency of the method was monitored using a process control consisting of three bacteriophages (T4, M13 and MS2 with different morphology and genome types. Defined amounts of the bacteriophages were added to the sample and their abundance was assessed by quantitative PCR during the preparation procedure. RESULTS: The procedure was applied to a pooled faecal sample of five pigs. From this sample, 69,613 sequence reads were generated. All of the added bacteriophages were identified by sequence analysis of the reads. In total, 7.7% of the reads showed significant sequence identities with published viral sequences. They mainly originated from bacteriophages (73.9% and mammalian viruses (23.9%; 0.8% of the sequences showed identities to plant viruses. The most abundant detected porcine viruses were kobuvirus, rotavirus C, astrovirus, enterovirus B, sapovirus and picobirnavirus. In addition, sequences with identities to the chimpanzee stool-associated circular ssDNA virus were identified. Whole genome analysis indicates that this virus, tentatively designated as pig stool-associated circular ssDNA virus (PigSCV, represents a novel pig virus. CONCLUSION: The established protocol enables the simultaneous detection of DNA and RNA viruses in pig faeces including the identification of so far unknown viruses. It may be applied in studies investigating aetiology, epidemiology and ecology of diseases. The implemented process control serves as quality control, ensures

  1. Nanostructures via DNA scaffold metallization

    OpenAIRE

    Ning, C.; Zinchenko, A.; Baigl, D.; Pyshkina, O.; Sergeyev, V.; Endo, Kazunaka; Yoshikawa, K.

    2005-01-01

    The critical role of polymers in process of noble metals nanostructures formation is well known, however, the use of DNA chain template in this process is yet largely unknown. In this study we demonstrate different ways of silver deposition on DNA template and report the influence of silver nanostructures formation on DNA conformational state. Metallization of DNA chain proceeds by two different scenarios depending on DNA conformation. If DNA chain is unfolded (elongated) chain, silver reduct...

  2. DNA damage and polyploidization.

    Science.gov (United States)

    Chow, Jeremy; Poon, Randy Y C

    2010-01-01

    A growing body of evidence indicates that polyploidization triggers chromosomal instability and contributes to tumorigenesis. DNA damage is increasingly being recognized for its roles in promoting polyploidization. Although elegant mechanisms known as the DNA damage checkpoints are responsible for halting the cell cycle after DNA damage, agents that uncouple the checkpoints can induce unscheduled entry into mitosis. Likewise, defects of the checkpoints in several disorders permit mitotic entry even in the presence of DNA damage. Forcing cells with damaged DNA into mitosis causes severe chromosome segregation defects, including lagging chromosomes, chromosomal fragments and chromosomal bridges. The presence of these lesions in the cleavage plane is believed to abort cytokinesis. It is postulated that if cytokinesis failure is coupled with defects of the p53-dependent postmitotic checkpoint pathway, cells can enter S phase and become polyploids. Progress in the past several years has unraveled some of the underlying principles of these pathways and underscored the important role of DNA damage in polyploidization. Furthermore, polyploidization per se may also be an important determinant of sensitivity to DNA damage, thereby may offer an opportunity for novel therapies.

  3. Quantitive DNA Fiber Mapping

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Chun-Mei; Wang, Mei; Greulich-Bode, Karin M.; Weier, Jingly F.; Weier, Heinz-Ulli G.

    2008-01-28

    Several hybridization-based methods used to delineate single copy or repeated DNA sequences in larger genomic intervals take advantage of the increased resolution and sensitivity of free chromatin, i.e., chromatin released from interphase cell nuclei. Quantitative DNA fiber mapping (QDFM) differs from the majority of these methods in that it applies FISH to purified, clonal DNA molecules which have been bound with at least one end to a solid substrate. The DNA molecules are then stretched by the action of a receding meniscus at the water-air interface resulting in DNA molecules stretched homogeneously to about 2.3 kb/{micro}m. When non-isotopically, multicolor-labeled probes are hybridized to these stretched DNA fibers, their respective binding sites are visualized in the fluorescence microscope, their relative distance can be measured and converted into kilobase pairs (kb). The QDFM technique has found useful applications ranging from the detection and delineation of deletions or overlap between linked clones to the construction of high-resolution physical maps to studies of stalled DNA replication and transcription.

  4. Regulating DNA Self-assembly by DNA-Surface Interactions.

    Science.gov (United States)

    Liu, Longfei; Li, Yulin; Wang, Yong; Zheng, Jianwei; Mao, Chengde

    2017-12-14

    DNA self-assembly provides a powerful approach for preparation of nanostructures. It is often studied in bulk solution and involves only DNA-DNA interactions. When confined to surfaces, DNA-surface interactions become an additional, important factor to DNA self-assembly. However, the way in which DNA-surface interactions influence DNA self-assembly is not well studied. In this study, we showed that weak DNA-DNA interactions could be stabilized by DNA-surface interactions to allow large DNA nanostructures to form. In addition, the assembly can be conducted isothermally at room temperature in as little as 5 seconds. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Chimeric proteins for detection and quantitation of DNA mutations, DNA sequence variations, DNA damage and DNA mismatches

    Science.gov (United States)

    McCutchen-Maloney, Sandra L.

    2002-01-01

    Chimeric proteins having both DNA mutation binding activity and nuclease activity are synthesized by recombinant technology. The proteins are of the general formula A-L-B and B-L-A where A is a peptide having DNA mutation binding activity, L is a linker and B is a peptide having nuclease activity. The chimeric proteins are useful for detection and identification of DNA sequence variations including DNA mutations (including DNA damage and mismatches) by binding to the DNA mutation and cutting the DNA once the DNA mutation is detected.

  6. "Artifactual" arsenate DNA

    DEFF Research Database (Denmark)

    Nielsen, Peter E

    2012-01-01

    The recent claim by Wolfe-Simon et al. that the Halomonas bacterial strain GFAJ-1 when grown in arsenate-containing medium with limiting phosphate is able to substitute phosphate with arsenate in biomolecules including nucleic acids and in particular DNA(1) arose much skepticism, primarily due...... to the very limited chemical stability of arsenate esters (see ref. 2 and references therein). A major part of the criticisms was concerned with the insufficient (bio)chemical evidence in the Wolfe-Simon study for the actual chemical incorporation of arsenate in DNA (and/or RNA). Redfield et al. now present...... evidence that the identification of arsenate DNA was artifactual....

  7. Multiplex Real-Time PCR Assay Targeting Eight Parasites Customized to the Korean Population: Potential Use for Detection in Diarrheal Stool Samples from Gastroenteritis Patients.

    Directory of Open Access Journals (Sweden)

    Eun Jeong Won

    Full Text Available Intestinal parasitic diseases occur worldwide and can cause diarrhea or gastroenteritis; however, their diagnosis is quite difficult, especially in low-endemism countries. We developed a multiplex real-time PCR assay for detection of eight intestinal parasites and prospectively evaluated it for patients with gastroenteritis. The assay targeted Cryptosporidium parvum, Giardia lamblia, Entamoeba histolytica, Blastocystis hominis, Dientamoeba fragilis, Clonorchis sinensis, Metagonimus yokogawai, and Gymnophalloides seoi. Performance characteristics were evaluated based on recovery after DNA extraction, analytical sensitivity, specificity, reproducibility, cross-reactivity, and interference characteristics. Clinical performance was validated against microscopy on 123 diarrheal samples. The assay demonstrated strong correlations between DNA concentrations and Ct values (R2, 0.9924-0.9998, and had a high PCR efficiency (83.3%-109.5%. Polymerase chain reactions detected as few as 10-30 copies of genomic DNA, and coefficient of variance was 0-7%. There was no cross-reactivity to the other 54 microorganisms tested. Interference occurred only in presence of high concentrations of erythrocytes or leukocytes. This assay had a higher correct identification rate (100.0% vs. 90.2% and lower incorrect ID rate (0.0% vs. 9.8% when compared to microscopy. Overall, this assay showed a higher sensitivity (100.0%; 95% confidence interval [CI] of 80.5-100.0 than microscopy (29.4%; 95% CI 10.31-55.96, and the specificity levels were comparable for both methods (100.0%; 95% CI 96.58-100.0. This newly developed multiplex real-time PCR assay offers a potential use for detecting intestinal parasitic pathogens customized to the Korean population.

  8. DNA from keratinous tissue

    DEFF Research Database (Denmark)

    Bengtsson, Camilla F.; Olsen, Maja E.; Brandt, Luise Ørsted

    2011-01-01

    Keratinous tissues such as nail, hair, horn, scales and feather have been used as a source of DNA for over 20 years. Particular benefits of such tissues include the ease with which they can be sampled, the relative stability of DNA in such tissues once sampled, and, in the context of ancient...... genetic analyses, the fact that sampling generally causes minimal visual damage to valuable specimens. Even when freshly sampled, however, the DNA quantity and quality in the fully keratinized parts of such tissues is extremely poor in comparison to other tissues such as blood and muscle – although little...... systematic research has been undertaken to characterize how such degradation may relate to sample source. In this review paper we present the current understanding of the quality and limitations of DNA in two key keratinous tissues, nail and hair. The findings indicate that although some fragments of nuclear...

  9. DNA fusion gene vaccines

    DEFF Research Database (Denmark)

    Holst, Peter Johannes; Bassi, Maria Rosaria; Thomsen, Allan Randrup

    2010-01-01

    DNA vaccines are versatile and safe, but limited immunogenicity has prevented their use in the clinical setting. Experimentally, immunogenicity may be enhanced by the use of new delivery technologies, by coadministration of cytokines and pathogen-associated molecular patterns, or by fusion...... of antigens into molecular domains that enhance antigen presentation. More specifically, the immunogenicity of DNA vaccines may benefit from increased protein synthesis, increased T-cell help and MHC class I presentation, and the addition of a range of specific cytokines and pathogen-associated molecular...... with viral-vectored vaccines, various synergistic components may need to be incorporated into DNA vaccines. From the perspective of the future clinical use of DNA vaccines, it has been suggested that antigen presentation should be improved and cytokine coadministration attempted. However, even...

  10. DNA Sampling Hook

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The DNA Sampling Hook is a significant improvement on a method of obtaining a tissue sample from a live fish in situ from an aquatic environment. A tissue sample...

  11. Retroviral DNA Integration

    Science.gov (United States)

    2016-01-01

    The integration of a DNA copy of the viral RNA genome into host chromatin is the defining step of retroviral replication. This enzymatic process is catalyzed by the virus-encoded integrase protein, which is conserved among retroviruses and LTR-retrotransposons. Retroviral integration proceeds via two integrase activities: 3′-processing of the viral DNA ends, followed by the strand transfer of the processed ends into host cell chromosomal DNA. Herein we review the molecular mechanism of retroviral DNA integration, with an emphasis on reaction chemistries and architectures of the nucleoprotein complexes involved. We additionally discuss the latest advances on anti-integrase drug development for the treatment of AIDS and the utility of integrating retroviral vectors in gene therapy applications. PMID:27198982

  12. DNA damage and carcinogenesis

    International Nuclear Information System (INIS)

    Stelow, R.B.

    1980-01-01

    Although cancer may arise as a result of many different types of molecular changes, there is little reason to doubt that changes to DNA are one of the more important ones in cancer initiation. Although DNA repair mechanisms seem able to eliminate a very large fraction of deleterious changes to DNA, we not only have little insight into the molecular mechanisms involved in such repair, but have a negligible amount of information to permit us to estimate the shape of dose response relations at low doses. The case of skin cancer is a special one, in that the average population is exposed to sufficient solar uv so that the effects of small increments in uv dose may be estimated. An approximate 85% reduction in DNA repair increases skin cancer incidence 10 4 fold

  13. DNA-Origami

    DEFF Research Database (Denmark)

    Voigt, Niels Vinther; Tørring, Thomas; Gothelf, Kurt Vesterager

    2010-01-01

    DNA-nanostrukturer giver nye muligheder for studier af individuelle molekyler. Ved at udnytte DNAs unikke selvsamlende egenskaber kan man designe systemer, hvorpå der kan studeres kemiske reaktioner, fluoroforer og biiomolekyler på enkeltmolekyle-niveau....

  14. DNA Microarray Technology

    Science.gov (United States)

    Skip to main content DNA Microarray Technology Enter Search Term(s): Español Research Funding An Overview Bioinformatics Current Grants Education and Training Funding Extramural Research News Features Funding Divisions Funding ...

  15. DNA sequencing conference, 2

    Energy Technology Data Exchange (ETDEWEB)

    Cook-Deegan, R.M. [Georgetown Univ., Kennedy Inst. of Ethics, Washington, DC (United States); Venter, J.C. [National Inst. of Neurological Disorders and Strokes, Bethesda, MD (United States); Gilbert, W. [Harvard Univ., Cambridge, MA (United States); Mulligan, J. [Stanford Univ., CA (United States); Mansfield, B.K. [Oak Ridge National Lab., TN (United States)

    1991-06-19

    This conference focused on DNA sequencing, genetic linkage mapping, physical mapping, informatics and bioethics. Several were used to study this sequencing and mapping. This article also discusses computer hardware and software aiding in the mapping of genes.

  16. Close encounters with DNA

    Science.gov (United States)

    Maffeo, C.; Yoo, J.; Comer, J.; Wells, D. B.; Luan, B.; Aksimentiev, A.

    2014-01-01

    Over the past ten years, the all-atom molecular dynamics method has grown in the scale of both systems and processes amenable to it and in its ability to make quantitative predictions about the behavior of experimental systems. The field of computational DNA research is no exception, witnessing a dramatic increase in the size of systems simulated with atomic resolution, the duration of individual simulations and the realism of the simulation outcomes. In this topical review, we describe the hallmark physical properties of DNA from the perspective of all-atom simulations. We demonstrate the amazing ability of such simulations to reveal the microscopic physical origins of experimentally observed phenomena and we review the frustrating limitations associated with imperfections of present atomic force fields and inadequate sampling. The review is focused on the following four physical properties of DNA: effective electric charge, response to an external mechanical force, interaction with other DNA molecules and behavior in an external electric field. PMID:25238560

  17. Gomphid DNA sequence data

    Data.gov (United States)

    U.S. Environmental Protection Agency — DNA sequence data for several genetic loci. This dataset is not publicly accessible because: It's already publicly available on GenBank. It can be accessed through...

  18. HPV DNA test

    Science.gov (United States)

    ... test; Cancer of cervix - HPV DNA test References Hacker NF. Cervical dysplasia and cancer. In: Hacker NF, Gambone JC, Hobel CJ, eds. Hacker and Moore's Essentials of Obstetrics and Gynecology . 6th ...

  19. Close encounters with DNA.

    Science.gov (United States)

    Maffeo, C; Yoo, J; Comer, J; Wells, D B; Luan, B; Aksimentiev, A

    2014-10-15

    Over the past ten years, the all-atom molecular dynamics method has grown in the scale of both systems and processes amenable to it and in its ability to make quantitative predictions about the behavior of experimental systems. The field of computational DNA research is no exception, witnessing a dramatic increase in the size of systems simulated with atomic resolution, the duration of individual simulations and the realism of the simulation outcomes. In this topical review, we describe the hallmark physical properties of DNA from the perspective of all-atom simulations. We demonstrate the amazing ability of such simulations to reveal the microscopic physical origins of experimentally observed phenomena. We also discuss the frustrating limitations associated with imperfections of present atomic force fields and inadequate sampling. The review is focused on the following four physical properties of DNA: effective electric charge, response to an external mechanical force, interaction with other DNA molecules and behavior in an external electric field.

  20. FBI's DNA analysis program

    Science.gov (United States)

    Brown, John R.

    1994-03-01

    Forensic DNA profiling technology is a significant law enforcement tool due to its superior discriminating power. Applying the principles of population genetics to the DNA profile obtained in violent crime investigations results in low frequency of occurrence estimates for the DNA profile. These estimates often range from a frequency of occurrence of 1 in 50 unrelated individuals to 1 in a million unrelated individuals or even smaller. It is this power to discriminate among individuals in the population that has propelled forensic DNA technology to the forefront of forensic testing in violent crime cases. Not only is the technology extremely powerful in including or excluding a criminal suspect as the perpetrator, but it also gives rise to the potential of identifying criminal suspects in cases where the investigators of unknown suspect cases have exhausted all other available leads.

  1. Making DNA Fingerprints.

    Science.gov (United States)

    Nunley, Kathie F.

    1996-01-01

    Presents an activity to simulate electrophoresis using everyday items. Uses adding machine paper to construct a set of DNA fingerprints that can be used to solve crime cases designed by students in any biology class. (JRH)

  2. Radiation damage in DNA

    International Nuclear Information System (INIS)

    Lafleur, V.

    1978-01-01

    A number of experiments are described with the purpose to obtain a better insight in the chemical nature and the biological significance of radiation-induced damage in DNA, with some emphasis on the significance of alkali-labile sites. It is shown that not only reactions of OH radicals but also of H radicals introduce breaks and other inactivating damage in single-standed phiX174 DNA. It is found that phosphate buffer is very suitable for the study of the reactions of H radicals with DNA, as the H 2 PO 4 - ions convert the hydrated electrons into H radicals. The hydrated electron, which does react with DNA, does not cause a detectable inactivation. (Auth.)

  3. DNA to DNA transcription might exist in eukaryotic cells

    OpenAIRE

    Li, Gao-De

    2016-01-01

    Till now, in biological sciences, the term, transcription, mainly refers to DNA to RNA transcription. But our recently published experimental findings obtained from Plasmodium falciparum strongly suggest the existence of DNA to DNA transcription in the genome of eukaryotic cells, which could shed some light on the functions of certain noncoding DNA in the human and other eukaryotic genomes.

  4. Patterning nanocrystals using DNA

    Energy Technology Data Exchange (ETDEWEB)

    Williams, Shara Carol [Univ. of California, Berkeley, CA (United States)

    2003-01-01

    One of the goals of nanotechnology is to enable programmed self-assembly of patterns made of various materials with nanometer-sized control. This dissertation describes the results of experiments templating arrangements of gold and semiconductor nanocrystals using 2'-deoxyribonucleic acid (DNA). Previously, simple DNA-templated linear arrangements of two and three nanocrystals structures have been made.[1] Here, we have sought to assemble larger and more complex nanostructures. Gold-DNA conjugates with 50 to 100 bases self-assembled into planned arrangements using strands of DNA containing complementary base sequences. We used two methods to increase the complexity of the arrangements: using branched synthetic doublers within the DNA covalent backbone to create discrete nanocrystal groupings, and incorporating the nanocrystals into a previously developed DNA lattice structure [2][3] that self-assembles from tiles made of DNA double-crossover molecules to create ordered nanoparticle arrays. In the first project, the introduction of a covalently-branched synthetic doubler reagent into the backbone of DNA strands created a branched DNA ''trimer.'' This DNA trimer templated various structures that contained groupings of three and four gold nanoparticles, giving promising, but inconclusive transmission electron microscopy (TEM) results. Due to the presence of a variety of possible structures in the reaction mixtures, and due to the difficulty of isolating the desired structures, the TEM and gel electrophoresis results for larger structures having four particles, and for structures containing both 5 and 10 nm gold nanoparticles were inconclusive. Better results may come from using optical detection methods, or from improved sample preparation. In the second project, we worked toward making two-dimensional ordered arrays of nanocrystals. We replicated and improved upon previous results for making DNA lattices, increasing the size of the lattices

  5. Das DNA-Puzzle

    Science.gov (United States)

    Kirchner, Stefan

    Im Jahre 1953 wurde von James Watson und Francis Crick erstmalig der strukturelle Aufbau der sogenannten DNA (Desoxyribonukleinsäure) beschrieben, welche das Erbgut jedes Lebewesens enthält. Der wesentliche Teil des Erbguts wird dabei durch eine sehr lange Folge der vier Basen Adenin (A), Cytosin (C), Guanin (G) und Thymin (T) codiert. Seit einigen Jahren ist es möglich, die Folge der vier Basen zu einer gegebenen DNA zu bestimmen. Biologen bezeichnen diesen Vorgang als Sequenzierung.

  6. PDA: Pooled DNA analyzer

    Directory of Open Access Journals (Sweden)

    Lin Chin-Yu

    2006-04-01

    Full Text Available Abstract Background Association mapping using abundant single nucleotide polymorphisms is a powerful tool for identifying disease susceptibility genes for complex traits and exploring possible genetic diversity. Genotyping large numbers of SNPs individually is performed routinely but is cost prohibitive for large-scale genetic studies. DNA pooling is a reliable and cost-saving alternative genotyping method. However, no software has been developed for complete pooled-DNA analyses, including data standardization, allele frequency estimation, and single/multipoint DNA pooling association tests. This motivated the development of the software, 'PDA' (Pooled DNA Analyzer, to analyze pooled DNA data. Results We develop the software, PDA, for the analysis of pooled-DNA data. PDA is originally implemented with the MATLAB® language, but it can also be executed on a Windows system without installing the MATLAB®. PDA provides estimates of the coefficient of preferential amplification and allele frequency. PDA considers an extended single-point association test, which can compare allele frequencies between two DNA pools constructed under different experimental conditions. Moreover, PDA also provides novel chromosome-wide multipoint association tests based on p-value combinations and a sliding-window concept. This new multipoint testing procedure overcomes a computational bottleneck of conventional haplotype-oriented multipoint methods in DNA pooling analyses and can handle data sets having a large pool size and/or large numbers of polymorphic markers. All of the PDA functions are illustrated in the four bona fide examples. Conclusion PDA is simple to operate and does not require that users have a strong statistical background. The software is available at http://www.ibms.sinica.edu.tw/%7Ecsjfann/first%20flow/pda.htm.

  7. Racemic DNA Crystallography

    OpenAIRE

    Mandal , Pradeep K.; Collie , Gavin W.; Kauffmann , Brice; Huc , Ivan

    2014-01-01

    International audience; Racemates increase the chances of crystallization by allowing molecular contacts to be formed in a greater number of ways. With the advent of protein synthesis, the production of protein racemates and racemic-protein crystallography are now possible. Curiously, racemic DNA crystallography had not been investigated despite the commercial availability of Land D-deoxyribo-oligonucleotides. Here, we report a study into racemic DNA crystallography showing the strong propens...

  8. Celebrating DNA's Repair Crew.

    Science.gov (United States)

    Kunkel, Thomas A

    2015-12-03

    This year, the Nobel Prize in Chemistry has been awarded to Tomas Lindahl, Aziz Sancar, and Paul Modrich for their seminal studies of the mechanisms by which cells from bacteria to man repair DNA damage that is generated by normal cellular metabolism and stress from the environment. These studies beautifully illustrate the remarkable power of DNA repair to influence life from evolution through disease susceptibility. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Introduction to DNA methods

    International Nuclear Information System (INIS)

    Delincee, H.

    1991-01-01

    The purpose of this session is to discuss the various possibilities for detecting modifications in DNA after irradiation and whether these changes can be utilized as an indicator for the irradiation treatment of foods. The requirement to be fulfilled is that the method be able to distinguish irradiated food without the presence of a control sample, thus the measured response after irradiation must be large enough to supersede background levels from other treatments. Much work has been performed on the effects of radiation on DNA, particularly due to its importance in radiation biology. The main lesions of DNA as a result of irradiation are base damage, damage of the sugar moiety, single strand and double strand breaks. Crosslinking between bases also occurs, e.g. production of thymine dimers, or between DNA and protein. A valuable review on how to utilize these DNA changes for detection purposes has already appeared. Tables 1, 2 and 3 list the proposed methods of detecting changes in irradiated DNA, some identified products as examples for a possible irradiation indicator, in the case of immunoassay the substance used as antigen, and some selected literature references. In this short review, it is not intended to provide a complete literature survey

  10. Variations in brain DNA

    Directory of Open Access Journals (Sweden)

    Jesus eAvila

    2014-11-01

    Full Text Available It is assumed that DNA sequences are conserved in the diverse cell types present in a multicellular organism like the human being. Thus, in order to compare the sequences in the genome of DNA from different individuals, nucleic acid is commonly isolated from a single tissue. In this regard, blood cells are widely used for this purpose because of their availability. Thus blood DNA has been used to study genetic familiar diseases that affect other tissues and organs, such as the liver, heart, and brain. While this approach is valid for the identification of familial diseases in which mutations are present in parental germinal cells and, therefore, in all the cells of a given organism, it is not suitable to identify sporadic diseases in which mutations might occur in specific somatic cells. This review addresses somatic DNA variations in different tissues or cells (mainly in the brain of single individuals and discusses whether the dogma of DNA invariance between cell types is indeed correct. We will also discuss how single nucleotide somatic variations arise, focusing on the presence of specific DNA mutations in the brain.

  11. Blood extracellular DNA after irradiation

    International Nuclear Information System (INIS)

    Vladimirov, V.G.; Tishchenko, L.I.; Surkova, E.A.; Vasil'eva, I.N.

    1993-01-01

    It has been shown that blood extracellular DNA of irradiated rats largely consists of the low-molecular DNA and its oligomers. Molecular masses of oligomers are multiple to molecular mass of monomer fragment with nucleosome size. The low-molecular DNA has linear form. The average content of GC-pairs in low-molecular DNA is higher than in total rat's DNA (48.5% against 41.5%). The low-molecular DNA is a part of complex containing RNA, acidic proteins and lipids. It is assumed that the formation of low-molecular DNA is a result of Ca/Mg - dependent nuclear endonuclease action

  12. DNA Knots: Theory and Experiments

    Science.gov (United States)

    Sumners, D. W.

    Cellular DNA is a long, thread-like molecule with remarkably complex topology. Enzymes that manipulate the geometry and topology of cellular DNA perform many vital cellular processes (including segregation of daughter chromosomes, gene regulation, DNA repair, and generation of antibody diversity). Some enzymes pass DNA through itself via enzyme-bridged transient breaks in the DNA; other enzymes break the DNA apart and reconnect it to different ends. In the topological approach to enzymology, circular DNA is incubated with an enzyme, producing an enzyme signature in the form of DNA knots and links. By observing the changes in DNA geometry (supercoiling) and topology (knotting and linking) due to enzyme action, the enzyme binding and mechanism can often be characterized. This paper will discuss some personal research history, and the tangle model for the analysis of site-specific recombination experiments on circular DNA.

  13. Bacterial community in ancient permafrost alluvium at the Mammoth Mountain (Eastern Siberia).

    Science.gov (United States)

    Brouchkov, Anatoli; Kabilov, Marsel; Filippova, Svetlana; Baturina, Olga; Rogov, Victor; Galchenko, Valery; Mulyukin, Andrey; Fursova, Oksana; Pogorelko, Gennady

    2017-12-15

    Permanently frozen (approx. 3.5Ma) alluvial Neogene sediments exposed in the Aldan river valley at the Mammoth Mountain (Eastern Siberia) are unique, ancient, and poorly studied permafrost environments. So far, the structure of the indigenous bacterial community has remained unknown. Use of 16S metagenomic analysis with total DNA isolation using DNA Spin Kit for Soil (MO-Bio) and QIAamp DNA Stool Mini Kit (Qiagen) has revealed the major and minor bacterial lineages in the permafrost alluvium sediments. In sum, 61 Operational Taxonomic Units (OTUs) with 31,239 reads (Qiagen kit) and 15,404 reads (Mo-Bio kit) could be assigned to the known taxa. Only three phyla, Bacteroidetes, Proteobacteria and Firmicutes, comprised >5% of the OTUs abundance and accounted for 99% of the total reads. OTUs pertaining to the top families (Chitinophagaceae, Caulobacteraceae, Sphingomonadaceae, Bradyrhizobiaceae, Halomonadaceae) held >90% of reads. The abundance of Actinobacteria was less (0.7%), whereas members of other phyla (Deinococcus-Thermus, Cyanobacteria/Chloroplast, Fusobacteria, and Acidobacteria) constituted a minor fraction of reads. The bacterial community in the studied ancient alluvium differs from other permafrost sediments, mainly by predominance of Bacteroidetes (>52%). The diversity of this preserved bacterial community has the potential to cause effects unknown if prompted to thaw and spread with changing climate. Therefore, this study elicits further reason to study how reintroduction of these ancient bacteria could affect the surrounding ecosystem, including current bacterial species. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Transcription-induced DNA supercoiling: New roles of intranucleosomal DNA loops in DNA repair and transcription.

    Science.gov (United States)

    Gerasimova, N S; Pestov, N A; Kulaeva, O I; Clark, D J; Studitsky, V M

    2016-05-26

    RNA polymerase II (Pol II) transcription through chromatin is accompanied by formation of small intranucleosomal DNA loops. Pol II captured within a small loop drives accumulation of DNA supercoiling, facilitating further transcription. DNA breaks relieve supercoiling and induce Pol II arrest, allowing detection of DNA damage hidden in chromatin structure.

  15. Eukaryotic DNA Replicases

    KAUST Repository

    Zaher, Manal S.; Oke, Muse; Hamdan, Samir

    2014-01-01

    The current model of the eukaryotic DNA replication fork includes three replicative DNA polymerases, polymerase α/primase complex (Pol α), polymerase δ (Pol δ), and polymerase ε (Pol ε). The primase synthesizes 8–12 nucleotide RNA primers that are extended by the DNA polymerization activity of Pol α into 30–35 nucleotide RNA-DNA primers. Replication factor C (RFC) opens the polymerase clamp-like processivity factor, proliferating cell nuclear antigen (PCNA), and loads it onto the primer-template. Pol δ utilizes PCNA to mediate highly processive DNA synthesis, while Pol ε has intrinsic high processivity that is modestly stimulated by PCNA. Pol ε replicates the leading strand and Pol δ replicates the lagging strand in a division of labor that is not strict. The three polymerases are comprised of multiple subunits and share unifying features in their large catalytic and B subunits. The remaining subunits are evolutionarily not related and perform diverse functions. The catalytic subunits are members of family B, which are distinguished by their larger sizes due to inserts in their N- and C-terminal regions. The sizes of these inserts vary among the three polymerases, and their functions remain largely unknown. Strikingly, the quaternary structures of Pol α, Pol δ, and Pol ε are arranged similarly. The catalytic subunits adopt a globular structure that is linked via its conserved C-terminal region to the B subunit. The remaining subunits are linked to the catalytic and B subunits in a highly flexible manner.

  16. Eukaryotic DNA Replicases

    KAUST Repository

    Zaher, Manal S.

    2014-11-21

    The current model of the eukaryotic DNA replication fork includes three replicative DNA polymerases, polymerase α/primase complex (Pol α), polymerase δ (Pol δ), and polymerase ε (Pol ε). The primase synthesizes 8–12 nucleotide RNA primers that are extended by the DNA polymerization activity of Pol α into 30–35 nucleotide RNA-DNA primers. Replication factor C (RFC) opens the polymerase clamp-like processivity factor, proliferating cell nuclear antigen (PCNA), and loads it onto the primer-template. Pol δ utilizes PCNA to mediate highly processive DNA synthesis, while Pol ε has intrinsic high processivity that is modestly stimulated by PCNA. Pol ε replicates the leading strand and Pol δ replicates the lagging strand in a division of labor that is not strict. The three polymerases are comprised of multiple subunits and share unifying features in their large catalytic and B subunits. The remaining subunits are evolutionarily not related and perform diverse functions. The catalytic subunits are members of family B, which are distinguished by their larger sizes due to inserts in their N- and C-terminal regions. The sizes of these inserts vary among the three polymerases, and their functions remain largely unknown. Strikingly, the quaternary structures of Pol α, Pol δ, and Pol ε are arranged similarly. The catalytic subunits adopt a globular structure that is linked via its conserved C-terminal region to the B subunit. The remaining subunits are linked to the catalytic and B subunits in a highly flexible manner.

  17. Long-term frozen storage of urine samples: a trouble to get PCR results in Schistosoma spp. DNA detection?

    Science.gov (United States)

    Fernández-Soto, Pedro; Velasco Tirado, Virginia; Carranza Rodríguez, Cristina; Pérez-Arellano, José Luis; Muro, Antonio

    2013-01-01

    Human schistosomiasis remains a serious worldwide public health problem. At present, a sensitive and specific assay for routine diagnosis of schistosome infection is not yet available. The potential for detecting schistosome-derived DNA by PCR-based methods in human clinical samples is currently being investigated as a diagnostic tool with potential application in routine schistosomiasis diagnosis. Collection of diagnostic samples such as stool or blood is usually difficult in some populations. However, urine is a biological sample that can be collected in a non-invasive method, easy to get from people of all ages and easy in management, but as a sample for PCR diagnosis is still not widely used. This could be due to the high variability in the reported efficiency of detection as a result of the high variation in urine samples' storage or conditions for handling and DNA preservation and extraction methods. We evaluate different commercial DNA extraction methods from a series of long-term frozen storage human urine samples from patients with parasitological confirmed schistosomiasis in order to assess the PCR effectiveness for Schistosoma spp. detection. Patients urine samples were frozen for 18 months up to 7 years until use. Results were compared with those obtained in PCR assays using fresh healthy human urine artificially contaminated with Schistosoma mansoni DNA and urine samples from mice experimentally infected with S. mansoni cercariae stored frozen for at least 12 months before use. PCR results in fresh human artificial urine samples using different DNA based extraction methods were much more effective than those obtained when long-term frozen human urine samples were used as the source of DNA template. Long-term frozen human urine samples are probably not a good source for DNA extraction for use as a template in PCR detection of Schistosoma spp., regardless of the DNA method of extraction used.

  18. Long-term frozen storage of urine samples: a trouble to get PCR results in Schistosoma spp. DNA detection?

    Directory of Open Access Journals (Sweden)

    Pedro Fernández-Soto

    Full Text Available BACKGROUND: Human schistosomiasis remains a serious worldwide public health problem. At present, a sensitive and specific assay for routine diagnosis of schistosome infection is not yet available. The potential for detecting schistosome-derived DNA by PCR-based methods in human clinical samples is currently being investigated as a diagnostic tool with potential application in routine schistosomiasis diagnosis. Collection of diagnostic samples such as stool or blood is usually difficult in some populations. However, urine is a biological sample that can be collected in a non-invasive method, easy to get from people of all ages and easy in management, but as a sample for PCR diagnosis is still not widely used. This could be due to the high variability in the reported efficiency of detection as a result of the high variation in urine samples' storage or conditions for handling and DNA preservation and extraction methods. METHODOLOGY/PRINCIPAL FINDINGS: We evaluate different commercial DNA extraction methods from a series of long-term frozen storage human urine samples from patients with parasitological confirmed schistosomiasis in order to assess the PCR effectiveness for Schistosoma spp. detection. Patients urine samples were frozen for 18 months up to 7 years until use. Results were compared with those obtained in PCR assays using fresh healthy human urine artificially contaminated with Schistosoma mansoni DNA and urine samples from mice experimentally infected with S. mansoni cercariae stored frozen for at least 12 months before use. PCR results in fresh human artificial urine samples using different DNA based extraction methods were much more effective than those obtained when long-term frozen human urine samples were used as the source of DNA template. CONCLUSIONS/SIGNIFICANCE: Long-term frozen human urine samples are probably not a good source for DNA extraction for use as a template in PCR detection of Schistosoma spp., regardless of the DNA

  19. DNA replication stress restricts ribosomal DNA copy number

    Science.gov (United States)

    Salim, Devika; Bradford, William D.; Freeland, Amy; Cady, Gillian; Wang, Jianmin

    2017-01-01

    Ribosomal RNAs (rRNAs) in budding yeast are encoded by ~100–200 repeats of a 9.1kb sequence arranged in tandem on chromosome XII, the ribosomal DNA (rDNA) locus. Copy number of rDNA repeat units in eukaryotic cells is maintained far in excess of the requirement for ribosome biogenesis. Despite the importance of the repeats for both ribosomal and non-ribosomal functions, it is currently not known how “normal” copy number is determined or maintained. To identify essential genes involved in the maintenance of rDNA copy number, we developed a droplet digital PCR based assay to measure rDNA copy number in yeast and used it to screen a yeast conditional temperature-sensitive mutant collection of essential genes. Our screen revealed that low rDNA copy number is associated with compromised DNA replication. Further, subculturing yeast under two separate conditions of DNA replication stress selected for a contraction of the rDNA array independent of the replication fork blocking protein, Fob1. Interestingly, cells with a contracted array grew better than their counterparts with normal copy number under conditions of DNA replication stress. Our data indicate that DNA replication stresses select for a smaller rDNA array. We speculate that this liberates scarce replication factors for use by the rest of the genome, which in turn helps cells complete DNA replication and continue to propagate. Interestingly, tumors from mini chromosome maintenance 2 (MCM2)-deficient mice also show a loss of rDNA repeats. Our data suggest that a reduction in rDNA copy number may indicate a history of DNA replication stress, and that rDNA array size could serve as a diagnostic marker for replication stress. Taken together, these data begin to suggest the selective pressures that combine to yield a “normal” rDNA copy number. PMID:28915237

  20. DNA replication stress restricts ribosomal DNA copy number.

    Science.gov (United States)

    Salim, Devika; Bradford, William D; Freeland, Amy; Cady, Gillian; Wang, Jianmin; Pruitt, Steven C; Gerton, Jennifer L

    2017-09-01

    Ribosomal RNAs (rRNAs) in budding yeast are encoded by ~100-200 repeats of a 9.1kb sequence arranged in tandem on chromosome XII, the ribosomal DNA (rDNA) locus. Copy number of rDNA repeat units in eukaryotic cells is maintained far in excess of the requirement for ribosome biogenesis. Despite the importance of the repeats for both ribosomal and non-ribosomal functions, it is currently not known how "normal" copy number is determined or maintained. To identify essential genes involved in the maintenance of rDNA copy number, we developed a droplet digital PCR based assay to measure rDNA copy number in yeast and used it to screen a yeast conditional temperature-sensitive mutant collection of essential genes. Our screen revealed that low rDNA copy number is associated with compromised DNA replication. Further, subculturing yeast under two separate conditions of DNA replication stress selected for a contraction of the rDNA array independent of the replication fork blocking protein, Fob1. Interestingly, cells with a contracted array grew better than their counterparts with normal copy number under conditions of DNA replication stress. Our data indicate that DNA replication stresses select for a smaller rDNA array. We speculate that this liberates scarce replication factors for use by the rest of the genome, which in turn helps cells complete DNA replication and continue to propagate. Interestingly, tumors from mini chromosome maintenance 2 (MCM2)-deficient mice also show a loss of rDNA repeats. Our data suggest that a reduction in rDNA copy number may indicate a history of DNA replication stress, and that rDNA array size could serve as a diagnostic marker for replication stress. Taken together, these data begin to suggest the selective pressures that combine to yield a "normal" rDNA copy number.

  1. DNA replication stress restricts ribosomal DNA copy number.

    Directory of Open Access Journals (Sweden)

    Devika Salim

    2017-09-01

    Full Text Available Ribosomal RNAs (rRNAs in budding yeast are encoded by ~100-200 repeats of a 9.1kb sequence arranged in tandem on chromosome XII, the ribosomal DNA (rDNA locus. Copy number of rDNA repeat units in eukaryotic cells is maintained far in excess of the requirement for ribosome biogenesis. Despite the importance of the repeats for both ribosomal and non-ribosomal functions, it is currently not known how "normal" copy number is determined or maintained. To identify essential genes involved in the maintenance of rDNA copy number, we developed a droplet digital PCR based assay to measure rDNA copy number in yeast and used it to screen a yeast conditional temperature-sensitive mutant collection of essential genes. Our screen revealed that low rDNA copy number is associated with compromised DNA replication. Further, subculturing yeast under two separate conditions of DNA replication stress selected for a contraction of the rDNA array independent of the replication fork blocking protein, Fob1. Interestingly, cells with a contracted array grew better than their counterparts with normal copy number under conditions of DNA replication stress. Our data indicate that DNA replication stresses select for a smaller rDNA array. We speculate that this liberates scarce replication factors for use by the rest of the genome, which in turn helps cells complete DNA replication and continue to propagate. Interestingly, tumors from mini chromosome maintenance 2 (MCM2-deficient mice also show a loss of rDNA repeats. Our data suggest that a reduction in rDNA copy number may indicate a history of DNA replication stress, and that rDNA array size could serve as a diagnostic marker for replication stress. Taken together, these data begin to suggest the selective pressures that combine to yield a "normal" rDNA copy number.

  2. DNA repair synthesis in human fibroblasts requires DNA polymerase delta

    International Nuclear Information System (INIS)

    Nishida, C.; Reinhard, P.; Linn, S.

    1988-01-01

    When UV-irradiated cultured diploid human fibroblasts were permeabilized with Brij-58 then separated from soluble material by centrifugation, conservative DNA repair synthesis could be restored by a soluble factor obtained from the supernatant of similarly treated HeLa cells. Extensive purification of this factor yielded a 10.2 S, 220,000-dalton polypeptide with the DNA polymerase and 3'- to 5'-exonuclease activities reported for DNA polymerase delta II. Monoclonal antibody to KB cell DNA polymerase alpha, while binding to HeLa DNA polymerase alpha, did not bind to the HeLa DNA polymerase delta. Moreover, at micromolar concentrations N2-(p-n-butylphenyl)-2'-deoxyguanosine 5'-triphosphate (BuPdGTP) and 2-(p-n-butylanilino)-2'-deoxyadenosine 5'-triphosphate (BuAdATP) were potent inhibitors of DNA polymerase alpha, but did not inhibit the DNA polymerase delta. Neither purified DNA polymerase alpha nor beta could promote repair DNA synthesis in the permeabilized cells. Furthermore, under conditions which inhibited purified DNA polymerase alpha by greater than 90%, neither monoclonal antibodies to DNA polymerase alpha, BuPdGTP, nor BuAdATP was able to inhibit significantly the DNA repair synthesis mediated by the DNA polymerase delta. Thus, it appears that a major portion of DNA repair synthesis induced by UV irradiation might be catalyzed by DNA polymerase delta. When xeroderma pigmentosum human diploid fibroblasts were utilized, DNA repair synthesis dependent upon ultraviolet light could be restored by addition of both T4 endonuclease V and DNA polymerase delta, but not by addition of either one alone

  3. Pilot study for early prognosis of Azoospermia in relation to Y-STR ...

    African Journals Online (AJOL)

    Methods: Buccal swab samples were taken from two groups of individuals (50 fertile and 50 Azoospermic patients), then DNA was isolated using QIAamp DNA Micro kit. DNA quantification was done using a Real-time PCR utilizing Quantifiler Kit. PCR was done using PowerPlex Y PCR Amplification Kit, then amplified ...

  4. Principles of DNA architectonics: design of DNA-based nanoobjects

    International Nuclear Information System (INIS)

    Vinogradova, O A; Pyshnyi, D V

    2012-01-01

    The methods of preparation of monomeric DNA blocks that serve as key building units for the construction of complex DNA objects are described. Examples are given of the formation of DNA blocks based on native and modified oligonucleotide components using hydrogen bonding and nucleic acid-specific types of bonding and also some affinity interactions with RNA, proteins, ligands. The static discrete and periodic two- and three-dimensional DNA objects reported to date are described systematically. Methods used to prove the structures of DNA objects and the prospects for practical application of nanostructures based on DNA and its analogues in biology, medicine and biophysics are considered. The bibliography includes 195 references.

  5. DNA Topoisomerases in Transcription

    DEFF Research Database (Denmark)

    Rødgaard, Morten Terpager

    2015-01-01

    This Ph.D. thesis summarizes the main results of my studies on the interplay between DNA topoisomerases and transcription. The work was performed from 2011 to 2015 at Aarhus University in the Laboratory of Genome Research, and was supervised by associate professor Anni H. Andersen. Most of the ex......This Ph.D. thesis summarizes the main results of my studies on the interplay between DNA topoisomerases and transcription. The work was performed from 2011 to 2015 at Aarhus University in the Laboratory of Genome Research, and was supervised by associate professor Anni H. Andersen. Most...... topoisomerase-DNA cleavage complex. The second study is an investigation of how topoisomerases influence gene regulation by keeping the genome in an optimal topological state....

  6. Duplication in DNA Sequences

    Science.gov (United States)

    Ito, Masami; Kari, Lila; Kincaid, Zachary; Seki, Shinnosuke

    The duplication and repeat-deletion operations are the basis of a formal language theoretic model of errors that can occur during DNA replication. During DNA replication, subsequences of a strand of DNA may be copied several times (resulting in duplications) or skipped (resulting in repeat-deletions). As formal language operations, iterated duplication and repeat-deletion of words and languages have been well studied in the literature. However, little is known about single-step duplications and repeat-deletions. In this paper, we investigate several properties of these operations, including closure properties of language families in the Chomsky hierarchy and equations involving these operations. We also make progress toward a characterization of regular languages that are generated by duplicating a regular language.

  7. DNA methylation in obesity

    Directory of Open Access Journals (Sweden)

    Małgorzata Pokrywka

    2014-11-01

    Full Text Available The number of overweight and obese people is increasing at an alarming rate, especially in the developed and developing countries. Obesity is a major risk factor for diabetes, cardiovascular disease, and cancer, and in consequence for premature death. The development of obesity results from the interplay of both genetic and environmental factors, which include sedentary life style and abnormal eating habits. In the past few years a number of events accompanying obesity, affecting expression of genes which are not directly connected with the DNA base sequence (e.g. epigenetic changes, have been described. Epigenetic processes include DNA methylation, histone modifications such as acetylation, methylation, phosphorylation, ubiquitination, and sumoylation, as well as non-coding micro-RNA (miRNA synthesis. In this review, the known changes in the profile of DNA methylation as a factor affecting obesity and its complications are described.

  8. Repeated DNA sequences in fungi

    Energy Technology Data Exchange (ETDEWEB)

    Dutta, S K

    1974-11-01

    Several fungal species, representatives of all broad groups like basidiomycetes, ascomycetes and phycomycetes, were examined for the nature of repeated DNA sequences by DNA:DNA reassociation studies using hydroxyapatite chromatography. All of the fungal species tested contained 10 to 20 percent repeated DNA sequences. There are approximately 100 to 110 copies of repeated DNA sequences of approximately 4 x 10/sup 7/ daltons piece size of each. Repeated DNA sequence homoduplexes showed on average 5/sup 0/C difference of T/sub e/50 (temperature at which 50 percent duplexes dissociate) values from the corresponding homoduplexes of unfractionated whole DNA. It is suggested that a part of repetitive sequences in fungi constitutes mitochondrial DNA and a part of it constitutes nuclear DNA. (auth)

  9. DNA Nanotechnology for Cancer Therapy

    Science.gov (United States)

    Kumar, Vinit; Palazzolo, Stefano; Bayda, Samer; Corona, Giuseppe; Toffoli, Giuseppe; Rizzolio, Flavio

    2016-01-01

    DNA nanotechnology is an emerging and exciting field, and represents a forefront frontier for the biomedical field. The specificity of the interactions between complementary base pairs makes DNA an incredible building material for programmable and very versatile two- and three-dimensional nanostructures called DNA origami. Here, we analyze the DNA origami and DNA-based nanostructures as a drug delivery system. Besides their physical-chemical nature, we dissect the critical factors such as stability, loading capability, release and immunocompatibility, which mainly limit in vivo applications. Special attention was dedicated to highlighting the boundaries to be overcome to bring DNA nanostructures closer to the bedside of patients. PMID:27022418

  10. DNA profiling of trace DNA recovered from bedding.

    Science.gov (United States)

    Petricevic, Susan F; Bright, Jo-Anne; Cockerton, Sarah L

    2006-05-25

    Trace DNA is often detected on handled items and worn clothing examined in forensic laboratories. In this study, the potential transfer of trace DNA to bedding by normal contact, when an individual sleeps in a bed, is examined. Volunteers slept one night on a new, lower bed sheet in their own bed and one night in a bed foreign to them. Samples from the sheets were collected and analysed by DNA profiling. The results indicate that the DNA profile of an individual can be obtained from bedding after one night of sleeping in a bed. The DNA profile of the owner of the bed could also be detected in the foreign bed experiments. Since mixed DNA profiles can be obtained from trace DNA on bedding, caution should be exercised when drawing conclusions from DNA profiling results obtained from such samples. This transfer may have important repercussions in sexual assault investigations.

  11. DNA tagged microparticles

    Science.gov (United States)

    Farquar, George R.; Leif, Roald N.; Wheeler, Elizabeth

    2016-03-22

    In one embodiment, a product includes a plurality of particles, each particle including: a carrier that includes a non-toxic material; and at least one DNA barcode coupled to the carrier, where the particles each have a diameter in a range from about 1 nanometer to about 100 microns.

  12. Comparison of a new multiplex real-time PCR with the Kato Katz thick smear and copro-antigen ELISA for the detection and differentiation of Taenia spp. in human stools.

    Science.gov (United States)

    Ng-Nguyen, Dinh; Stevenson, Mark A; Dorny, Pierre; Gabriël, Sarah; Vo, Tinh Van; Nguyen, Van-Anh Thi; Phan, Trong Van; Hii, Sze Fui; Traub, Rebecca J

    2017-07-01

    Taenia solium, the cause of neurocysticercosis (NCC), has significant socioeconomic impacts on communities in developing countries. This disease, along with taeniasis is estimated to infect 2.5 to 5 million people globally. Control of T. solium NCC necessitates accurate diagnosis and treatment of T. solium taeniasis carriers. In areas where all three species of Taenia tapeworms (T. solium, Taenia saginata and Taenia asiatica) occur sympatrically, conventional microscope- and copro-antigen based diagnostic methods are unable to distinguish between these three Taenia species. Molecular diagnostic tools have been developed to overcome this limitation; however, conventional PCR-based techniques remain unsuitable for large-scale deployment in community-based surveys. Moreover, a real-time PCR (qPCR) for the discrimination of all three species of Taenia in human stool does not exist. This study describes the development and validation of a new triplex Taq-Man probe-based qPCR for the detection and discrimination of all three Taenia human tapeworms in human stools collected from communities in the Central Highlands of Vietnam. The diagnostic characteristics of the test are compared with conventional Kato Katz (KK) thick smear and copro-antigen ELISA (cAgELISA) method utilizing fecal samples from a community based cross-sectional study. Using this new multiplex real-time PCR we provide an estimate of the true prevalence of taeniasis in the source population for the community based cross-sectional study. Primers and TaqMan probes for the specific amplification of T. solium, T. saginata and T. asiatica were designed and successfully optimized to target the internal transcribed spacer I (ITS-1) gene of T. solium and the cytochrome oxidase subunit I (COX-1) gene of T. saginata and T. asiatica. The newly designed triplex qPCR (T3qPCR) was compared to KK and cAgELISA for the detection of Taenia eggs in stool samples collected from 342 individuals in Dak Lak province, Central

  13. Comparison of a new multiplex real-time PCR with the Kato Katz thick smear and copro-antigen ELISA for the detection and differentiation of Taenia spp. in human stools.

    Directory of Open Access Journals (Sweden)

    Dinh Ng-Nguyen

    2017-07-01

    Full Text Available Taenia solium, the cause of neurocysticercosis (NCC, has significant socioeconomic impacts on communities in developing countries. This disease, along with taeniasis is estimated to infect 2.5 to 5 million people globally. Control of T. solium NCC necessitates accurate diagnosis and treatment of T. solium taeniasis carriers. In areas where all three species of Taenia tapeworms (T. solium, Taenia saginata and Taenia asiatica occur sympatrically, conventional microscope- and copro-antigen based diagnostic methods are unable to distinguish between these three Taenia species. Molecular diagnostic tools have been developed to overcome this limitation; however, conventional PCR-based techniques remain unsuitable for large-scale deployment in community-based surveys. Moreover, a real-time PCR (qPCR for the discrimination of all three species of Taenia in human stool does not exist. This study describes the development and validation of a new triplex Taq-Man probe-based qPCR for the detection and discrimination of all three Taenia human tapeworms in human stools collected from communities in the Central Highlands of Vietnam. The diagnostic characteristics of the test are compared with conventional Kato Katz (KK thick smear and copro-antigen ELISA (cAgELISA method utilizing fecal samples from a community based cross-sectional study. Using this new multiplex real-time PCR we provide an estimate of the true prevalence of taeniasis in the source population for the community based cross-sectional study.Primers and TaqMan probes for the specific amplification of T. solium, T. saginata and T. asiatica were designed and successfully optimized to target the internal transcribed spacer I (ITS-1 gene of T. solium and the cytochrome oxidase subunit I (COX-1 gene of T. saginata and T. asiatica. The newly designed triplex qPCR (T3qPCR was compared to KK and cAgELISA for the detection of Taenia eggs in stool samples collected from 342 individuals in Dak Lak province

  14. Comparison of a new multiplex real-time PCR with the Kato Katz thick smear and copro-antigen ELISA for the detection and differentiation of Taenia spp. in human stools

    Science.gov (United States)

    Stevenson, Mark A.; Dorny, Pierre; Gabriël, Sarah; Vo, Tinh Van; Nguyen, Van-Anh Thi; Phan, Trong Van; Hii, Sze Fui; Traub, Rebecca J.

    2017-01-01

    Background Taenia solium, the cause of neurocysticercosis (NCC), has significant socioeconomic impacts on communities in developing countries. This disease, along with taeniasis is estimated to infect 2.5 to 5 million people globally. Control of T. solium NCC necessitates accurate diagnosis and treatment of T. solium taeniasis carriers. In areas where all three species of Taenia tapeworms (T. solium, Taenia saginata and Taenia asiatica) occur sympatrically, conventional microscope- and copro-antigen based diagnostic methods are unable to distinguish between these three Taenia species. Molecular diagnostic tools have been developed to overcome this limitation; however, conventional PCR-based techniques remain unsuitable for large-scale deployment in community-based surveys. Moreover, a real-time PCR (qPCR) for the discrimination of all three species of Taenia in human stool does not exist. This study describes the development and validation of a new triplex Taq-Man probe-based qPCR for the detection and discrimination of all three Taenia human tapeworms in human stools collected from communities in the Central Highlands of Vietnam. The diagnostic characteristics of the test are compared with conventional Kato Katz (KK) thick smear and copro-antigen ELISA (cAgELISA) method utilizing fecal samples from a community based cross-sectional study. Using this new multiplex real-time PCR we provide an estimate of the true prevalence of taeniasis in the source population for the community based cross-sectional study. Methodology/Principal findings Primers and TaqMan probes for the specific amplification of T. solium, T. saginata and T. asiatica were designed and successfully optimized to target the internal transcribed spacer I (ITS-1) gene of T. solium and the cytochrome oxidase subunit I (COX-1) gene of T. saginata and T. asiatica. The newly designed triplex qPCR (T3qPCR) was compared to KK and cAgELISA for the detection of Taenia eggs in stool samples collected from 342

  15. The dynamic interplay between DNA topoisomerases and DNA topology.

    Science.gov (United States)

    Seol, Yeonee; Neuman, Keir C

    2016-11-01

    Topological properties of DNA influence its structure and biochemical interactions. Within the cell, DNA topology is constantly in flux. Transcription and other essential processes, including DNA replication and repair, not only alter the topology of the genome but also introduce additional complications associated with DNA knotting and catenation. These topological perturbations are counteracted by the action of topoisomerases, a specialized class of highly conserved and essential enzymes that actively regulate the topological state of the genome. This dynamic interplay among DNA topology, DNA processing enzymes, and DNA topoisomerases is a pervasive factor that influences DNA metabolism in vivo. Building on the extensive structural and biochemical characterization over the past four decades that has established the fundamental mechanistic basis of topoisomerase activity, scientists have begun to explore the unique roles played by DNA topology in modulating and influencing the activity of topoisomerases. In this review we survey established and emerging DNA topology-dependent protein-DNA interactions with a focus on in vitro measurements of the dynamic interplay between DNA topology and topoisomerase activity.

  16. Conformation-dependent DNA attraction

    Science.gov (United States)

    Li, Weifeng; Nordenskiöld, Lars; Zhou, Ruhong; Mu, Yuguang

    2014-05-01

    Understanding how DNA molecules interact with other biomolecules is related to how they utilize their functions and is therefore critical for understanding their structure-function relationships. For a long time, the existence of Z-form DNA (a left-handed double helical version of DNA, instead of the common right-handed B-form) has puzzled the scientists, and the definitive biological significance of Z-DNA has not yet been clarified. In this study, the effects of DNA conformation in DNA-DNA interactions are explored by molecular dynamics simulations. Using umbrella sampling, we find that for both B- and Z-form DNA, surrounding Mg2+ ions always exert themselves to screen the Coulomb repulsion between DNA phosphates, resulting in very weak attractive force. On the contrary, a tight and stable bound state is discovered for Z-DNA in the presence of Mg2+ or Na+, benefiting from their hydrophobic nature. Based on the contact surface and a dewetting process analysis, a two-stage binding process of Z-DNA is outlined: two Z-DNA first attract each other through charge screening and Mg2+ bridges to phosphate groups in the same way as that of B-DNA, after which hydrophobic contacts of the deoxyribose groups are formed via a dewetting effect, resulting in stable attraction between two Z-DNA molecules. The highlighted hydrophobic nature of Z-DNA interaction from the current study may help to understand the biological functions of Z-DNA in gene transcription.Understanding how DNA molecules interact with other biomolecules is related to how they utilize their functions and is therefore critical for understanding their structure-function relationships. For a long time, the existence of Z-form DNA (a left-handed double helical version of DNA, instead of the common right-handed B-form) has puzzled the scientists, and the definitive biological significance of Z-DNA has not yet been clarified. In this study, the effects of DNA conformation in DNA-DNA interactions are explored by

  17. Melting curves of gammairradiated DNA

    International Nuclear Information System (INIS)

    Hofer, H.; Altmann, H.; Kehrer, M.

    1978-08-01

    Melting curves of gammairradiated DNA and data derived of them, are reported. The diminished stability is explained by basedestruction. DNA denatures completely at room temperature, if at least every fifth basepair is broken or weakened by irradiation. (author)

  18. An Introduction to DNA Fingerprinting.

    Science.gov (United States)

    Hepfer, Carol Ely; And Others

    1993-01-01

    Provides background information on DNA fingerprinting, and describes exercises for introducing general biology students at the high school or college level to the methodology and applications of DNA fingerprinting. (PR)

  19. Esitleti kakskeelset luulekogu "Luule DNA"

    Index Scriptorium Estoniae

    2007-01-01

    Magrelli, Valerio. Luule DNA = Il DNA della poesia / tõlkinud [ja saatesõna:] Maarja Kangro ja Kalju Kruusa. Tallinn : Koma, 2006. Sisaldab autori teksti. Esitlus 24. jaan. Kirjanike majas Tallinnas

  20. Conformation-dependent DNA attraction.

    Science.gov (United States)

    Li, Weifeng; Nordenskiöld, Lars; Zhou, Ruhong; Mu, Yuguang

    2014-06-21

    Understanding how DNA molecules interact with other biomolecules is related to how they utilize their functions and is therefore critical for understanding their structure-function relationships. For a long time, the existence of Z-form DNA (a left-handed double helical version of DNA, instead of the common right-handed B-form) has puzzled the scientists, and the definitive biological significance of Z-DNA has not yet been clarified. In this study, the effects of DNA conformation in DNA-DNA interactions are explored by molecular dynamics simulations. Using umbrella sampling, we find that for both B- and Z-form DNA, surrounding Mg(2+) ions always exert themselves to screen the Coulomb repulsion between DNA phosphates, resulting in very weak attractive force. On the contrary, a tight and stable bound state is discovered for Z-DNA in the presence of Mg(2+) or Na(+), benefiting from their hydrophobic nature. Based on the contact surface and a dewetting process analysis, a two-stage binding process of Z-DNA is outlined: two Z-DNA first attract each other through charge screening and Mg(2+) bridges to phosphate groups in the same way as that of B-DNA, after which hydrophobic contacts of the deoxyribose groups are formed via a dewetting effect, resulting in stable attraction between two Z-DNA molecules. The highlighted hydrophobic nature of Z-DNA interaction from the current study may help to understand the biological functions of Z-DNA in gene transcription.

  1. Alterations of ultraviolet irradiated DNA

    International Nuclear Information System (INIS)

    Davila, C.; Garces, F.

    1980-01-01

    Thymine dimers production has been studied in several DNA- 3 H irradiated at various wave lenght of U.V. Light. The influence of dimers on the hydrodynamic and optic properties, thermal structural stability and transformant capacity of DNA have been studied too. At last the recognition and excision of dimers by the DNA-UV-Endonuclease and DNA-Polimerase-I was also studied. (author)

  2. Interfacing DNA nanodevices with biology

    DEFF Research Database (Denmark)

    Vinther, Mathias; Kjems, Jørgen

    2016-01-01

    in biology and biomedicine acting as a molecular ‘nanorobot’ or smart drug interacting with the cellular machinery. In this review, we will explore and examine the perspective of DNA nanotechnology for such use. We summarize which requirements DNA nanostructures must fulfil to function in cellular...... environments and inside living organisms. In addition, we highlight recent advances in interfacing DNA nanostructures with biology....

  3. Human Genome Research: Decoding DNA

    Science.gov (United States)

    dropdown arrow Site Map A-Z Index Menu Synopsis Human Genome Research: Decoding DNA Resources with of the DNA double helix during April 2003. James D. Watson, Francis Crick, and Maurice Wilkins were company Celera announced the completion of a "working draft" reference DNA sequence of the human

  4. My journey to DNA repair.

    Science.gov (United States)

    Lindahl, Tomas

    2013-02-01

    I completed my medical studies at the Karolinska Institute in Stockholm but have always been devoted to basic research. My longstanding interest is to understand fundamental DNA repair mechanisms in the fields of cancer therapy, inherited human genetic disorders and ancient DNA. I initially measured DNA decay, including rates of base loss and cytosine deamination. I have discovered several important DNA repair proteins and determined their mechanisms of action. The discovery of uracil-DNA glycosylase defined a new category of repair enzymes with each specialized for different types of DNA damage. The base excision repair pathway was first reconstituted with human proteins in my group. Cell-free analysis for mammalian nucleotide excision repair of DNA was also developed in my laboratory. I found multiple distinct DNA ligases in mammalian cells, and led the first genetic and biochemical work on DNA ligases I, III and IV. I discovered the mammalian exonucleases DNase III (TREX1) and IV (FEN1). Interestingly, expression of TREX1 was altered in some human autoimmune diseases. I also showed that the mutagenic DNA adduct O(6)-methylguanine (O(6)mG) is repaired without removing the guanine from DNA, identifying a surprising mechanism by which the methyl group is transferred to a residue in the repair protein itself. A further novel process of DNA repair discovered by my research group is the action of AlkB as an iron-dependent enzyme carrying out oxidative demethylation. Copyright © 2013. Production and hosting by Elsevier Ltd.

  5. DNA Origami-Graphene Hybrid Nanopore for DNA Detection.

    Science.gov (United States)

    Barati Farimani, Amir; Dibaeinia, Payam; Aluru, Narayana R

    2017-01-11

    DNA origami nanostructures can be used to functionalize solid-state nanopores for single molecule studies. In this study, we characterized a nanopore in a DNA origami-graphene heterostructure for DNA detection. The DNA origami nanopore is functionalized with a specific nucleotide type at the edge of the pore. Using extensive molecular dynamics (MD) simulations, we computed and analyzed the ionic conductivity of nanopores in heterostructures carpeted with one or two layers of DNA origami on graphene. We demonstrate that a nanopore in DNA origami-graphene gives rise to distinguishable dwell times for the four DNA base types, whereas for a nanopore in bare graphene, the dwell time is almost the same for all types of bases. The specific interactions (hydrogen bonds) between DNA origami and the translocating DNA strand yield different residence times and ionic currents. We also conclude that the speed of DNA translocation decreases due to the friction between the dangling bases at the pore mouth and the sequencing DNA strands.

  6. Efficient Sleeping Beauty DNA Transposition From DNA Minicircles

    Directory of Open Access Journals (Sweden)

    Nynne Sharma

    2013-01-01

    Full Text Available DNA transposon-based vectors have emerged as new potential delivery tools in therapeutic gene transfer. Such vectors are now showing promise in hematopoietic stem cells and primary human T cells, and clinical trials with transposon-engineered cells are on the way. However, the use of plasmid DNA as a carrier of the vector raises safety concerns due to the undesirable administration of bacterial sequences. To optimize vectors based on the Sleeping Beauty (SB DNA transposon for clinical use, we examine here SB transposition from DNA minicircles (MCs devoid of the bacterial plasmid backbone. Potent DNA transposition, directed by the hyperactive SB100X transposase, is demonstrated from MC donors, and the stable transfection rate is significantly enhanced by expressing the SB100X transposase from MCs. The stable transfection rate is inversely related to the size of circular donor, suggesting that a MC-based SB transposition system benefits primarily from an increased cellular uptake and/or enhanced expression which can be observed with DNA MCs. DNA transposon and transposase MCs are easily produced, are favorable in size, do not carry irrelevant DNA, and are robust substrates for DNA transposition. In accordance, DNA MCs should become a standard source of DNA transposons not only in therapeutic settings but also in the daily use of the SB system.

  7. DNA-DNA hybridization determined in micro-wells using covalent attachment of DNA

    DEFF Research Database (Denmark)

    Christensen, H.; Angen, Øystein; Mutters, R.

    2000-01-01

    The present study was aimed at reducing the time and labour used to perform DNA-DNA hybridizations for classification of bacteria at the species level. A micro-well-format DNA hybridization method was developed and validated. DNA extractions were performed by a small-scale method and DNA...... was sheared mechanically into fragments of between 400 and 700 bases. The hybridization conditions were calibrated according to DNA similarities obtained by the spectrophotometric method using strains within the family Pasteurellaceae, Optimal conditions were obtained with 300 ng DNA added per well and bound...... by covalent attachment to NucleoLink. Hybridization was performed with 500 ng DNA, 5% (w/w) of which was labelled with photo-activatable biotin (competitive hybridization) for 2.5 h at 65 degrees C in 2 x SSC followed by stringent washing with 2 x SSC at the same temperature. The criteria for acceptance...

  8. Dine marker har DNA

    DEFF Research Database (Denmark)

    Eckholdt, Annette; Winding, Anne; Krogh, Paul Henning

    2017-01-01

    Ordet "biodiversitet" og at det er noget, vi skal have mere af, nævnes hyppigt. Men hvad er biodiversitet, og hvordan måles det? Agrologisk har bedt et par eksperter fra Aarhus Universitet forklare, hvordan et DNA-aftryk af jord og vand kan erstatte optællinger i felten og sige noget om biodivers......Ordet "biodiversitet" og at det er noget, vi skal have mere af, nævnes hyppigt. Men hvad er biodiversitet, og hvordan måles det? Agrologisk har bedt et par eksperter fra Aarhus Universitet forklare, hvordan et DNA-aftryk af jord og vand kan erstatte optællinger i felten og sige noget om...

  9. Fleet DNA (Presentation)

    Energy Technology Data Exchange (ETDEWEB)

    Walkokwicz, K.; Duran, A.

    2014-06-01

    The Fleet DNA project objectives include capturing and quantifying drive cycle and technology variation for the multitude of medium- and heavy-duty vocations; providing a common data storage warehouse for medium- and heavy-duty vehicle fleet data across DOE activities and laboratories; and integrating existing DOE tools, models, and analyses to provide data-driven decision making capabilities. Fleet DNA advantages include: for Government - providing in-use data for standard drive cycle development, R&D, tech targets, and rule making; for OEMs - real-world usage datasets provide concrete examples of customer use profiles; for fleets - vocational datasets help illustrate how to maximize return on technology investments; for Funding Agencies - ways are revealed to optimize the impact of financial incentive offers; and for researchers -a data source is provided for modeling and simulation.

  10. Repulsive DNA-DNA interactions accelerate viral DNA packaging in phage Phi29.

    Science.gov (United States)

    Keller, Nicholas; delToro, Damian; Grimes, Shelley; Jardine, Paul J; Smith, Douglas E

    2014-06-20

    We use optical tweezers to study the effect of attractive versus repulsive DNA-DNA interactions on motor-driven viral packaging. Screening of repulsive interactions accelerates packaging, but induction of attractive interactions by spermidine(3+) causes heterogeneous dynamics. Acceleration is observed in a fraction of complexes, but most exhibit slowing and stalling, suggesting that attractive interactions promote nonequilibrium DNA conformations that impede the motor. Thus, repulsive interactions facilitate packaging despite increasing the energy of the theoretical optimum spooled DNA conformation.

  11. Radiobiology with DNA ligands

    International Nuclear Information System (INIS)

    Weinreich, R.; Argentini, M.; Guenther, I.; Koziorowski, J.; Larsson, B.; Nievergelt-Egido, M.C.; Salt, C.; Wyer, L.; Dos Santos, D.F.; Hansen, H.J.

    1997-01-01

    The paper deals with the following topics: labelling of DNA ligands and other tumour-affinic compounds with 4.15-d 124 I, radiotoxicity of Hoechst 33258 and 33342 and of iodinated Hoechst 33258 in cell cultures, preparation of 76 Br-, 123 I-, and 221 At-labelled 5-halo-2'-deoxyuridine, chemical syntheses of boron derivatives of Hoechst 33258.III., Gadolinium neutron capture therapy

  12. DNA nanotechnology and fluorescence applications.

    Science.gov (United States)

    Schlichthaerle, Thomas; Strauss, Maximilian T; Schueder, Florian; Woehrstein, Johannes B; Jungmann, Ralf

    2016-06-01

    Structural DNA nanotechnology allow researchers to use the unique molecular recognition properties of DNA strands to construct nanoscale objects with almost arbitrary complexity in two and three dimensions. Abstracted as molecular breadboards, DNA nanostructures enable nanometer-precise placement of guest molecules such as proteins, fluorophores, or nanoparticles. These assemblies can be used to study biological phenomena with unprecedented control over number, spacing, and molecular identity. Here, we give a general introduction to structural DNA nanotechnology and more specifically discuss applications of DNA nanostructures in the field of fluorescence and plasmonics. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Next generation DNA led technologies

    CERN Document Server

    Jyothsna, G; Kashyap, Amita

    2016-01-01

    This brief highlights advances in DNA technologies and their wider applications. DNA is the source of life and has been studied since a generation, but very little is known as yet. Several sophisticated technologies of the current era have laid their foundations on the principle of DNA based mechanisms. DNA based technologies are bringing a new revolution of Advanced Science and Technology. Forensic Investigation, Medical Diagnosis, Paternity Disputes, Individual Identity, Health insurance, Motor Insurance have incorporated the DNA testing and profiling technologies for settling the issues.

  14. DNA AND ITS METAPHORES

    Directory of Open Access Journals (Sweden)

    Jan Domaradzki

    2015-04-01

    Full Text Available The aim of the present paper is to describe the main metaphors presented in genetic discourse: DNA as text, information, language, book, code, project/blueprint, map, computer, music, and cooking. It also analyses the social implication of these metaphors. The author of this article argues that metaphors are double-edged swords: while they brighten difficult and abstract genetic concepts, they also lead to the misunderstanding and misinterpretation of the reality. The reason for this is that most of these metaphors are of deterministic, reductionist, and fatalistic character. Consequently, they shift the attention from complexity of genetic processes. Moreover, as they appeal to emotions, ascetics, and morality they may involve exaggeration: while they bring hope, they also create an atmosphere of fear over the misuse of genetic knowledge. The author of this article states that the genetic metaphors do not simply reflect the social ideas on DNA, but also shape our understanding of genetics and imagination on the social application of genetic knowledge. Due to this reason, DNA should be understood not only as a biological code, but as a cultural as well.

  15. DNA adducts-chemical addons

    Directory of Open Access Journals (Sweden)

    T R Rajalakshmi

    2015-01-01

    Full Text Available DNA adduct is a piece of DNA covalently bond to a chemical (safrole, benzopyrenediol epoxide, acetaldehyde. This process could be the start of a cancerous cell. When a chemical binds to DNA, it gets damaged resulting in abnormal replication. This could be the start of a mutation and without proper DNA repair, this can lead to cancer. It is this chemical that binds with the DNA is our prime area of concern. Instead of performing the whole body analysis for diagnosing cancer, this test could be carried out for early detection of cancer. When scanning tunneling microscope is used, the DNA results can be obtained earlier. DNA adducts in scientific experiments are used as biomarkers.

  16. Mitochondrial DNA repair and aging

    International Nuclear Information System (INIS)

    Mandavilli, Bhaskar S.; Santos, Janine H.; Van Houten, Bennett

    2002-01-01

    The mitochondrial electron transport chain plays an important role in energy production in aerobic organisms and is also a significant source of reactive oxygen species that damage DNA, RNA and proteins in the cell. Oxidative damage to the mitochondrial DNA is implicated in various degenerative diseases, cancer and aging. The importance of mitochondrial ROS in age-related degenerative diseases is further strengthened by studies using animal models, Caenorhabditis elegans, Drosophila and yeast. Research in the last several years shows that mitochondrial DNA is more susceptible to various carcinogens and ROS when compared to nuclear DNA. DNA damage in mammalian mitochondria is repaired by base excision repair (BER). Studies have shown that mitochondria contain all the enzymes required for BER. Mitochondrial DNA damage, if not repaired, leads to disruption of electron transport chain and production of more ROS. This vicious cycle of ROS production and mtDNA damage ultimately leads to energy depletion in the cell and apoptosis

  17. Mitochondrial DNA repair and aging

    Energy Technology Data Exchange (ETDEWEB)

    Mandavilli, Bhaskar S.; Santos, Janine H.; Van Houten, Bennett

    2002-11-30

    The mitochondrial electron transport chain plays an important role in energy production in aerobic organisms and is also a significant source of reactive oxygen species that damage DNA, RNA and proteins in the cell. Oxidative damage to the mitochondrial DNA is implicated in various degenerative diseases, cancer and aging. The importance of mitochondrial ROS in age-related degenerative diseases is further strengthened by studies using animal models, Caenorhabditis elegans, Drosophila and yeast. Research in the last several years shows that mitochondrial DNA is more susceptible to various carcinogens and ROS when compared to nuclear DNA. DNA damage in mammalian mitochondria is repaired by base excision repair (BER). Studies have shown that mitochondria contain all the enzymes required for BER. Mitochondrial DNA damage, if not repaired, leads to disruption of electron transport chain and production of more ROS. This vicious cycle of ROS production and mtDNA damage ultimately leads to energy depletion in the cell and apoptosis.

  18. Radiation damage of DNA. Model for direct ionization of DNA

    International Nuclear Information System (INIS)

    Kobayashi, Kazuo; Tagawa, Seiichi

    2004-01-01

    Current aspects of radiation damage of DNA, particularly induced by the direct effect of radiation, and author's method of pulse radiolysis are described in relation to behavior of ions formed by radiation and active principles to induce the strand break. In irradiation of DNA solution in water, the direct effect of radiation is derived from ionization of DNA itself and indirect one, from the reaction between DNA and radicals generated from water molecules and the former direct one has been scarcely investigated due to difficulty of experimental approach. Radicals generated in sugar moiety of DNA are shown important in the strand break by recent studies on crystalline DNA irradiated by X-ray, DNA solution by electron and photon beams, hydrated DNA by γ-ray and by high linear energy transfer (LET) ion. Author's pulse radiolysis studies have revealed behaviors of guanine and adenine radical cations in dynamics of DNA oxidation. Since reactions described are the model, the experimental approach is thought necessary for elucidation of the actually occurring DNA damage in living cells. (N.I.)

  19. DNA stable-isotope probing (DNA-SIP).

    Science.gov (United States)

    Dunford, Eric A; Neufeld, Josh D

    2010-08-02

    DNA stable-isotope probing (DNA-SIP) is a powerful technique for identifying active microorganisms that assimilate particular carbon substrates and nutrients into cellular biomass. As such, this cultivation-independent technique has been an important methodology for assigning metabolic function to the diverse communities inhabiting a wide range of terrestrial and aquatic environments. Following the incubation of an environmental sample with stable-isotope labelled compounds, extracted nucleic acid is subjected to density gradient ultracentrifugation and subsequent gradient fractionation to separate nucleic acids of differing densities. Purification of DNA from cesium chloride retrieves labelled and unlabelled DNA for subsequent molecular characterization (e.g. fingerprinting, microarrays, clone libraries, metagenomics). This JoVE video protocol provides visual step-by-step explanations of the protocol for density gradient ultracentrifugation, gradient fractionation and recovery of labelled DNA. The protocol also includes sample SIP data and highlights important tips and cautions that must be considered to ensure a successful DNA-SIP analysis.

  20. Dynamics of DNA conformations and DNA-protein interaction

    DEFF Research Database (Denmark)

    Metzler, R.; Ambjörnsson, T.; Lomholt, Michael Andersen

    2005-01-01

    Optical tweezers, atomic force microscopes, patch clamping, or fluorescence techniques make it possible to study both the equilibrium conformations and dynamics of single DNA molecules as well as their interaction with binding proteins. In this paper we address the dynamics of local DNA...... denaturation (bubble breathing), deriving its dynamic response to external physical parameters and the DNA sequence in terms of the bubble relaxation time spectrum and the autocorrelation function of bubble breathing. The interaction with binding proteins that selectively bind to the DNA single strand exposed...... in a denaturation bubble are shown to involve an interesting competition of time scales, varying between kinetic blocking of protein binding up to full binding protein-induced denaturation of the DNA. We will also address the potential to use DNA physics for the design of nanosensors. Finally, we report recent...

  1. DNA adducts as molecular dosimeters

    International Nuclear Information System (INIS)

    Lucier, G.W.

    1990-01-01

    There is compelling evidence that DNA adducts play an important role in the actions of many pulmonary carcinogens. During the last ten years sensitive methods (antibodies and 32 P-postlabeling) have been developed that permit detection of DNA adducts in tissues of animals or humans exposed to low levels of some genotoxic carcinogens. This capability has led to approaches designed to more reliably estimate the shape of the dose-response curve in the low dose region for a few carcinogens. Moreover, dosimetry comparisions can, in some cases, be made between animals and humans which help in judging the adequacy of animal models for human risk assessments. There are several points that need to be considered in the evaluation of DNA adducts as a molecular dosimeter. For example, DNA adduct formation is only one of many events that are needed for tumor development and some potent carcinogens do not form DNA adducts; i.e., TCDD. Other issues that need to be considered are DNA adduct heterogeneity, DNA repair, relationship of DNA adducts to somatic mutation and cell specificity in DNA adduct formation and persistence. Molecular epidemiology studies often require quantitation of adducts in cells such as lymphocytes which may or may not be reliable surrogates for adduct concentrations in target issues. In summary, accurate quantitation of low levels of DNA adducts may provide data useful in species to species extrapolation of risk including the development of more meaningful human monitoring programs

  2. DNA-based watermarks using the DNA-Crypt algorithm

    Science.gov (United States)

    Heider, Dominik; Barnekow, Angelika

    2007-01-01

    Background The aim of this paper is to demonstrate the application of watermarks based on DNA sequences to identify the unauthorized use of genetically modified organisms (GMOs) protected by patents. Predicted mutations in the genome can be corrected by the DNA-Crypt program leaving the encrypted information intact. Existing DNA cryptographic and steganographic algorithms use synthetic DNA sequences to store binary information however, although these sequences can be used for authentication, they may change the target DNA sequence when introduced into living organisms. Results The DNA-Crypt algorithm and image steganography are based on the same watermark-hiding principle, namely using the least significant base in case of DNA-Crypt and the least significant bit in case of the image steganography. It can be combined with binary encryption algorithms like AES, RSA or Blowfish. DNA-Crypt is able to correct mutations in the target DNA with several mutation correction codes such as the Hamming-code or the WDH-code. Mutations which can occur infrequently may destroy the encrypted information, however an integrated fuzzy controller decides on a set of heuristics based on three input dimensions, and recommends whether or not to use a correction code. These three input dimensions are the length of the sequence, the individual mutation rate and the stability over time, which is represented by the number of generations. In silico experiments using the Ypt7 in Saccharomyces cerevisiae shows that the DNA watermarks produced by DNA-Crypt do not alter the translation of mRNA into protein. Conclusion The program is able to store watermarks in living organisms and can maintain the original information by correcting mutations itself. Pairwise or multiple sequence alignments show that DNA-Crypt produces few mismatches between the sequences similar to all steganographic algorithms. PMID:17535434

  3. Superimposed Code Theorectic Analysis of DNA Codes and DNA Computing

    Science.gov (United States)

    2010-03-01

    that the hybridization that occurs between a DNA strand and its Watson - Crick complement can be used to perform mathematical computation. This research...ssDNA single stranded DNA WC Watson – Crick A Adenine C Cytosine G Guanine T Thymine ... Watson - Crick (WC) duplex, e.g., TCGCA TCGCA . Note that non-WC duplexes can form and such a formation is called a cross-hybridization. Cross

  4. DNA-based watermarks using the DNA-Crypt algorithm

    Directory of Open Access Journals (Sweden)

    Barnekow Angelika

    2007-05-01

    Full Text Available Abstract Background The aim of this paper is to demonstrate the application of watermarks based on DNA sequences to identify the unauthorized use of genetically modified organisms (GMOs protected by patents. Predicted mutations in the genome can be corrected by the DNA-Crypt program leaving the encrypted information intact. Existing DNA cryptographic and steganographic algorithms use synthetic DNA sequences to store binary information however, although these sequences can be used for authentication, they may change the target DNA sequence when introduced into living organisms. Results The DNA-Crypt algorithm and image steganography are based on the same watermark-hiding principle, namely using the least significant base in case of DNA-Crypt and the least significant bit in case of the image steganography. It can be combined with binary encryption algorithms like AES, RSA or Blowfish. DNA-Crypt is able to correct mutations in the target DNA with several mutation correction codes such as the Hamming-code or the WDH-code. Mutations which can occur infrequently may destroy the encrypted information, however an integrated fuzzy controller decides on a set of heuristics based on three input dimensions, and recommends whether or not to use a correction code. These three input dimensions are the length of the sequence, the individual mutation rate and the stability over time, which is represented by the number of generations. In silico experiments using the Ypt7 in Saccharomyces cerevisiae shows that the DNA watermarks produced by DNA-Crypt do not alter the translation of mRNA into protein. Conclusion The program is able to store watermarks in living organisms and can maintain the original information by correcting mutations itself. Pairwise or multiple sequence alignments show that DNA-Crypt produces few mismatches between the sequences similar to all steganographic algorithms.

  5. DNA-based watermarks using the DNA-Crypt algorithm.

    Science.gov (United States)

    Heider, Dominik; Barnekow, Angelika

    2007-05-29

    The aim of this paper is to demonstrate the application of watermarks based on DNA sequences to identify the unauthorized use of genetically modified organisms (GMOs) protected by patents. Predicted mutations in the genome can be corrected by the DNA-Crypt program leaving the encrypted information intact. Existing DNA cryptographic and steganographic algorithms use synthetic DNA sequences to store binary information however, although these sequences can be used for authentication, they may change the target DNA sequence when introduced into living organisms. The DNA-Crypt algorithm and image steganography are based on the same watermark-hiding principle, namely using the least significant base in case of DNA-Crypt and the least significant bit in case of the image steganography. It can be combined with binary encryption algorithms like AES, RSA or Blowfish. DNA-Crypt is able to correct mutations in the target DNA with several mutation correction codes such as the Hamming-code or the WDH-code. Mutations which can occur infrequently may destroy the encrypted information, however an integrated fuzzy controller decides on a set of heuristics based on three input dimensions, and recommends whether or not to use a correction code. These three input dimensions are the length of the sequence, the individual mutation rate and the stability over time, which is represented by the number of generations. In silico experiments using the Ypt7 in Saccharomyces cerevisiae shows that the DNA watermarks produced by DNA-Crypt do not alter the translation of mRNA into protein. The program is able to store watermarks in living organisms and can maintain the original information by correcting mutations itself. Pairwise or multiple sequence alignments show that DNA-Crypt produces few mismatches between the sequences similar to all steganographic algorithms.

  6. The DNA Files

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1998-06-09

    The DNA Files is a radio documentary which disseminates genetics information over public radio. The documentaries explore subjects which include the following: How genetics affects society. How human life began and how it evolved. Could new prenatal genetic tests hold the key to disease prevention later in life? Would a national genetic data base sacrifice individual privacy? and Should genes that may lead to the cure for cancer be privately owned? This report serves as a project update for the second quarter of 1998. It includes the spring/summer 1998 newsletter, the winter 1998 newsletter, the program clock, and the latest flyer.

  7. Towards a DNA Nanoprocessor: Reusable Tile-Integrated DNA Circuits.

    Science.gov (United States)

    Gerasimova, Yulia V; Kolpashchikov, Dmitry M

    2016-08-22

    Modern electronic microprocessors use semiconductor logic gates organized on a silicon chip to enable efficient inter-gate communication. Here, arrays of communicating DNA logic gates integrated on a single DNA tile were designed and used to process nucleic acid inputs in a reusable format. Our results lay the foundation for the development of a DNA nanoprocessor, a small and biocompatible device capable of performing complex analyses of DNA and RNA inputs. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. DNA-Conjugated Organic Chromophores in DNA Stacking Interactions

    DEFF Research Database (Denmark)

    Filichev, Vyacheslav V.; Pedersen, Erik Bjerregaard

    2009-01-01

    Since the discovery of the intercalation of acridine derivatives into DNA (1961), chemists have synthesized many intercalators tethered to DNA. Advances in the chemical synthesis of modified nucleosides along with progress in oligonucleotide synthesis have made it possible to introduce organic ch...... review presents those efforts in the design of intercalators/organic chromophores as oligonucleotide conjugates that form a foundation for the generation of novel nucleic acid architectures......Since the discovery of the intercalation of acridine derivatives into DNA (1961), chemists have synthesized many intercalators tethered to DNA. Advances in the chemical synthesis of modified nucleosides along with progress in oligonucleotide synthesis have made it possible to introduce organic...

  9. DNA methylation and memory formation.

    Science.gov (United States)

    Day, Jeremy J; Sweatt, J David

    2010-11-01

    Memory formation and storage require long-lasting changes in memory-related neuronal circuits. Recent evidence indicates that DNA methylation may serve as a contributing mechanism in memory formation and storage. These emerging findings suggest a role for an epigenetic mechanism in learning and long-term memory maintenance and raise apparent conundrums and questions. For example, it is unclear how DNA methylation might be reversed during the formation of a memory, how changes in DNA methylation alter neuronal function to promote memory formation, and how DNA methylation patterns differ between neuronal structures to enable both consolidation and storage of memories. Here we evaluate the existing evidence supporting a role for DNA methylation in memory, discuss how DNA methylation may affect genetic and neuronal function to contribute to behavior, propose several future directions for the emerging subfield of neuroepigenetics, and begin to address some of the broader implications of this work.

  10. Monitoring Biodiversity using Environmental DNA

    DEFF Research Database (Denmark)

    Thomsen, Philip Francis

    DNA). Especially the advance in DNA sequencing technology has revolutionized this field and opened new frontiers in ecology, evolution and environmental sciences. Also, it is becoming a powerful tool for field biologist, with new and efficient methods for monitoring biodiversity. This thesis focuses on the use...... of eDNA in monitoring of biodiversity in different settings. First, it is shown that a diversity of rare freshwater animals – representing amphibians, fish, mammals, insects and crustaceans – can be detected based on eDNA obtained directly from 15 ml water samples of lakes, ponds and streams...... setting, showing that eDNA obtained directly from ½ l seawater samples can account for marine fish biodiversity using NGS. Promisingly, eDNA covered the fish diversity better than any of 9 methods, conventionally used in marine fish surveys. Additionally, it is shown that even short 100-bp. fish e...

  11. Loss of maintenance DNA methylation results in abnormal DNA origin firing during DNA replication

    Energy Technology Data Exchange (ETDEWEB)

    Haruta, Mayumi [Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Shimada, Midori, E-mail: midorism@med.nagoya-cu.ac.jp [Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Nishiyama, Atsuya; Johmura, Yoshikazu [Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Le Tallec, Benoît; Debatisse, Michelle [Institut Curie, Centre de Recherche, 26 rue d’Ulm, CNRS UMR 3244, 75248 ParisCedex 05 (France); Nakanishi, Makoto, E-mail: mkt-naka@med.nagoya-cu.ac.jp [Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan)

    2016-01-22

    The mammalian maintenance methyltransferase DNMT1 [DNA (cytosine-5-)-methyltransferase 1] mediates the inheritance of the DNA methylation pattern during replication. Previous studies have shown that depletion of DNMT1 causes a severe growth defect and apoptosis in differentiated cells. However, the detailed mechanisms behind this phenomenon remain poorly understood. Here we show that conditional ablation of Dnmt1 in murine embryonic fibroblasts (MEFs) resulted in an aberrant DNA replication program showing an accumulation of late-S phase replication and causing severely defective growth. Furthermore, we found that the catalytic activity and replication focus targeting sequence of DNMT1 are required for a proper DNA replication program. Taken together, our findings suggest that the maintenance of DNA methylation by DNMT1 plays a critical role in proper regulation of DNA replication in mammalian cells. - Highlights: • DNMT1 depletion results in an abnormal DNA replication program. • Aberrant DNA replication is independent of the DNA damage checkpoint in DNMT1cKO. • DNMT1 catalytic activity and RFT domain are required for proper DNA replication. • DNMT1 catalytic activity and RFT domain are required for cell proliferation.

  12. Human DNA ligase III bridges two DNA ends to promote specific intermolecular DNA end joining

    Science.gov (United States)

    Kukshal, Vandna; Kim, In-Kwon; Hura, Gregory L.; Tomkinson, Alan E.; Tainer, John A.; Ellenberger, Tom

    2015-01-01

    Mammalian DNA ligase III (LigIII) functions in both nuclear and mitochondrial DNA metabolism. In the nucleus, LigIII has functional redundancy with DNA ligase I whereas LigIII is the only mitochondrial DNA ligase and is essential for the survival of cells dependent upon oxidative respiration. The unique LigIII zinc finger (ZnF) domain is not required for catalytic activity but senses DNA strand breaks and stimulates intermolecular ligation of two DNAs by an unknown mechanism. Consistent with this activity, LigIII acts in an alternative pathway of DNA double strand break repair that buttresses canonical non-homologous end joining (NHEJ) and is manifest in NHEJ-defective cancer cells, but how LigIII acts in joining intermolecular DNA ends versus nick ligation is unclear. To investigate how LigIII efficiently joins two DNAs, we developed a real-time, fluorescence-based assay of DNA bridging suitable for high-throughput screening. On a nicked duplex DNA substrate, the results reveal binding competition between the ZnF and the oligonucleotide/oligosaccharide-binding domain, one of three domains constituting the LigIII catalytic core. In contrast, these domains collaborate and are essential for formation of a DNA-bridging intermediate by adenylated LigIII that positions a pair of blunt-ended duplex DNAs for efficient and specific intermolecular ligation. PMID:26130724

  13. Loss of maintenance DNA methylation results in abnormal DNA origin firing during DNA replication

    International Nuclear Information System (INIS)

    Haruta, Mayumi; Shimada, Midori; Nishiyama, Atsuya; Johmura, Yoshikazu; Le Tallec, Benoît; Debatisse, Michelle; Nakanishi, Makoto

    2016-01-01

    The mammalian maintenance methyltransferase DNMT1 [DNA (cytosine-5-)-methyltransferase 1] mediates the inheritance of the DNA methylation pattern during replication. Previous studies have shown that depletion of DNMT1 causes a severe growth defect and apoptosis in differentiated cells. However, the detailed mechanisms behind this phenomenon remain poorly understood. Here we show that conditional ablation of Dnmt1 in murine embryonic fibroblasts (MEFs) resulted in an aberrant DNA replication program showing an accumulation of late-S phase replication and causing severely defective growth. Furthermore, we found that the catalytic activity and replication focus targeting sequence of DNMT1 are required for a proper DNA replication program. Taken together, our findings suggest that the maintenance of DNA methylation by DNMT1 plays a critical role in proper regulation of DNA replication in mammalian cells. - Highlights: • DNMT1 depletion results in an abnormal DNA replication program. • Aberrant DNA replication is independent of the DNA damage checkpoint in DNMT1cKO. • DNMT1 catalytic activity and RFT domain are required for proper DNA replication. • DNMT1 catalytic activity and RFT domain are required for cell proliferation.

  14. DNA controlled assembly of liposomes

    DEFF Research Database (Denmark)

    Vogel, Stefan; Jakobsen, Ulla; Simonsen, Adam Cohen

    2009-01-01

    DNA-encoding of solid nanoparticles requires surfacechemistry, which is often tedious and not generally applicable. In the present study non-covalently attached DNA are used to assemble soft nanoparticles (liposomes) in solution. This process displays remarkably sharp thermal transitions from...... assembled to disassembled state for which reason this method allows easy and fast detection of polynucleotides (e.g. DNA or RNA), including single nucleotide polymorphisms as well as insertions and deletions....

  15. Biosensors for DNA sequence detection

    Science.gov (United States)

    Vercoutere, Wenonah; Akeson, Mark

    2002-01-01

    DNA biosensors are being developed as alternatives to conventional DNA microarrays. These devices couple signal transduction directly to sequence recognition. Some of the most sensitive and functional technologies use fibre optics or electrochemical sensors in combination with DNA hybridization. In a shift from sequence recognition by hybridization, two emerging single-molecule techniques read sequence composition using zero-mode waveguides or electrical impedance in nanoscale pores.

  16. DNA damage, homology-directed repair, and DNA methylation.

    Directory of Open Access Journals (Sweden)

    Concetta Cuozzo

    2007-07-01

    Full Text Available To explore the link between DNA damage and gene silencing, we induced a DNA double-strand break in the genome of Hela or mouse embryonic stem (ES cells using I-SceI restriction endonuclease. The I-SceI site lies within one copy of two inactivated tandem repeated green fluorescent protein (GFP genes (DR-GFP. A total of 2%-4% of the cells generated a functional GFP by homology-directed repair (HR and gene conversion. However, approximately 50% of these recombinants expressed GFP poorly. Silencing was rapid and associated with HR and DNA methylation of the recombinant gene, since it was prevented in Hela cells by 5-aza-2'-deoxycytidine. ES cells deficient in DNA methyl transferase 1 yielded as many recombinants as wild-type cells, but most of these recombinants expressed GFP robustly. Half of the HR DNA molecules were de novo methylated, principally downstream to the double-strand break, and half were undermethylated relative to the uncut DNA. Methylation of the repaired gene was independent of the methylation status of the converting template. The methylation pattern of recombinant molecules derived from pools of cells carrying DR-GFP at different loci, or from an individual clone carrying DR-GFP at a single locus, was comparable. ClustalW analysis of the sequenced GFP molecules in Hela and ES cells distinguished recombinant and nonrecombinant DNA solely on the basis of their methylation profile and indicated that HR superimposed novel methylation profiles on top of the old patterns. Chromatin immunoprecipitation and RNA analysis revealed that DNA methyl transferase 1 was bound specifically to HR GFP DNA and that methylation of the repaired segment contributed to the silencing of GFP expression. Taken together, our data support a mechanistic link between HR and DNA methylation and suggest that DNA methylation in eukaryotes marks homologous recombined segments.

  17. Fluorescence Microscopy of Nanochannel-Confined DNA.

    Science.gov (United States)

    Westerlund, Fredrik; Persson, Fredrik; Fritzsche, Joachim; Beech, Jason P; Tegenfeldt, Jonas O

    2018-01-01

    Stretching of DNA in nanoscale confinement allows for several important studies. The genetic contents of the DNA can be visualized on the single DNA molecule level and both the polymer physics of confined DNA and also DNA/protein and other DNA/DNA-binding molecule interactions can be explored. This chapter describes the basic steps to fabricate the nanostructures, perform the experiments and analyze the data.

  18. Aging and DNA repair capability. [Review

    Energy Technology Data Exchange (ETDEWEB)

    Tice, R R

    1977-01-01

    A review of the literature on DNA repair processes in relation to aging is presented under the following headings: DNA repair processes; age-related occurrence of unrepaired DNA lesions; DNA repair capability as a function of age; tissue-specific DNA repair capability; acceleration of the aging process by exposure to DNA damaging agents; human genetic syndromes; and longevity and DNA repair processes. (HLW)

  19. DNA modification by alkylating compounds

    Energy Technology Data Exchange (ETDEWEB)

    Kruglyakova, E.E.

    1985-09-01

    Results are given for research on the physico-chemical properties of alkylating compounds - nitroso alkyl ureas (NAU) which possess a broad spectrum of biological activity, such as mutagenic, carcinogenic, and anti-tumor action that is due to the alkylation and carbamoylation of DNA as well as other cellular components. Identified chemical products of NAU interaction with DNA and its components are cited. Structural conversions of a DNA macromolecule resulting from its chemical modification are examined. NAU are used to discuss possible biological consequences of DNA modification. 148 references.

  20. Supercoil Formation During DNA Melting

    Science.gov (United States)

    Sayar, Mehmet; Avsaroglu, Baris; Kabakcioglu, Alkan

    2009-03-01

    Supercoil formation plays a key role in determining the structure-function relationship in DNA. Biological and technological processes, such as protein synthesis, polymerase chain reaction, and microarrays relys on separation of the two strands in DNA, which is coupled to the unwinding of the supercoiled structure. This problem has been studied theoretically via Peyrard-Bishop and Poland-Scheraga type models, which include a simple representation of the DNA structural properties. In recent years, computational models, which provide a more realtistic representaion of DNA molecule, have been used to study the melting behavior of short DNA chains. Here, we will present a new coarse-grained model of DNA which is capable of simulating sufficiently long DNA chains for studying the supercoil formation during melting, without sacrificing the local structural properties. Our coarse-grained model successfully reproduces the local geometry of the DNA molecule, such as the 3'-5' directionality, major-minor groove structure, and the helical pitch. We will present our initial results on the dynamics of supercoiling during DNA melting.

  1. Monogenic diseases of DNA repair

    DEFF Research Database (Denmark)

    Keijzers, Guido; Bakula, Daniela; Scheibye-Knudsen, Morten

    2017-01-01

    Maintaining the stability of the genome is essential for all organisms, and it is not surprising that damage to DNA has been proposed as an explanation for multiple chronic diseases.1-5 Conserving a pristine genome is therefore of central importance to our health. To overcome the genotoxic stress...... of a growing number of human diseases. Notably, many of these monogenic DNA-repair disorders display features of accelerated aging, supporting the notion that genome maintenance is a key factor for organismal longevity. This review focuses on the physiological consequences of loss of DNA repair, particularly...... in the context of monogenic DNA-repair diseases....

  2. DNA nanotechnology-enabled biosensors.

    Science.gov (United States)

    Chao, Jie; Zhu, Dan; Zhang, Yinan; Wang, Lianhui; Fan, Chunhai

    2016-02-15

    Biosensors employ biological molecules to recognize the target and utilize output elements which can translate the biorecognition event into electrical, optical or mass-sensitive signals to determine the quantities of the target. DNA-based biosensors, as a sub-field to biosensor, utilize DNA strands with short oligonucleotides as probes for target recognition. Although DNA-based biosensors have offered a promising alternative for fast, simple and cheap detection of target molecules, there still exist key challenges including poor stability and reproducibility that hinder their competition with the current gold standard for DNA assays. By exploiting the self-recognition properties of DNA molecules, researchers have dedicated to make versatile DNA nanostructures in a highly rigid, controllable and functionalized manner, which offers unprecedented opportunities for developing DNA-based biosensors. In this review, we will briefly introduce the recent advances on design and fabrication of static and dynamic DNA nanostructures, and summarize their applications for fabrication and functionalization of DNA-based biosensors. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Statistical Approaches for DNA Barcoding

    DEFF Research Database (Denmark)

    Nielsen, Rasmus; Matz, M.

    2006-01-01

    The use of DNA as a tool for species identification has become known as "DNA barcoding" (Floyd et al., 2002; Hebert et al., 2003; Remigio and Hebert, 2003). The basic idea is straightforward: a small amount of DNA is extracted from the specimen, amplified and sequenced. The gene region sequenced...... is chosen so that it is nearly identical among individuals of the same species, but different between species, and therefore its sequence, can serve as an identification tag for the species ("DNA barcode"). By matching the sequence obtained from an unidentified specimen ("query" sequence) to the database...

  4. Radiobiological significance of DNA repair

    International Nuclear Information System (INIS)

    Kuzin, A.M.

    1978-01-01

    A short outline is given on the history of the problem relating to the repair of radiation injuries, specifically its molecular mechanisms. The most urgent problems which currently confront the researchers are noted. This is a further study on the role of DNA repair in post-radiation recovery, search for ways to activate and suppress DNA repair, investigations into the activity balance of various repair enzymes as well as the problem of errors in the structure of repairing DNA. An important role is attached to the investigations of DNA repair in solving a number of practical problems

  5. DNA repair: keeping it together

    DEFF Research Database (Denmark)

    Lisby, Michael; Rothstein, Rodney

    2004-01-01

    A protein scaffold has been identified that holds a chromosome together in the event of a DNA double-strand break. This scaffold is dependent on Rad52 and the Rad50-Mre11-Xrs2 complex and withstands the pulling forces of the mitotic spindle during DNA damage checkpoint arrest.......A protein scaffold has been identified that holds a chromosome together in the event of a DNA double-strand break. This scaffold is dependent on Rad52 and the Rad50-Mre11-Xrs2 complex and withstands the pulling forces of the mitotic spindle during DNA damage checkpoint arrest....

  6. Loss of maintenance DNA methylation results in abnormal DNA origin firing during DNA replication.

    Science.gov (United States)

    Haruta, Mayumi; Shimada, Midori; Nishiyama, Atsuya; Johmura, Yoshikazu; Le Tallec, Benoît; Debatisse, Michelle; Nakanishi, Makoto

    2016-01-22

    The mammalian maintenance methyltransferase DNMT1 [DNA (cytosine-5-)-methyltransferase 1] mediates the inheritance of the DNA methylation pattern during replication. Previous studies have shown that depletion of DNMT1 causes a severe growth defect and apoptosis in differentiated cells. However, the detailed mechanisms behind this phenomenon remain poorly understood. Here we show that conditional ablation of Dnmt1 in murine embryonic fibroblasts (MEFs) resulted in an aberrant DNA replication program showing an accumulation of late-S phase replication and causing severely defective growth. Furthermore, we found that the catalytic activity and replication focus targeting sequence of DNMT1 are required for a proper DNA replication program. Taken together, our findings suggest that the maintenance of DNA methylation by DNMT1 plays a critical role in proper regulation of DNA replication in mammalian cells. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Clinical strains of acinetobacter classified by DNA-DNA hybridization

    International Nuclear Information System (INIS)

    Tjernberg, I.; Ursing, J.

    1989-01-01

    A collection of Acinetobacter strains consisting of 168 consecutive clinical strains and 30 type and reference strains was studied by DNA-DNA hybridization and a few phenotypic tests. The field strains could be allotted to 13 DNA groups. By means of reference strains ten of these could be identified with groups described by Bouvet and Grimont (1986), while three groups were new; they were given the numbers 13-15. The type strain of A. radioresistens- recently described by Nishimura et al. (1988) - was shown to be a member of DNA group 12, which comprised 31 clinical isolates. Of the 19 strains of A. junii, eight showed hemolytic acitivity on sheep and human blood agar and an additional four strains on human blood agar only. Strains of this species have previously been regarded as non-hemolytic. Reciprocal DNA pairing data for the reference strains of the DNA gropus were treated by UPGMA clustering. The reference strains for A. calcoaceticus, A. baumannii and DNA groups 3 and 13 formed a cluster with about 70% relatedness within the cluster. Other DNA groups joined at levels below 60%. (author)

  8. Clinical strains of acinetobacter classified by DNA-DNA hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Tjernberg, I; Ursing, J [Department of Medical Microbiology, University of Lund, Malmoe General Hospital, Malmoe (Sweden)

    1989-01-01

    A collection of Acinetobacter strains consisting of 168 consecutive clinical strains and 30 type and reference strains was studied by DNA-DNA hybridization and a few phenotypic tests. The field strains could be allotted to 13 DNA groups. By means of reference strains ten of these could be identified with groups described by Bouvet and Grimont (1986), while three groups were new; they were given the numbers 13-15. The type strain of A. radioresistens- recently described by Nishimura et al. (1988) - was shown to be a member of DNA group 12, which comprised 31 clinical isolates. Of the 19 strains of A. junii, eight showed hemolytic acitivity on sheep and human blood agar and an additional four strains on human blood agar only. Strains of this species have previously been regarded as non-hemolytic. Reciprocal DNA pairing data for the reference strains of the DNA gropus were treated by UPGMA clustering. The reference strains for A. calcoaceticus, A. baumannii and DNA groups 3 and 13 formed a cluster with about 70% relatedness within the cluster. Other DNA groups joined at levels below 60%. (author).

  9. Repulsive DNA-DNA interactions accelerate viral DNA packaging in phage phi29

    OpenAIRE

    Keller, Nicholas; delToro, Damian; Grimes, Shelley; Jardine, Paul J.; Smith, Douglas E.

    2014-01-01

    We use optical tweezers to study the effect of attractive versus repulsive DNA-DNA interactions on motor-driven viral packaging. Screening of repulsive interactions accelerates packaging, but induction of attractive interactions by spermidine3+ causes heterogeneous dynamics. Acceleration is observed in a fraction of complexes, but most exhibit slowing and stalling, suggesting that attractive interactions promote nonequilibrium DNA conformations that impede the motor. Thus, repulsive interacti...

  10. Master equation approach to DNA breathing in heteropolymer DNA

    DEFF Research Database (Denmark)

    Ambjörnsson, Tobias; Banik, Suman K; Lomholt, Michael A

    2007-01-01

    After crossing an initial barrier to break the first base-pair (bp) in double-stranded DNA, the disruption of further bps is characterized by free energies up to a few k(B)T. Thermal motion within the DNA double strand therefore causes the opening of intermittent single-stranded denaturation zones......, the DNA bubbles. The unzipping and zipping dynamics of bps at the two zipper forks of a bubble, where the single strand of the denatured zone joins the still intact double strand, can be monitored by single molecule fluorescence or NMR methods. We here establish a dynamic description of this DNA breathing...... in a heteropolymer DNA with given sequence in terms of a master equation that governs the time evolution of the joint probability distribution for the bubble size and position along the sequence. The transfer coefficients are based on the Poland-Scheraga free energy model. We derive the autocorrelation function...

  11. DNA damage in neurodegenerative diseases

    Energy Technology Data Exchange (ETDEWEB)

    Coppedè, Fabio, E-mail: fabio.coppede@med.unipi.it; Migliore, Lucia, E-mail: lucia.migliore@med.unipi.it

    2015-06-15

    Highlights: • Oxidative DNA damage is one of the earliest detectable events in the neurodegenerative process. • The mitochondrial DNA is more vulnerable to oxidative attack than the nuclear DNA. • Cytogenetic damage has been largely documented in Alzheimer's disease patients. • The question of whether DNA damage is cause or consequence of neurodegeneration is still open. • Increasing evidence links DNA damage and repair with epigenetic phenomena. - Abstract: Following the observation of increased oxidative DNA damage in nuclear and mitochondrial DNA extracted from post-mortem brain regions of patients affected by neurodegenerative diseases, the last years of the previous century and the first decade of the present one have been largely dedicated to the search of markers of DNA damage in neuronal samples and peripheral tissues of patients in early, intermediate or late stages of neurodegeneration. Those studies allowed to demonstrate that oxidative DNA damage is one of the earliest detectable events in neurodegeneration, but also revealed cytogenetic damage in neurodegenerative conditions, such as for example a tendency towards chromosome 21 malsegregation in Alzheimer's disease. As it happens for many neurodegenerative risk factors the question of whether DNA damage is cause or consequence of the neurodegenerative process is still open, and probably both is true. The research interest in markers of oxidative stress was shifted, in recent years, towards the search of epigenetic biomarkers of neurodegenerative disorders, following the accumulating evidence of a substantial contribution of epigenetic mechanisms to learning, memory processes, behavioural disorders and neurodegeneration. Increasing evidence is however linking DNA damage and repair with epigenetic phenomena, thereby opening the way to a very attractive and timely research topic in neurodegenerative diseases. We will address those issues in the context of Alzheimer's disease

  12. Authentication of forensic DNA samples.

    Science.gov (United States)

    Frumkin, Dan; Wasserstrom, Adam; Davidson, Ariane; Grafit, Arnon

    2010-02-01

    Over the past twenty years, DNA analysis has revolutionized forensic science, and has become a dominant tool in law enforcement. Today, DNA evidence is key to the conviction or exoneration of suspects of various types of crime, from theft to rape and murder. However, the disturbing possibility that DNA evidence can be faked has been overlooked. It turns out that standard molecular biology techniques such as PCR, molecular cloning, and recently developed whole genome amplification (WGA), enable anyone with basic equipment and know-how to produce practically unlimited amounts of in vitro synthesized (artificial) DNA with any desired genetic profile. This artificial DNA can then be applied to surfaces of objects or incorporated into genuine human tissues and planted in crime scenes. Here we show that the current forensic procedure fails to distinguish between such samples of blood, saliva, and touched surfaces with artificial DNA, and corresponding samples with in vivo generated (natural) DNA. Furthermore, genotyping of both artificial and natural samples with Profiler Plus((R)) yielded full profiles with no anomalies. In order to effectively deal with this problem, we developed an authentication assay, which distinguishes between natural and artificial DNA based on methylation analysis of a set of genomic loci: in natural DNA, some loci are methylated and others are unmethylated, while in artificial DNA all loci are unmethylated. The assay was tested on natural and artificial samples of blood, saliva, and touched surfaces, with complete success. Adopting an authentication assay for casework samples as part of the forensic procedure is necessary for maintaining the high credibility of DNA evidence in the judiciary system.

  13. Effects of radiation on DNA

    International Nuclear Information System (INIS)

    Medina, V.F.O.

    1978-01-01

    Irradiation has been shown to depress DNA (deoxyribonucleic acid) synthesis resulting in deficient DNA synthesis. In one experiment, Hela S 3 cells completed the next division after a dose of 500 rads to 200 kw X-rays. Another experiment showed that the amount of DNA synthesized was dependent on the stage in the generation cycle at which the cells are irradiated (Giffites and Tolmach, 1975). DNA synthesis was measured by radioactive thymidine incorporation. The smallest deficiency (20-35%) after a dose of 500 rad X-ray was observed in Hela S 3 cells irradiated in early G 1 or early G 2 , while the greatest deficiency (55-70*) after 500 rad X-ray was found in cells irradiated at mitosis or at the Gsub(1)/S transition. Using velocity sedimentation in alkaline gradients of the DNA from hamster, Elkind, et al 1972, studied repair processes as a function of X-ray dose. DNA containing material released by alkaline lysis was found initially contained in a complex-containing lipid, the sedimentation of which was anomalous relative to denatured RNA from unirradated cells. Doses of X-rays small enough to be in the range that permits high survival (100-800 rads) speed the resolution of single-stranded DNA from this DNA complex, giving rise to a species having a number average molecular weight of 2 x 10 8 daltons. Larger doses greater than 1000 to 2000 rads resulted in a degradation of these DNA strands. Incubation after irradiation resulted in the rapid repair of damage, although the rate of repair of damage to the complex resulted in a reassociation of lipid and DNA. This evidence supports the possibility that a large DNA-membrane structure is a principal target of radiation

  14. DNA damage in neurodegenerative diseases

    International Nuclear Information System (INIS)

    Coppedè, Fabio; Migliore, Lucia

    2015-01-01

    Highlights: • Oxidative DNA damage is one of the earliest detectable events in the neurodegenerative process. • The mitochondrial DNA is more vulnerable to oxidative attack than the nuclear DNA. • Cytogenetic damage has been largely documented in Alzheimer's disease patients. • The question of whether DNA damage is cause or consequence of neurodegeneration is still open. • Increasing evidence links DNA damage and repair with epigenetic phenomena. - Abstract: Following the observation of increased oxidative DNA damage in nuclear and mitochondrial DNA extracted from post-mortem brain regions of patients affected by neurodegenerative diseases, the last years of the previous century and the first decade of the present one have been largely dedicated to the search of markers of DNA damage in neuronal samples and peripheral tissues of patients in early, intermediate or late stages of neurodegeneration. Those studies allowed to demonstrate that oxidative DNA damage is one of the earliest detectable events in neurodegeneration, but also revealed cytogenetic damage in neurodegenerative conditions, such as for example a tendency towards chromosome 21 malsegregation in Alzheimer's disease. As it happens for many neurodegenerative risk factors the question of whether DNA damage is cause or consequence of the neurodegenerative process is still open, and probably both is true. The research interest in markers of oxidative stress was shifted, in recent years, towards the search of epigenetic biomarkers of neurodegenerative disorders, following the accumulating evidence of a substantial contribution of epigenetic mechanisms to learning, memory processes, behavioural disorders and neurodegeneration. Increasing evidence is however linking DNA damage and repair with epigenetic phenomena, thereby opening the way to a very attractive and timely research topic in neurodegenerative diseases. We will address those issues in the context of Alzheimer's disease

  15. DNA Movies and Panspermia

    Directory of Open Access Journals (Sweden)

    Victor Norris

    2011-10-01

    Full Text Available There are several ways that our species might try to send a message to another species separated from us by space and/or time. Synthetic biology might be used to write an epitaph to our species, or simply “Kilroy was here”, in the genome of a bacterium via the patterns of either (1 the codons to exploit Life's non-equilibrium character or (2 the bases themselves to exploit Life's quasi-equilibrium character. We suggest here how DNA movies might be designed using such patterns. We also suggest that a search for mechanisms to create and preserve such patterns might lead to a better understanding of modern cells. Finally, we argue that the cutting-edge microbiology and synthetic biology needed for the Kilroy project would put origin-of-life studies in the vanguard of research.

  16. DNA: Polymer and molecular code

    Science.gov (United States)

    Shivashankar, G. V.

    1999-10-01

    The thesis work focusses upon two aspects of DNA, the polymer and the molecular code. Our approach was to bring single molecule micromanipulation methods to the study of DNA. It included a home built optical microscope combined with an atomic force microscope and an optical tweezer. This combined approach led to a novel method to graft a single DNA molecule onto a force cantilever using the optical tweezer and local heating. With this method, a force versus extension assay of double stranded DNA was realized. The resolution was about 10 picoN. To improve on this force measurement resolution, a simple light backscattering technique was developed and used to probe the DNA polymer flexibility and its fluctuations. It combined the optical tweezer to trap a DNA tethered bead and the laser backscattering to detect the beads Brownian fluctuations. With this technique the resolution was about 0.1 picoN with a millisecond access time, and the whole entropic part of the DNA force-extension was measured. With this experimental strategy, we measured the polymerization of the protein RecA on an isolated double stranded DNA. We observed the progressive decoration of RecA on the l DNA molecule, which results in the extension of l , due to unwinding of the double helix. The dynamics of polymerization, the resulting change in the DNA entropic elasticity and the role of ATP hydrolysis were the main parts of the study. A simple model for RecA assembly on DNA was proposed. This work presents a first step in the study of genetic recombination. Recently we have started a study of equilibrium binding which utilizes fluorescence polarization methods to probe the polymerization of RecA on single stranded DNA. In addition to the study of material properties of DNA and DNA-RecA, we have developed experiments for which the code of the DNA is central. We studied one aspect of DNA as a molecular code, using different techniques. In particular the programmatic use of template specificity makes

  17. DNA Extraction Techniques for Use in Education

    Science.gov (United States)

    Hearn, R. P.; Arblaster, K. E.

    2010-01-01

    DNA extraction provides a hands-on introduction to DNA and enables students to gain real life experience and practical knowledge of DNA. Students gain a sense of ownership and are more enthusiastic when they use their own DNA. A cost effective, simple protocol for DNA extraction and visualization was devised. Buccal mucosal epithelia provide a…

  18. Forensic trace DNA: A review

    NARCIS (Netherlands)

    R.A.H. van Oorschot (Roland ); K. Ballantyne (Kaye); R.J. Mitchell (R. John)

    2010-01-01

    textabstractDNA analysis is frequently used to acquire information from biological material to aid enquiries associated with criminal offences, disaster victim identification and missing persons investigations. As the relevance and value of DNA profiling to forensic investigations has increased, so

  19. Aktionslæringens DNA

    DEFF Research Database (Denmark)

    Madsen, Benedicte

    Aktionslæringen DNA giver en række redskaber til læring i fællesskaber, uanset om der arbejdes med individuelle eller kollektive projekter i offentlig eller privat regi. Metoden danner modvægt til de mere individuelistiske traditioner inden for voksenpædagogikken. DNA-metaforen bruges bogen igennem...

  20. LEGO-like DNA Structures

    DEFF Research Database (Denmark)

    Gothelf, Kurt Vesterager

    2012-01-01

    -dimensional (3D) DNA structures by self-assembly of single-stranded DNA “bricks.” The method opens a new route to complex self-assembled (3D) nanostructures that may serve as addressable templates for placing guest molecules with high precision, with possible applications in biophysics, medicine...

  1. Replication of bacteriophage lambda DNA

    International Nuclear Information System (INIS)

    Tsurimoto, T.; Matsubara, K.

    1983-01-01

    In this paper results of studies on the mechanism of bacteriophage lambda replication using molecular biological and biochemical approaches are reported. The purification of the initiator proteins, O and P, and the role of the O and P proteins in the initiation of lambda DNA replication through interactions with specific DNA sequences are described. 47 references, 15 figures

  2. DNA adducts in senescent cells

    International Nuclear Information System (INIS)

    Gaubatz, J.W.

    1987-01-01

    Perturbations in DNA repair and other metabolic processes during development and aging might affect the steady-state level of genomic damage. The persistence or accumulation of DNA lesions in postmitotic cells could have a significant impact on proper cellular function, interfering with gene regulation for example. To test the notion that DNA damage increases as a function of age in non-dividing cells, DNA was purified from heart tissue of C57BL/6Nia mice at different ages and analyzed by post labeling techniques to detect DNA adducts. In the present experiments, four-dimensional, thin-layer chromatography was used to isolate aromatic adducts that were labeled with carrier-free (γ- 32 P) ATP under DNA-P excess conditions. The complexity and frequency of aromatic adducts varied between DNA samples. Several adducts were present in all preparations and were clearly more abundant in nucleotide maps of mature and old heart DNA. However, a direct correlation with age was not observed. In contrast, experiments in which aromatic adducts were first isolated by phase-transfer to 1-butanol, then labeled with excess (γ- 32 P)ATP indicated that there was an age-related increase in these adducts. The results are consistent with their earlier studies that showed alkyl adducts increased during aging of mouse myocardium and suggest that a common repair pathway might be involved

  3. Authenticity in ancient DNA studies

    DEFF Research Database (Denmark)

    Gilbert, M Thomas P; Willerslev, Eske

    2006-01-01

    Ancient DNA studies represent a powerful tool that can be used to obtain genetic insights into the past. However, despite the publication of large numbers of apparently successful ancient DNA studies, a number of problems exist with the field that are often ignored. Therefore, questions exist as ...

  4. Bubble coalescence in breathing DNA

    DEFF Research Database (Denmark)

    Novotný, Tomas; Pedersen, Jonas Nyvold; Ambjörnsson, Tobias

    2007-01-01

    We investigate the coalescence of two DNA bubbles initially located at weak segments and separated by a more stable barrier region in a designed construct of double-stranded DNA. The characteristic time for bubble coalescence and the corresponding distribution are derived, as well as the distribu...... vicious walkers in opposite potentials....

  5. DNA nanotechnology: a future perspective

    Science.gov (United States)

    2013-01-01

    In addition to its genetic function, DNA is one of the most distinct and smart self-assembling nanomaterials. DNA nanotechnology exploits the predictable self-assembly of DNA oligonucleotides to design and assemble innovative and highly discrete nanostructures. Highly ordered DNA motifs are capable of providing an ultra-fine framework for the next generation of nanofabrications. The majority of these applications are based upon the complementarity of DNA base pairing: adenine with thymine, and guanine with cytosine. DNA provides an intelligent route for the creation of nanoarchitectures with programmable and predictable patterns. DNA strands twist along one helix for a number of bases before switching to the other helix by passing through a crossover junction. The association of two crossovers keeps the helices parallel and holds them tightly together, allowing the assembly of bigger structures. Because of the DNA molecule's unique and novel characteristics, it can easily be applied in a vast variety of multidisciplinary research areas like biomedicine, computer science, nano/optoelectronics, and bionanotechnology. PMID:23497147

  6. Multiscale modelling of DNA mechanics

    Czech Academy of Sciences Publication Activity Database

    Dršata, Tomáš; Lankaš, Filip

    2015-01-01

    Roč. 27, č. 32 (2015), 323102/1-323102/12 ISSN 0953-8984 R&D Projects: GA ČR(CZ) GA14-21893S Institutional support: RVO:61388963 Keywords : DNA elasticity * DNA coarse-grained models * molecular dynamics simulations Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.209, year: 2015

  7. DNA damage by Auger emitters

    International Nuclear Information System (INIS)

    Martin, R.F.; d'Cunha, Glenn; Gibbs, Richard; Murray, Vincent; Pardee, Marshall; Allen, B.J.

    1988-01-01

    125 I atoms can be introduced at specific locations along a defined DNA target molecule, either by site-directed incorporation of an 125 I-labelled deoxynucleotide or by binding of an 125 I-labelled sequence-selective DNA ligand. After allowing accumulation of 125 I decay-induced damage to the DNA, application of DNA sequencing techniques enables positions of strand breaks to be located relative to the site of decay, at a resolution corresponding to the distance between adjacent nucleotides [0.34 nm]. Thus, DNA provides a molecular framework to analyse the extent of damage following [averaged] individual decay events. Results can be compared with energy deposition data generated by computer-simulation methods developed by Charlton et al. The DNA sequencing technique also provides information about the chemical nature of the termini of the DNA chains produced following Auger decay-induced damage. In addition to reviewing the application of this approach to the analysis of 125 I decay induced DNA damage, some more recent results obtained by using 67 Ga are also presented. (author)

  8. DNA-extractie zonder remming

    NARCIS (Netherlands)

    Bonants, P.J.M.; Lee, van der T.A.J.

    2011-01-01

    Moleculaire technieken voor de detectie en identificatie van plantenpathogenen maken gebruik van het DNA of RNA van de ziekteverwekker. Voor een aantal substraten, zoals grond, is de extractie van amplificeerbaar nucleïnezuur een probleem. Tijdens de DNA-extractie uit sommige moeilijke substraten

  9. The journey of DNA repair.

    Science.gov (United States)

    Saini, Natalie

    2015-12-01

    21 years ago, the DNA Repair Enzyme was declared "Molecule of the Year". Today, we are celebrating another "year of repair", with the 2015 Nobel Prize in Chemistry being awarded to Aziz Sancar, Tomas Lindahl and Paul Modrich for their collective work on the different DNA repair pathways.

  10. The DnaA Tale

    DEFF Research Database (Denmark)

    Hansen, Flemming G.; Atlung, Tove

    2018-01-01

    , and translation efficiency, as well as, the DnaA protein, its concentration, its binding to DnaA-boxes, and its binding of ATP or ADP. Furthermore, we will discuss the different models for regulation of initiation which have been proposed over the years, with particular emphasis on the Initiator Titration Model....

  11. Effects of radiation on DNA

    International Nuclear Information System (INIS)

    Braddock, M.

    1985-07-01

    The hydroxyl radical (OH radical) is the most damaging radical produced by the effect of ionizing radiation in water. The rate of reaction of the OH radical with purified, native and isodisperse DNA has been determined as compared with calf thymus DNA. This has been achieved by direct observation of the rate of formation of the DNA-OH radical adduct, and by competition with SCN - . Results obtained from direct observation are consistent with calculations which have been performed using the encounter frequency model of Braams and Ebert. However, results obtained for OH radical with DNA derived from competition plots suggest a rate constant somewhat lower than that obtained from direct observation. The relative merits of both techniques are discussed. In order to study the effect of energy deposited directly in the DNA, dry films of purified plasmid DNA have been irradiated in a system where the indirect effects of radical interaction have been minimized. The present results indicate that with different molecular lengths of plasmid DNA, non-random breakage may occur, and that additional damage may be brought about at sites of previously existing damage. Differences in the sensitivity of plasmid DNA molecules of varying lengths to radiation induced double strand breaks have been demonstrated. (author)

  12. The journey of DNA repair

    OpenAIRE

    Saini, Natalie

    2015-01-01

    21 years ago, the DNA Repair Enzyme was declared “Molecule of the Year”. Today, we are celebrating another “year of repair”, with the 2015 Nobel Prize in Chemistry being awarded to Aziz Sancar, Tomas Lindahl and Paul Modrich for their collective work on the different DNA repair pathways.

  13. Drug Addiction and DNA Modifications.

    Science.gov (United States)

    Brown, Amber N; Feng, Jian

    2017-01-01

    Drug addiction is a complex disorder which can be influenced by both genetic and environmental factors. Research has shown that epigenetic modifications can translate environmental signals into changes in gene expression, suggesting that epigenetic changes may underlie the causes and possibly treatment of substance use disorders. This chapter will focus on epigenetic modifications to DNA, which include DNA methylation and several recently defined additional DNA epigenetic changes. We will discuss the functions of DNA modifications and methods for detecting them, followed by a description of the research investigating the function and consequences of drug-induced changes in DNA methylation patterns. Understanding these epigenetic changes may provide us translational tools for the diagnosis and treatment of addiction in the future.

  14. DNA methylation in metabolic disorders

    DEFF Research Database (Denmark)

    Barres, Romain; Zierath, Juleen R

    2011-01-01

    DNA methylation is a major epigenetic modification that controls gene expression in physiologic and pathologic states. Metabolic diseases such as diabetes and obesity are associated with profound alterations in gene expression that are caused by genetic and environmental factors. Recent reports...... have provided evidence that environmental factors at all ages could modify DNA methylation in somatic tissues, which suggests that DNA methylation is a more dynamic process than previously appreciated. Because of the importance of lifestyle factors in metabolic disorders, DNA methylation provides...... a mechanism by which environmental factors, including diet and exercise, can modify genetic predisposition to disease. This article considers the current evidence that defines a role for DNA methylation in metabolic disorders....

  15. Ancient DNA from marine mammals

    DEFF Research Database (Denmark)

    Foote, Andrew David; Hofreiter, Michael; Morin, Philip A.

    2012-01-01

    such as bone, tooth, baleen, skin, fur, whiskers and scrimshaw using ancient DNA (aDNA) approaches provide an oppor- tunity for investigating such changes over evolutionary and ecological timescales. Here, we review the application of aDNA techniques to the study of marine mammals. Most of the studies have...... focused on detecting changes in genetic diversity following periods of exploitation and environmental change. To date, these studies have shown that even small sample sizes can provide useful information on historical genetic diversity. Ancient DNA has also been used in investigations of changes...... in distribution and range of marine mammal species; we review these studies and discuss the limitations of such ‘presence only’ studies. Combining aDNA data with stable isotopes can provide further insights into changes in ecology and we review past studies and suggest future potential applications. We also...

  16. DNA typing by capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, N.

    1997-10-08

    Capillary electrophoresis is becoming more and more important in nucleic acid analysis including DNA sequencing, typing and disease gene measurements. This work summarized the background of DNA typing. The recent development of capillary electrophoresis was also discussed. The second part of the thesis showed the principle of DNA typing based on using the allelic ladder as the absolute standard ladder in capillary electrophoresis system. Future work will be focused on demonstrating DNA typing on multiplex loci and examples of disease diagnosis in the on-line format of PCR-CE. Also capillary array electrophoresis system should allow high throughput, fast speed DNA typing. Only the introduction and conclusions for this report are available here. A reprint was removed for separate processing.

  17. Multiscale modelling of DNA mechanics

    International Nuclear Information System (INIS)

    Dršata, Tomáš; Lankaš, Filip

    2015-01-01

    Mechanical properties of DNA are important not only in a wide range of biological processes but also in the emerging field of DNA nanotechnology. We review some of the recent developments in modeling these properties, emphasizing the multiscale nature of the problem. Modern atomic resolution, explicit solvent molecular dynamics simulations have contributed to our understanding of DNA fine structure and conformational polymorphism. These simulations may serve as data sources to parameterize rigid base models which themselves have undergone major development. A consistent buildup of larger entities involving multiple rigid bases enables us to describe DNA at more global scales. Free energy methods to impose large strains on DNA, as well as bead models and other approaches, are also briefly discussed. (topical review)

  18. Molecular mechanisms of DNA photodamage

    Energy Technology Data Exchange (ETDEWEB)

    Starrs, S.M

    2000-05-01

    Photodamage in DNA, caused by ultraviolet (UV) light, can occur by direct excitation of the nucleobases or indirectly via the action of photosensitisers. Such, DNA photodamage can be potentially mutagenic or lethal. Among the methods available for detecting UV-induced DNA damage, gel sequencing protocols, utilising synthetic oligodeoxyribonucleotides as targets for UV radiation, allow photolesions to be mapped at nucleotide resolution. This approach has been applied to investigate both DNA damage mechanisms. Following a general overview of DNA photoreactivity, and a description of the main experimental procedures, Chapter 3 identifies the origin of an anomalous mobility shift observed in purine chemical sequence ladders that can confuse the interpretation of DNA cleavage results; measures to abolish this shift are also described. Chapters 4 and 5 examine the alkali-labile DNA damage photosensitised by representative nonsteroidal antiinflammatory drugs (NSAIDs) and the fluoroquinolone antibiotics. Suprofen was the most photoactive NSAID studied, producing different patterns of guanine-specific damage in single-stranded and duplex DNA. Uniform modification of guanine bases, typifying attack by singlet oxygen, was observed in single-stranded oligodeoxyribonucleotides. In duplex molecules, modification was limited to the 5'-G of GG doublets, which is indicative of an electron transfer. The effect of quenchers and photoproduct analysis substantiated these findings. The quinolone, nalidixic acid, behaves similarly. The random base cleavage photosensitised by the fluoroquinolones, has been attributed to free radicals produced during their photodecomposition. Chapter 6 addresses the photoreactivity of purines within unusual DNA structures formed by the repeat sequences (GGA){sub n} and (GA){sub n}, and a minihairpin. There was no definitive evidence for enhanced purine reactivity caused by direct excitation. Finally, Chapter 7 investigates the mutagenic potential of a

  19. Molecular mechanisms of DNA photodamage

    International Nuclear Information System (INIS)

    Starrs, S.M.

    2000-05-01

    Photodamage in DNA, caused by ultraviolet (UV) light, can occur by direct excitation of the nucleobases or indirectly via the action of photosensitisers. Such, DNA photodamage can be potentially mutagenic or lethal. Among the methods available for detecting UV-induced DNA damage, gel sequencing protocols, utilising synthetic oligodeoxyribonucleotides as targets for UV radiation, allow photolesions to be mapped at nucleotide resolution. This approach has been applied to investigate both DNA damage mechanisms. Following a general overview of DNA photoreactivity, and a description of the main experimental procedures, Chapter 3 identifies the origin of an anomalous mobility shift observed in purine chemical sequence ladders that can confuse the interpretation of DNA cleavage results; measures to abolish this shift are also described. Chapters 4 and 5 examine the alkali-labile DNA damage photosensitised by representative nonsteroidal antiinflammatory drugs (NSAIDs) and the fluoroquinolone antibiotics. Suprofen was the most photoactive NSAID studied, producing different patterns of guanine-specific damage in single-stranded and duplex DNA. Uniform modification of guanine bases, typifying attack by singlet oxygen, was observed in single-stranded oligodeoxyribonucleotides. In duplex molecules, modification was limited to the 5'-G of GG doublets, which is indicative of an electron transfer. The effect of quenchers and photoproduct analysis substantiated these findings. The quinolone, nalidixic acid, behaves similarly. The random base cleavage photosensitised by the fluoroquinolones, has been attributed to free radicals produced during their photodecomposition. Chapter 6 addresses the photoreactivity of purines within unusual DNA structures formed by the repeat sequences (GGA) n and (GA) n , and a minihairpin. There was no definitive evidence for enhanced purine reactivity caused by direct excitation. Finally, Chapter 7 investigates the mutagenic potential of a dimeric

  20. RAD51 interconnects between DNA replication, DNA repair and immunity.

    Science.gov (United States)

    Bhattacharya, Souparno; Srinivasan, Kalayarasan; Abdisalaam, Salim; Su, Fengtao; Raj, Prithvi; Dozmorov, Igor; Mishra, Ritu; Wakeland, Edward K; Ghose, Subroto; Mukherjee, Shibani; Asaithamby, Aroumougame

    2017-05-05

    RAD51, a multifunctional protein, plays a central role in DNA replication and homologous recombination repair, and is known to be involved in cancer development. We identified a novel role for RAD51 in innate immune response signaling. Defects in RAD51 lead to the accumulation of self-DNA in the cytoplasm, triggering a STING-mediated innate immune response after replication stress and DNA damage. In the absence of RAD51, the unprotected newly replicated genome is degraded by the exonuclease activity of MRE11, and the fragmented nascent DNA accumulates in the cytosol, initiating an innate immune response. Our data suggest that in addition to playing roles in homologous recombination-mediated DNA double-strand break repair and replication fork processing, RAD51 is also implicated in the suppression of innate immunity. Thus, our study reveals a previously uncharacterized role of RAD51 in initiating immune signaling, placing it at the hub of new interconnections between DNA replication, DNA repair, and immunity. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  1. DNA Topology and the Initiation of Virus DNA Packaging.

    Directory of Open Access Journals (Sweden)

    Choon Seok Oh

    Full Text Available During progeny assembly, viruses selectively package virion genomes from a nucleic acid pool that includes host nucleic acids. For large dsDNA viruses, including tailed bacteriophages and herpesviruses, immature viral DNA is recognized and translocated into a preformed icosahedral shell, the prohead. Recognition involves specific interactions between the viral packaging enzyme, terminase, and viral DNA recognition sites. Generally, viral DNA is recognized by terminase's small subunit (TerS. The large terminase subunit (TerL contains translocation ATPase and endonuclease domains. In phage lambda, TerS binds a sequence repeated three times in cosB, the recognition site. TerS binding to cosB positions TerL to cut the concatemeric DNA at the adjacent nicking site, cosN. TerL introduces staggered nicks in cosN, generating twelve bp cohesive ends. Terminase separates the cohesive ends and remains bound to the cosB-containing end, in a nucleoprotein structure called Complex I. Complex I docks on the prohead's portal vertex and translocation ensues. DNA topology plays a role in the TerSλ-cosBλ interaction. Here we show that a site, I2, located between cosN and cosB, is critically important for an early DNA packaging step. I2 contains a complex static bend. I2 mutations block DNA packaging. I2 mutant DNA is cut by terminase at cosN in vitro, but in vivo, no cos cleavage is detected, nor is there evidence for Complex I. Models for what packaging step might be blocked by I2 mutations are presented.

  2. The role of DNA dependent protein kinase in synapsis of DNA ends

    NARCIS (Netherlands)

    E.P.W.C. Weterings (Eric); N.S. Verkaik (Nicole); H.T. Brüggenwirth (Hennie); D.C. van Gent (Dik); J.H.J. Hoeijmakers (Jan)

    2003-01-01

    textabstractDNA dependent protein kinase (DNA-PK) plays a central role in the non-homologous end-joining pathway of DNA double strand break repair. Its catalytic subunit (DNA-PK(CS)) functions as a serine/threonine protein kinase. We show that DNA-PK forms a stable complex at DNA termini that blocks

  3. Human mitochondrial DNA (mtDNA) types in Malaysia

    International Nuclear Information System (INIS)

    Lian, L.H.; Koh, C.L.; Lim, M.E.

    2000-01-01

    Each human cell contains hundreds of mitochondria and thousands of double-stranded circular mtDNA. The delineation of human mtDNA variation and genetics over the past decade has provided unique and often startling insights into human evolution, degenerative diseases, and aging. Each mtDNA of 16,569 base pairs, encodes 13 polypeptides essential to the enzymes of the mitochondrial energy generating pathway, plus the necessary tRNAs and rRNAs. The highly polymorphic noncoding D-(displacement) loop region, also called the control region, is approximately 1.2 kb long. It contains two well-characterized hypervariable (HV-) regions, HV1 and HV2. MtDNA identification is usually based on these sequence differences. According to the TWTGDAM (Technical Working Group for DNA Analysis Methods), the minimum requirement for a mtDNA database for HV1 is from positions 16024 to 16365 and for HV2, from positions 00073 to 00340. The targeted Malaysian population subgroups for this study were mainly the Malays, Chinese, Indians, and indigenous Ibans, Bidayuhs, Kadazan-Dusuns, and Bajaus. Research methodologies undertaken included DNA extraction of samples from unrelated individuals, amplification of the specific regions via the polymerase chain reaction (PCR), and preparation of template DNA for sequencing by using an automated DNA sequencer. Sufficient nucleotide sequence data were generated from the mtDNA analysis. When the sequences were analyzed, sequence variations were found to be caused by nucleotide substitutions, insertions, and deletions. Of the three causes of the sequence variations, nucleotide substitutions (86.1%) accounted for the vast majority of polymorphism. It is noted that transitions (83.5%) were predominant when compared to the significantly lower frequencies of transversions (2.6%). Insertions (0.9%) and deletions (13.0%) were rather rare and found only in HV2. The data generated will also form the basis of a Malaysian DNA sequence database of mtDNA D

  4. DNA Replication Profiling Using Deep Sequencing.

    Science.gov (United States)

    Saayman, Xanita; Ramos-Pérez, Cristina; Brown, Grant W

    2018-01-01

    Profiling of DNA replication during progression through S phase allows a quantitative snap-shot of replication origin usage and DNA replication fork progression. We present a method for using deep sequencing data to profile DNA replication in S. cerevisiae.

  5. Programme DNA Lattices: Design, Synthesis and Applications

    National Research Council Canada - National Science Library

    Reif, John

    2006-01-01

    .... Self-assembled DNA nanostructures provide a methodology for bottom-up nanoscale construction of highly patterned systems, utilizing macromolecular DNA tiles" composed of branched DNA, self-assembled...

  6. Single gene retrieval from thermally degraded DNA

    Indian Academy of Sciences (India)

    Unknown

    DNA thermal degradation was shown to occur via a singlet oxygen pathway. A comparative study of the ther- mal degradation of cellular DNA and isolated DNA showed that cellular ..... definite level of energy (e.g. depurination active energy,.

  7. An optimised protocol for molecular identification of Eimeria from chickens☆

    Science.gov (United States)

    Kumar, Saroj; Garg, Rajat; Moftah, Abdalgader; Clark, Emily L.; Macdonald, Sarah E.; Chaudhry, Abdul S.; Sparagano, Olivier; Banerjee, Partha S.; Kundu, Krishnendu; Tomley, Fiona M.; Blake, Damer P.

    2014-01-01

    Molecular approaches supporting identification of Eimeria parasites infecting chickens have been available for more than 20 years, although they have largely failed to replace traditional measures such as microscopy and pathology. Limitations of microscopy-led diagnostics, including a requirement for specialist parasitological expertise and low sample throughput, are yet to be outweighed by the difficulties associated with accessing genomic DNA from environmental Eimeria samples. A key step towards the use of Eimeria species-specific PCR as a sensitive and reproducible discriminatory tool for use in the field is the production of a standardised protocol that includes sample collection and DNA template preparation, as well as primer selection from the numerous PCR assays now published. Such a protocol will facilitate development of valuable epidemiological datasets which may be easily compared between studies and laboratories. The outcome of an optimisation process undertaken in laboratories in India and the UK is described here, identifying four steps. First, samples were collected into a 2% (w/v) potassium dichromate solution. Second, oocysts were enriched by flotation in saturated saline. Third, genomic DNA was extracted using a QIAamp DNA Stool mini kit protocol including a mechanical homogenisation step. Finally, nested PCR was carried out using previously published primers targeting the internal transcribed spacer region 1 (ITS-1). Alternative methods tested included sample processing in the presence of faecal material, DNA extraction using a traditional phenol/chloroform protocol, the use of SCAR multiplex PCR (one tube and two tube versions) and speciation using the morphometric tool COCCIMORPH for the first time with field samples. PMID:24138724

  8. Molecular biological mechanisms I. DNA repair

    International Nuclear Information System (INIS)

    Friedl, A.A.

    2000-01-01

    Cells of all living systems possess a variety of mechanisms that allow to repair spontaneous and exogeneously induced DNA damage. DNA repair deficiencies may invoke enhanced sensitivity towards DNA-damaging agents such as ionizing radiation. They may also enhance the risk of cancer development, both spontaneously or after induction. This article reviews several DNA repair mechanisms, especially those dealing with DNA double-strand breaks, and describes hereditary diseases associated with DNA repair defects. (orig.) [de

  9. Pregnancy Constipation: Are Stool Softeners Safe?

    Science.gov (United States)

    ... lifestyle changes. For example: Drink plenty of fluids. Water is a good choice. Prune juice also can help. Include physical activity in your daily routine. Being active can help prevent pregnancy constipation. Include more fiber in your diet. Choose high- ...

  10. EXAMINATION OF FINGERNAIL CONTENTS AND STOOL FOR

    African Journals Online (AJOL)

    Computer Center

    intestinal helminths and protozoa. The objective of this study was to determine the level of fingernail ... prevalence rates of the major protozoan and helminthic infections (1,2). Intestinal parasites have been widely distributed in ..... Methods of Food-borne Disease. Control With Special reference to. Ethiopia. 1997; 32-59. 14.

  11. Stool Test: C. Difficile Toxin (For Parents)

    Science.gov (United States)

    ... sealable container before taking it to the lab. Plastic wrap also can be used to line the diaper of an infant or toddler who isn't yet using the toilet. The wrap should be placed so that urine ... be collected into a clean, dry plastic jar with a screw-cap lid. For best ...

  12. DNA Binding by the Ribosomal DNA Transcription Factor Rrn3 Is Essential for Ribosomal DNA Transcription*

    Science.gov (United States)

    Stepanchick, Ann; Zhi, Huijun; Cavanaugh, Alice H.; Rothblum, Katrina; Schneider, David A.; Rothblum, Lawrence I.

    2013-01-01

    The human homologue of yeast Rrn3 is an RNA polymerase I-associated transcription factor that is essential for ribosomal DNA (rDNA) transcription. The generally accepted model is that Rrn3 functions as a bridge between RNA polymerase I and the transcription factors bound to the committed template. In this model Rrn3 would mediate an interaction between the mammalian Rrn3-polymerase I complex and SL1, the rDNA transcription factor that binds to the core promoter element of the rDNA. In the course of studying the role of Rrn3 in recruitment, we found that Rrn3 was in fact a DNA-binding protein. Analysis of the sequence of Rrn3 identified a domain with sequence similarity to the DNA binding domain of heat shock transcription factor 2. Randomization, or deletion, of the amino acids in this region in Rrn3, amino acids 382–400, abrogated its ability to bind DNA, indicating that this domain was an important contributor to DNA binding by Rrn3. Control experiments demonstrated that these mutant Rrn3 constructs were capable of interacting with both rpa43 and SL1, two other activities demonstrated to be essential for Rrn3 function. However, neither of these Rrn3 mutants was capable of functioning in transcription in vitro. Moreover, although wild-type human Rrn3 complemented a yeast rrn3-ts mutant, the DNA-binding site mutant did not. These results demonstrate that DNA binding by Rrn3 is essential for transcription by RNA polymerase I. PMID:23393135

  13. DNA binding by the ribosomal DNA transcription factor rrn3 is essential for ribosomal DNA transcription.

    Science.gov (United States)

    Stepanchick, Ann; Zhi, Huijun; Cavanaugh, Alice H; Rothblum, Katrina; Schneider, David A; Rothblum, Lawrence I

    2013-03-29

    The human homologue of yeast Rrn3 is an RNA polymerase I-associated transcription factor that is essential for ribosomal DNA (rDNA) transcription. The generally accepted model is that Rrn3 functions as a bridge between RNA polymerase I and the transcription factors bound to the committed template. In this model Rrn3 would mediate an interaction between the mammalian Rrn3-polymerase I complex and SL1, the rDNA transcription factor that binds to the core promoter element of the rDNA. In the course of studying the role of Rrn3 in recruitment, we found that Rrn3 was in fact a DNA-binding protein. Analysis of the sequence of Rrn3 identified a domain with sequence similarity to the DNA binding domain of heat shock transcription factor 2. Randomization, or deletion, of the amino acids in this region in Rrn3, amino acids 382-400, abrogated its ability to bind DNA, indicating that this domain was an important contributor to DNA binding by Rrn3. Control experiments demonstrated that these mutant Rrn3 constructs were capable of interacting with both rpa43 and SL1, two other activities demonstrated to be essential for Rrn3 function. However, neither of these Rrn3 mutants was capable of functioning in transcription in vitro. Moreover, although wild-type human Rrn3 complemented a yeast rrn3-ts mutant, the DNA-binding site mutant did not. These results demonstrate that DNA binding by Rrn3 is essential for transcription by RNA polymerase I.

  14. DNA Damage, Mutagenesis and Cancer

    Directory of Open Access Journals (Sweden)

    Ashis K. Basu

    2018-03-01

    Full Text Available A large number of chemicals and several physical agents, such as UV light and γ-radiation, have been associated with the etiology of human cancer. Generation of DNA damage (also known as DNA adducts or lesions induced by these agents is an important first step in the process of carcinogenesis. Evolutionary processes gave rise to DNA repair tools that are efficient in repairing damaged DNA; yet replication of damaged DNA may take place prior to repair, particularly when they are induced at a high frequency. Damaged DNA replication may lead to gene mutations, which in turn may give rise to altered proteins. Mutations in an oncogene, a tumor-suppressor gene, or a gene that controls the cell cycle can generate a clonal cell population with a distinct advantage in proliferation. Many such events, broadly divided into the stages of initiation, promotion, and progression, which may occur over a long period of time and transpire in the context of chronic exposure to carcinogens, can lead to the induction of human cancer. This is exemplified in the long-term use of tobacco being responsible for an increased risk of lung cancer. This mini-review attempts to summarize this wide area that centers on DNA damage as it relates to the development of human cancer.

  15. Minisequencing mitochondrial DNA pathogenic mutations

    Directory of Open Access Journals (Sweden)

    Carracedo Ángel

    2008-04-01

    Full Text Available Abstract Background There are a number of well-known mutations responsible of common mitochondrial DNA (mtDNA diseases. In order to overcome technical problems related to the analysis of complete mtDNA genomes, a variety of different techniques have been proposed that allow the screening of coding region pathogenic mutations. Methods We here propose a minisequencing assay for the analysis of mtDNA mutations. In a single reaction, we interrogate a total of 25 pathogenic mutations distributed all around the whole mtDNA genome in a sample of patients suspected for mtDNA disease. Results We have detected 11 causal homoplasmic mutations in patients suspected for Leber disease, which were further confirmed by standard automatic sequencing. Mutations m.11778G>A and m.14484T>C occur at higher frequency than expected by change in the Galician (northwest Spain patients carrying haplogroup J lineages (Fisher's Exact test, P-value Conclusion We here developed a minisequencing genotyping method for the screening of the most common pathogenic mtDNA mutations which is simple, fast, and low-cost. The technique is robust and reproducible and can easily be implemented in standard clinical laboratories.

  16. Ancient and modern environmental DNA

    Science.gov (United States)

    Pedersen, Mikkel Winther; Overballe-Petersen, Søren; Ermini, Luca; Sarkissian, Clio Der; Haile, James; Hellstrom, Micaela; Spens, Johan; Thomsen, Philip Francis; Bohmann, Kristine; Cappellini, Enrico; Schnell, Ida Bærholm; Wales, Nathan A.; Carøe, Christian; Campos, Paula F.; Schmidt, Astrid M. Z.; Gilbert, M. Thomas P.; Hansen, Anders J.; Orlando, Ludovic; Willerslev, Eske

    2015-01-01

    DNA obtained from environmental samples such as sediments, ice or water (environmental DNA, eDNA), represents an important source of information on past and present biodiversity. It has revealed an ancient forest in Greenland, extended by several thousand years the survival dates for mainland woolly mammoth in Alaska, and pushed back the dates for spruce survival in Scandinavian ice-free refugia during the last glaciation. More recently, eDNA was used to uncover the past 50 000 years of vegetation history in the Arctic, revealing massive vegetation turnover at the Pleistocene/Holocene transition, with implications for the extinction of megafauna. Furthermore, eDNA can reflect the biodiversity of extant flora and fauna, both qualitatively and quantitatively, allowing detection of rare species. As such, trace studies of plant and vertebrate DNA in the environment have revolutionized our knowledge of biogeography. However, the approach remains marred by biases related to DNA behaviour in environmental settings, incomplete reference databases and false positive results due to contamination. We provide a review of the field. PMID:25487334

  17. One-step multiplex real-time RT-PCR assay for detecting and genotyping wild-type group A rotavirus strains and vaccine strains (Rotarix® and RotaTeq®) in stool samples

    Science.gov (United States)

    Mijatovic-Rustempasic, Slavica; Esona, Mathew D.; Tam, Ka Ian; Quaye, Osbourne; Bowen, Michael D.

    2016-01-01

    Background. Group A rotavirus (RVA) infection is the major cause of acute gastroenteritis (AGE) in young children worldwide. Introduction of two live-attenuated rotavirus vaccines, RotaTeq® and Rotarix®, has dramatically reduced RVA associated AGE and mortality in developed as well as in many developing countries. High-throughput methods are needed to genotype rotavirus wild-type strains and to identify vaccine strains in stool samples. Quantitative RT-PCR assays (qRT-PCR) offer several advantages including increased sensitivity, higher throughput, and faster turnaround time. Methods. In this study, a one-step multiplex qRT-PCR assay was developed to detect and genotype wild-type strains and vaccine (Rotarix® and RotaTeq®) rotavirus strains along with an internal processing control (Xeno or MS2 RNA). Real-time RT-PCR assays were designed for VP7 (G1, G2, G3, G4, G9, G12) and VP4 (P[4], P[6] and P[8]) genotypes. The multiplex qRT-PCR assay also included previously published NSP3 qRT-PCR for rotavirus detection and Rotarix® NSP2 and RotaTeq® VP6 qRT-PCRs for detection of Rotarix® and RotaTeq® vaccine strains respectively. The multiplex qRT-PCR assay was validated using 853 sequence confirmed stool samples and 24 lab cultured strains of different rotavirus genotypes. By using thermostable rTth polymerase enzyme, dsRNA denaturation, reverse transcription (RT) and amplification (PCR) steps were performed in single tube by uninterrupted thermocycling profile to reduce chances of sample cross contamination and for rapid generation of results. For quantification, standard curves were generated using dsRNA transcripts derived from RVA gene segments. Results. The VP7 qRT-PCRs exhibited 98.8–100% sensitivity, 99.7–100% specificity, 85–95% efficiency and a limit of detection of 4–60 copies per singleplex reaction. The VP7 qRT-PCRs exhibited 81–92% efficiency and limit of detection of 150–600 copies in multiplex reactions. The VP4 qRT-PCRs exhibited 98.8

  18. How We Make DNA Origami.

    Science.gov (United States)

    Wagenbauer, Klaus F; Engelhardt, Floris A S; Stahl, Evi; Hechtl, Vera K; Stömmer, Pierre; Seebacher, Fabian; Meregalli, Letizia; Ketterer, Philip; Gerling, Thomas; Dietz, Hendrik

    2017-10-05

    DNA origami has attracted substantial attention since its invention ten years ago, due to the seemingly infinite possibilities that it affords for creating customized nanoscale objects. Although the basic concept of DNA origami is easy to understand, using custom DNA origami in practical applications requires detailed know-how for designing and producing the particles with sufficient quality and for preparing them at appropriate concentrations with the necessary degree of purity in custom environments. Such know-how is not readily available for newcomers to the field, thus slowing down the rate at which new applications outside the field of DNA nanotechnology may emerge. To foster faster progress, we share in this article the experience in making and preparing DNA origami that we have accumulated over recent years. We discuss design solutions for creating advanced structural motifs including corners and various types of hinges that expand the design space for the more rigid multilayer DNA origami and provide guidelines for preventing undesired aggregation and on how to induce specific oligomerization of multiple DNA origami building blocks. In addition, we provide detailed protocols and discuss the expected results for five key methods that allow efficient and damage-free preparation of DNA origami. These methods are agarose-gel purification, filtration through molecular cut-off membranes, PEG precipitation, size-exclusion chromatography, and ultracentrifugation-based sedimentation. The guide for creating advanced design motifs and the detailed protocols with their experimental characterization that we describe here should lower the barrier for researchers to accomplish the full DNA origami production workflow. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. DNA-Controlled Assembly of Soft Nanoparticles

    DEFF Research Database (Denmark)

    Vogel, Stefan

    2015-01-01

    This book covers the emerging topic of DNA nanotechnology and DNA supramolecular chemistry in its broader sense. By taking DNA out of its biological role, this biomolecule has become a very versatile building block in materials chemistry, supramolecular chemistry and bio-nanotechnology. Many nove......-covalent systems, DNA origami, DNA based switches, DNA machines, and alternative structures and templates. This broad coverage is very appealing since it combines both the synthesis of modified DNA as well as designer concepts to successfully plan and make DNA nanostructures....

  20. Radicals of DNA and DNA nucleotides generated by ionising radiation

    International Nuclear Information System (INIS)

    Przybytniak, G.

    2004-01-01

    A first stage of cell processes leading to DNA damage of initiated by radical reactions. In a model system such transformations were generated by ionising radiation which involves production of electron loss and electron gain centers of the substrate and radical formation. Using cryogenic ESR spectroscopy it was found that the DNA nucleotides, which convert to radical anions upon electron capture undergo the separation of unpaired spin and charge due to protonation. Circular and linear dichroism studies enabled to conclude that iron ions(III) induce strong changes in the DNA helical structure indicating their coordination with nitrogen bases. The repair of DNA radicals produced via radiolytic oxidation, i.e. the guanine radical cation and the allyl type radical of thymine, is possible at elevated temperatures due to the involvement of sulphydryl groups. The influence of the thiol charge is then limited

  1. Environmental influences on DNA curvature

    DEFF Research Database (Denmark)

    Ussery, David; Higgins, C.F.; Bolshoy, A.

    1999-01-01

    DNA curvature plays an important role in many biological processes. To study environmentalinfluences on DNA curvature we compared the anomalous migration on polyacrylamide gels ofligation ladders of 11 specifically-designed oligonucleotides. At low temperatures (25 degreesC and below) most......, whilst spermine enhanced theanomalous migration of a different set of sequences. Sequences with a GGC motif exhibitedgreater curvature than predicted by the presently-used angles for the nearest-neighbour wedgemodel and are especially sensitive to Mg2+. The data have implications for models...... for DNAcurvature and for environmentally-sensitive DNA conformations in the regulation of geneexpression....

  2. Normalized cDNA libraries

    Science.gov (United States)

    Soares, Marcelo B.; Efstratiadis, Argiris

    1997-01-01

    This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3' noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to moderate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library.

  3. DNA Probe for Lactobacillus delbrueckii

    Science.gov (United States)

    Delley, Michèle; Mollet, Beat; Hottinger, Herbert

    1990-01-01

    From a genomic DNA library of Lactobacillus delbrueckii subsp. bulgaricus, a clone was isolated which complements a leucine auxotrophy of an Escherichia coli strain (GE891). Subsequent analysis of the clone indicated that it could serve as a specific DNA probe. Dot-blot hybridizations with over 40 different Lactobacillus strains showed that this clone specifically recognizes L. delbrueckii subsp. delbrueckii, bulgaricus, and lactis. The sensitivity of the method was tested by using an α-32P-labeled DNA probe. Images PMID:16348233

  4. DNA Uptake by Transformable Bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Lacks, Sanford A.

    1999-03-31

    The various processes of DNA uptake by cells can be categorized as: viral DNA entry, conjugation, or transformation. Within each category, a variety of mechanisms have been found. However, considerable similarities occur among the different mechanisms of conjugation and, especially, transformation. All of these natural mechanisms of DNA transfer are quite elaborate and involve multiple protein components, as the case may be, of the virus, the donor cell, and the recipient cell. The mechanisms of viral infection and conjugation will be discussed mainly with respect to their relevance to transformation.

  5. DNA UPTAKE BY TRANSFORMABLE BACTERIA

    Energy Technology Data Exchange (ETDEWEB)

    LACKS,S.A.

    1999-09-07

    The various processes of DNA uptake by cells can be categorized as: viral DNA entry, conjugation, or transformation. Within each category, a variety of mechanisms have been found. However, considerable similarities occur among the different mechanisms of conjugation and, especially, transformation. All of these natural mechanisms of DNA transfer are quite elaborate and involve multiple protein components, as the case may be, of the virus, the donor cell, and the recipient cell. The mechanisms of viral infection and conjugation will be discussed mainly with respect to their relevance to transformation.

  6. Recruitment of DNA methyltransferase I to DNA repair sites

    Science.gov (United States)

    Mortusewicz, Oliver; Schermelleh, Lothar; Walter, Joachim; Cardoso, M. Cristina; Leonhardt, Heinrich

    2005-01-01

    In mammalian cells, the replication of genetic and epigenetic information is directly coupled; however, little is known about the maintenance of epigenetic information in DNA repair. Using a laser microirradiation system to introduce DNA lesions at defined subnuclear sites, we tested whether the major DNA methyltransferase (Dnmt1) or one of the two de novo methyltransferases (Dnmt3a, Dnmt3b) are recruited to sites of DNA repair in vivo. Time lapse microscopy of microirradiated mammalian cells expressing GFP-tagged Dnmt1, Dnmt3a, or Dnmt3b1 together with red fluorescent protein-tagged proliferating cell nuclear antigen (PCNA) revealed that Dnmt1 and PCNA accumulate at DNA damage sites as early as 1 min after irradiation in S and non-S phase cells, whereas recruitment of Dnmt3a and Dnmt3b was not observed. Deletion analysis showed that Dnmt1 recruitment was mediated by the PCNA-binding domain. These data point to a direct role of Dnmt1 in the restoration of epigenetic information during DNA repair. PMID:15956212

  7. Bacterial identification and subtyping using DNA microarray and DNA sequencing.

    Science.gov (United States)

    Al-Khaldi, Sufian F; Mossoba, Magdi M; Allard, Marc M; Lienau, E Kurt; Brown, Eric D

    2012-01-01

    The era of fast and accurate discovery of biological sequence motifs in prokaryotic and eukaryotic cells is here. The co-evolution of direct genome sequencing and DNA microarray strategies not only will identify, isotype, and serotype pathogenic bacteria, but also it will aid in the discovery of new gene functions by detecting gene expressions in different diseases and environmental conditions. Microarray bacterial identification has made great advances in working with pure and mixed bacterial samples. The technological advances have moved beyond bacterial gene expression to include bacterial identification and isotyping. Application of new tools such as mid-infrared chemical imaging improves detection of hybridization in DNA microarrays. The research in this field is promising and future work will reveal the potential of infrared technology in bacterial identification. On the other hand, DNA sequencing by using 454 pyrosequencing is so cost effective that the promise of $1,000 per bacterial genome sequence is becoming a reality. Pyrosequencing technology is a simple to use technique that can produce accurate and quantitative analysis of DNA sequences with a great speed. The deposition of massive amounts of bacterial genomic information in databanks is creating fingerprint phylogenetic analysis that will ultimately replace several technologies such as Pulsed Field Gel Electrophoresis. In this chapter, we will review (1) the use of DNA microarray using fluorescence and infrared imaging detection for identification of pathogenic bacteria, and (2) use of pyrosequencing in DNA cluster analysis to fingerprint bacterial phylogenetic trees.

  8. Comparison of five DNA quantification methods

    DEFF Research Database (Denmark)

    Nielsen, Karsten; Mogensen, Helle Smidt; Hedman, Johannes

    2008-01-01

    Six commercial preparations of human genomic DNA were quantified using five quantification methods: UV spectrometry, SYBR-Green dye staining, slot blot hybridization with the probe D17Z1, Quantifiler Human DNA Quantification kit and RB1 rt-PCR. All methods measured higher DNA concentrations than...... Quantification kit in two experiments. The measured DNA concentrations with Quantifiler were 125 and 160% higher than expected based on the manufacturers' information. When the Quantifiler human DNA standard (Raji cell line) was replaced by the commercial human DNA preparation G147A (Promega) to generate the DNA...... standard curve in the Quantifiler Human DNA Quantification kit, the DNA quantification results of the human DNA preparations were 31% higher than expected based on the manufacturers' information. The results indicate a calibration problem with the Quantifiler human DNA standard for its use...

  9. Synchronization of DNA array replication kinetics

    Science.gov (United States)

    Manturov, Alexey O.; Grigoryev, Anton V.

    2016-04-01

    In the present work we discuss the features of the DNA replication kinetics at the case of multiplicity of simultaneously elongated DNA fragments. The interaction between replicated DNA fragments is carried out by free protons that appears at the every nucleotide attachment at the free end of elongated DNA fragment. So there is feedback between free protons concentration and DNA-polymerase activity that appears as elongation rate dependence. We develop the numerical model based on a cellular automaton, which can simulate the elongation stage (growth of DNA strands) for DNA elongation process with conditions pointed above and we study the possibility of the DNA polymerases movement synchronization. The results obtained numerically can be useful for DNA polymerase movement detection and visualization of the elongation process in the case of massive DNA replication, eg, under PCR condition or for DNA "sequencing by synthesis" sequencing devices evaluation.

  10. DNA repair deficiency in neurodegeneration

    DEFF Research Database (Denmark)

    Jeppesen, Dennis Kjølhede; Bohr, Vilhelm A; Stevnsner, Tinna V.

    2011-01-01

    Deficiency in repair of nuclear and mitochondrial DNA damage has been linked to several neurodegenerative disorders. Many recent experimental results indicate that the post-mitotic neurons are particularly prone to accumulation of unrepaired DNA lesions potentially leading to progressive...... neurodegeneration. Nucleotide excision repair is the cellular pathway responsible for removing helix-distorting DNA damage and deficiency in such repair is found in a number of diseases with neurodegenerative phenotypes, including Xeroderma Pigmentosum and Cockayne syndrome. The main pathway for repairing oxidative...... base lesions is base excision repair, and such repair is crucial for neurons given their high rates of oxygen metabolism. Mismatch repair corrects base mispairs generated during replication and evidence indicates that oxidative DNA damage can cause this pathway to expand trinucleotide repeats, thereby...

  11. Sorting fluorescent nanocrystals with DNA

    Energy Technology Data Exchange (ETDEWEB)

    Gerion, Daniele; Parak, Wolfgang J.; Williams, Shara C.; Zanchet, Daniela; Micheel, Christine M.; Alivisatos, A. Paul

    2001-12-10

    Semiconductor nanocrystals with narrow and tunable fluorescence are covalently linked to oligonucleotides. These biocompounds retain the properties of both nanocrystals and DNA. Therefore, different sequences of DNA can be coded with nanocrystals and still preserve their ability to hybridize to their complements. We report the case where four different sequences of DNA are linked to four nanocrystal samples having different colors of emission in the range of 530-640 nm. When the DNA-nanocrystal conjugates are mixed together, it is possible to sort each type of nanoparticle using hybridization on a defined micrometer -size surface containing the complementary oligonucleotide. Detection of sorting requires only a single excitation source and an epifluorescence microscope. The possibility of directing fluorescent nanocrystals towards specific biological targets and detecting them, combined with their superior photo-stability compared to organic dyes, opens the way to improved biolabeling experiments, such as gene mapping on a nanometer scale or multicolor microarray analysis.

  12. DNA photoprotection conveyed by sunscreen.

    Science.gov (United States)

    DeHaven, Charlene; Hayden, Patrick J; Armento, Alexander; Oldach, Jonathan

    2014-06-01

    Skin photoaging is the consequence of solar UV exposure, and DNA damage is an important part of this process. The objective of the current work was to demonstrate that in vitro skin models can be utilized to confirm that DNA damage protection is provided by sunscreens. Skin equivalents were exposed to full-spectrum UV light administered with a standard research solar simulator with and without pre-application of sunscreen. Cyclopyrimidine dimer (CPD) and sunburn cell (SBC) formation as well as CPD quantitation were evaluated to determine DNA damage protection provided by the sunscreen. Marked decreases in both CPDs and SBCs were observed when sunscreen was applied prior to UV exposure. Sunscreen application prior to full-spectrum solar UV exposure protects DNA from photodamage measured by CPD and SBC formation. This can be expected to lessen the risk of photoaging and malignant transformation. © 2014 Wiley Periodicals, Inc.

  13. DNA vaccines for aquacultured fish

    DEFF Research Database (Denmark)

    Lorenzen, Niels; LaPatra, S.E.

    2005-01-01

    of licensing and public acceptance of the technology. The potential benefits of DNA vaccines for farmed fish include improved animal welfare, reduced environmental impacts of aquaculture activities, increased food quality and quantity, and more sustainable production. Testing under commercial production......Deoxyribonucleic acid (DNA) vaccination is based on the administration of the gene encoding the vaccine antigen, rather than the antigen itself. Subsequent expression of the antigen by cells in the vaccinated hosts triggers the host immune system. Among the many experimental DNA vaccines tested...... in various animal species as well as in humans, the vaccines against rhabdovirus diseases in fish have given some of the most promising results. A single intramuscular (IM) injection of microgram amounts of DNA induces rapid and long-lasting protection in farmed salmonids against economically important...

  14. Mammalian DNA Repair. Final Report

    Energy Technology Data Exchange (ETDEWEB)

    Wood, Richard D.

    2003-01-24

    The Gordon Research Conference (GRC) on Mammalian DNA Repair was held at Harbortown Resort, Ventura Beach, CA. Emphasis was placed on current unpublished research and discussion of the future target areas in this field.

  15. (UVB)-induced DNA damage

    African Journals Online (AJOL)

    Jane

    2011-08-17

    dependent cytogenetic lesions were assessed by the micronucleus test (MNT). It was found that POE effectively reduced the extent of DNA breakages and cytogenetic lesions upon exposure to UVB (erythemal ultraviolet (EUV);.

  16. Mitogenomic analyses from ancient DNA

    DEFF Research Database (Denmark)

    Paijmans, Johanna L. A.; Gilbert, Tom; Hofreiter, Michael

    2013-01-01

    The analysis of ancient DNA is playing an increasingly important role in conservation genetic, phylogenetic and population genetic analyses, as it allows incorporating extinct species into DNA sequence trees and adds time depth to population genetics studies. For many years, these types of DNA...... analyses (whether using modern or ancient DNA) were largely restricted to the analysis of short fragments of the mitochondrial genome. However, due to many technological advances during the past decade, a growing number of studies have explored the power of complete mitochondrial genome sequences...... yielded major progress with regard to both the phylogenetic positions of extinct species, as well as resolving population genetics questions in both extinct and extant species....

  17. Integrated Sensing Using DNA Nanoarchitectures

    Science.gov (United States)

    2014-05-20

    Norton. Thiolated Dendrimers as Multi-Point Binding Headgroups for DNA Immobilization on Gold, Langmuir, (10 2011): 0. doi: 10.1021/la202444s...flat mica surface, the structure is planar (it is conformal, lacking rigidity as a 2 nm thick polymer sheet. The simulated structure is shown in...Morris, John R., and Norton, Michael L.; Thiolated Dendrimers as Multi-Point Binding Headgroups for DNA Immobilization on Gold, Langmuir, 27(20

  18. LET-effects in DNA

    International Nuclear Information System (INIS)

    Kraft, G.; Taucher-Scholz, G.; Heilmann, J.

    1994-11-01

    In this contribution, an introductory view on the physical properties of ions is given and the cellular response to high LET radiation is summarized. Then the measurements of strand break induction of DNA in solution and in intracellular DNA are reported and compared to cell survival. The possibility of changes in the quality of the lesions is discussed and finally the present status of model calculations in comparison to the experiments is given. (orig./HSI)

  19. DNA Binding Hydroxyl Radical Probes

    OpenAIRE

    Tang, Vicky J; Konigsfeld, Katie M; Aguilera, Joe A; Milligan, Jamie R

    2012-01-01

    The hydroxyl radical is the primary mediator of DNA damage by the indirect effect of ionizing radiation. It is a powerful oxidizing agent produced by the radiolysis of water and is responsible for a significant fraction of the DNA damage associated with ionizing radiation. There is therefore an interest in the development of sensitive assays for its detection. The hydroxylation of aromatic groups to produce fluorescent products has been used for this purpose. We have examined four different c...

  20. DNA probe for lactobacillus delbrueckii

    Energy Technology Data Exchange (ETDEWEB)

    Delley, M.; Mollet, B.; Hottinger, H. (Nestle Research Centre, Lausanne (Switzerland))

    1990-06-01

    From a genomic DNA library of Lactobacillus delbrueckii subsp. bulgaricus, a clone was isolated which complements a leucine auxotrophy of an Escherichia coli strain (GE891). Subsequent analysis of the clone indicated that it could serve as a specific DNA probe. Dot-blot hybridizations with over 40 different Lactobacillus strains showed that this clone specifically recognized L. delbrueckii subsp. delbrueckii, bulgaricus, and lactis. The sensitivity of the method was tested by using an {alpha}-{sup 32}P-labeled probe.

  1. DNA probe for lactobacillus delbrueckii

    International Nuclear Information System (INIS)

    Delley, M.; Mollet, B.; Hottinger, H.

    1990-01-01

    From a genomic DNA library of Lactobacillus delbrueckii subsp. bulgaricus, a clone was isolated which complements a leucine auxotrophy of an Escherichia coli strain (GE891). Subsequent analysis of the clone indicated that it could serve as a specific DNA probe. Dot-blot hybridizations with over 40 different Lactobacillus strains showed that this clone specifically recognized L. delbrueckii subsp. delbrueckii, bulgaricus, and lactis. The sensitivity of the method was tested by using an α- 32 P-labeled probe

  2. DNA Probe for Lactobacillus delbrueckii

    OpenAIRE

    Delley, Michèle; Mollet, Beat; Hottinger, Herbert

    1990-01-01

    From a genomic DNA library of Lactobacillus delbrueckii subsp. bulgaricus, a clone was isolated which complements a leucine auxotrophy of an Escherichia coli strain (GE891). Subsequent analysis of the clone indicated that it could serve as a specific DNA probe. Dot-blot hybridizations with over 40 different Lactobacillus strains showed that this clone specifically recognizes L. delbrueckii subsp. delbrueckii, bulgaricus, and lactis. The sensitivity of the method was tested by using an α-32P-l...

  3. Genetic Variation of Giardia lamblia Isolates from Food-handlers in Kashan, Central Iran

    Directory of Open Access Journals (Sweden)

    Hossein HOOSHYAR

    2017-02-01

    Full Text Available Background: Based on genotyping study of human isolates of Giardia lamblia; humans are mainly infected by two assemblages A and B. The present study was carried out to determine the sub-assemblages of G. lamblia isolated from food handlers referred to Kashan health centers, central Iran, 2015.Methods: In this cross-sectional study, 3653 stool samples collected from food-handlers that annually refer to health center for getting a health certification and examined microscopically for G. lamblia cyst. Totally, 44 isolates were selected from 47 Giardia positive samples. Cysts were partially purified by the sucrose density gradient method. After freeze-thaw cycles, genomic DNA was extracted using QIAamp Stool Mini kit. A single step PCR-RFLP method was used to amplify a 458bp fragment at the glutamate dehydrogenase (gdh locus, restriction enzymes BspLI and RsaI were used for distinguish between genotypes A and B and their subgroups.Results: Of 44 isolates, 24(54.5% were sub-assemblage AII, 9(20.5% group B including 7(15.9% BIII and 2(4.6% BIV sub-assemblage and 11(25% isolates showed a mixed pattern of AII and B. Sub-assemblage AI was not detected in this study.Conclusion: The higher rate of sub-assemblage AII demonstrated an anthroponotic origin of the infection so infected food-handlers could directly transmit this protozoan to consumers via contaminated food and water. For finding of pattern of transmission and distribution of Giardia assemblages and sub-assemblage, more studies in human and animal population in different regions are necessary.

  4. DNA extraction from herbarium specimens.

    Science.gov (United States)

    Drábková, Lenka Záveská

    2014-01-01

    With the expansion of molecular techniques, the historical collections have become widely used. Studying plant DNA using modern molecular techniques such as DNA sequencing plays an important role in understanding evolutionary relationships, identification through DNA barcoding, conservation status, and many other aspects of plant biology. Enormous herbarium collections are an important source of material especially for specimens from areas difficult to access or from taxa that are now extinct. The ability to utilize these specimens greatly enhances the research. However, the process of extracting DNA from herbarium specimens is often fraught with difficulty related to such variables as plant chemistry, drying method of the specimen, and chemical treatment of the specimen. Although many methods have been developed for extraction of DNA from herbarium specimens, the most frequently used are modified CTAB and DNeasy Plant Mini Kit protocols. Nine selected protocols in this chapter have been successfully used for high-quality DNA extraction from different kinds of plant herbarium tissues. These methods differ primarily with respect to their requirements for input material (from algae to vascular plants), type of the plant tissue (leaves with incrustations, sclerenchyma strands, mucilaginous tissues, needles, seeds), and further possible applications (PCR-based methods or microsatellites, AFLP).

  5. DNA recognition by synthetic constructs.

    Science.gov (United States)

    Pazos, Elena; Mosquera, Jesús; Vázquez, M Eugenio; Mascareñas, José L

    2011-09-05

    The interaction of transcription factors with specific DNA sites is key for the regulation of gene expression. Despite the availability of a large body of structural data on protein-DNA complexes, we are still far from fully understanding the molecular and biophysical bases underlying such interactions. Therefore, the development of non-natural agents that can reproduce the DNA-recognition properties of natural transcription factors remains a major and challenging goal in chemical biology. In this review we summarize the basics of double-stranded DNA recognition by transcription factors, and describe recent developments in the design and preparation of synthetic DNA binders. We mainly focus on synthetic peptides that have been designed by following the DNA interaction of natural proteins, and we discuss how the tools of organic synthesis can be used to make artificial constructs equipped with functionalities that introduce additional properties to the recognition process, such as sensing and controllability. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Fanconi anemia and DNA repair.

    Science.gov (United States)

    Grompe, M; D'Andrea, A

    2001-10-01

    Fanconi anemia (FA) is an autosomal recessive disorder caused by defects in at least eight distinct genes FANCA, B, C, D1, D2, E, F and G. The clinical phenotype of all FA complementation groups is similar and is characterized by progressive bone marrow failure, cancer proneness and typical birth defects. The principal cellular phenotype is hypersensitivity to DNA damage, particularly interstrand DNA crosslinks. The FA proteins constitute a multiprotein pathway whose precise biochemical function(s) remain unknown. Five of the FA proteins (FANCA, C, E, F and G) interact in a nuclear complex upstream of FANCD2. FANCB and FANCD1 have not yet been cloned, but it is likely that FANCB is part of the nuclear complex and that FANCD1 acts downstream of FANCD2. The FA nuclear complex regulates the mono-ubiquitination of FANCD2 in response to DNA damage, resulting in targeting of this protein into nuclear foci. These foci also contain BRCA1 and other DNA damage response proteins. In male meiosis, FANCD2 also co-localizes with BRCA1 at synaptonemal complexes. Together, these data suggest that the FA pathway functions primarily as a DNA damage response system, although its exact role (direct involvement in DNA repair versus indirect, facilitating role) has not yet been defined.

  7. DNA-scaffolded nanoparticle structures

    Energy Technology Data Exchange (ETDEWEB)

    Hoegberg, Bjoern; Olin, Haakan [Department of Engineering Physics and Mathematics, Mid Sweden University, SE-851 70 Sundsvall, Sweden (Sweden)

    2007-03-15

    DNA self-assembly is a powerful route to the production of very small, complex structures. When used in combination with nanoparticles it is likely to become a key technology in the production of nanoelectronics in the future. Previously, demonstrated nanoparticle assemblies have mainly been periodic and highly symmetric arrays, unsuited as building blocks for any complex circuits. With the invention of DNA-scaffolded origami reported earlier this year (Rothemund P W K 2006 Nature 440 (7082) 297-302), a new route to complex nanostructures using DNA has been opened. Here, we give a short review of the field and present the current status of our experiments were DNA origami is used in conjunction with nanoparticles. Gold nanoparticles are functionalized with thiolated single stranded DNA. Strands that are complementary to the gold particle strands can be positioned on the self-assembled DNA-structure in arbitrary patterns. This property should allow an accurate positioning of the particles by letting them hybridize on the lattice. We report on our recent experiments on this system and discuss open problems and future applications.

  8. DNA-scaffolded nanoparticle structures

    International Nuclear Information System (INIS)

    Hoegberg, Bjoern; Olin, Haakan

    2007-01-01

    DNA self-assembly is a powerful route to the production of very small, complex structures. When used in combination with nanoparticles it is likely to become a key technology in the production of nanoelectronics in the future. Previously, demonstrated nanoparticle assemblies have mainly been periodic and highly symmetric arrays, unsuited as building blocks for any complex circuits. With the invention of DNA-scaffolded origami reported earlier this year (Rothemund P W K 2006 Nature 440 (7082) 297-302), a new route to complex nanostructures using DNA has been opened. Here, we give a short review of the field and present the current status of our experiments were DNA origami is used in conjunction with nanoparticles. Gold nanoparticles are functionalized with thiolated single stranded DNA. Strands that are complementary to the gold particle strands can be positioned on the self-assembled DNA-structure in arbitrary patterns. This property should allow an accurate positioning of the particles by letting them hybridize on the lattice. We report on our recent experiments on this system and discuss open problems and future applications

  9. Mechanical design of DNA nanostructures

    Science.gov (United States)

    Castro, Carlos E.; Su, Hai-Jun; Marras, Alexander E.; Zhou, Lifeng; Johnson, Joshua

    2015-03-01

    Structural DNA nanotechnology is a rapidly emerging field that has demonstrated great potential for applications such as single molecule sensing, drug delivery, and templating molecular components. As the applications of DNA nanotechnology expand, a consideration of their mechanical behavior is becoming essential to understand how these structures will respond to physical interactions. This review considers three major avenues of recent progress in this area: (1) measuring and designing mechanical properties of DNA nanostructures, (2) designing complex nanostructures based on imposed mechanical stresses, and (3) designing and controlling structurally dynamic nanostructures. This work has laid the foundation for mechanically active nanomachines that can generate, transmit, and respond to physical cues in molecular systems.Structural DNA nanotechnology is a rapidly emerging field that has demonstrated great potential for applications such as single molecule sensing, drug delivery, and templating molecular components. As the applications of DNA nanotechnology expand, a consideration of their mechanical behavior is becoming essential to understand how these structures will respond to physical interactions. This review considers three major avenues of recent progress in this area: (1) measuring and designing mechanical properties of DNA nanostructures, (2) designing complex nanostructures based on imposed mechanical stresses, and (3) designing and controlling structurally dynamic nanostructures. This work has laid the foundation for mechanically active nanomachines that can generate, transmit, and respond to physical cues in molecular systems. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr07153k

  10. Satellite DNA: An Evolving Topic.

    Science.gov (United States)

    Garrido-Ramos, Manuel A

    2017-09-18

    Satellite DNA represents one of the most fascinating parts of the repetitive fraction of the eukaryotic genome. Since the discovery of highly repetitive tandem DNA in the 1960s, a lot of literature has extensively covered various topics related to the structure, organization, function, and evolution of such sequences. Today, with the advent of genomic tools, the study of satellite DNA has regained a great interest. Thus, Next-Generation Sequencing (NGS), together with high-throughput in silico analysis of the information contained in NGS reads, has revolutionized the analysis of the repetitive fraction of the eukaryotic genomes. The whole of the historical and current approaches to the topic gives us a broad view of the function and evolution of satellite DNA and its role in chromosomal evolution. Currently, we have extensive information on the molecular, chromosomal, biological, and population factors that affect the evolutionary fate of satellite DNA, knowledge that gives rise to a series of hypotheses that get on well with each other about the origin, spreading, and evolution of satellite DNA. In this paper, I review these hypotheses from a methodological, conceptual, and historical perspective and frame them in the context of chromosomal organization and evolution.

  11. DNA Damage and Pulmonary Hypertension

    Science.gov (United States)

    Ranchoux, Benoît; Meloche, Jolyane; Paulin, Roxane; Boucherat, Olivier; Provencher, Steeve; Bonnet, Sébastien

    2016-01-01

    Pulmonary hypertension (PH) is defined by a mean pulmonary arterial pressure over 25 mmHg at rest and is diagnosed by right heart catheterization. Among the different groups of PH, pulmonary arterial hypertension (PAH) is characterized by a progressive obstruction of distal pulmonary arteries, related to endothelial cell dysfunction and vascular cell proliferation, which leads to an increased pulmonary vascular resistance, right ventricular hypertrophy, and right heart failure. Although the primary trigger of PAH remains unknown, oxidative stress and inflammation have been shown to play a key role in the development and progression of vascular remodeling. These factors are known to increase DNA damage that might favor the emergence of the proliferative and apoptosis-resistant phenotype observed in PAH vascular cells. High levels of DNA damage were reported to occur in PAH lungs and remodeled arteries as well as in animal models of PH. Moreover, recent studies have demonstrated that impaired DNA-response mechanisms may lead to an increased mutagen sensitivity in PAH patients. Finally, PAH was linked with decreased breast cancer 1 protein (BRCA1) and DNA topoisomerase 2-binding protein 1 (TopBP1) expression, both involved in maintaining genome integrity. This review aims to provide an overview of recent evidence of DNA damage and DNA repair deficiency and their implication in PAH pathogenesis. PMID:27338373

  12. DNA testing in homicide investigations.

    Science.gov (United States)

    Prahlow, Joseph A; Cameron, Thomas; Arendt, Alexander; Cornelis, Kenneth; Bontrager, Anthony; Suth, Michael S; Black, Lisa; Tobey, Rebbecca; Pollock, Sharon; Stur, Shawn; Cotter, Kenneth; Gabrielse, Joel

    2017-10-01

    Objectives With the widespread use of DNA testing, police, death investigators, and attorneys need to be aware of the capabilities of this technology. This review provides an overview of scenarios where DNA evidence has played a major role in homicide investigations in order to highlight important educational issues for police, death investigators, forensic pathologists, and attorneys. Methods This was a nonrandom, observational, retrospective study. Data were obtained from the collective files of the authors from casework during a 15-year period, from 2000 through 2014. Results A series of nine scenarios, encompassing 11 deaths, is presented from the standpoint of the police and death investigation, the forensic pathology autopsy performance, the subsequent DNA testing of evidence, and, ultimately, the final adjudication of cases. Details of each case are presented, along with a discussion that focuses on important aspects of sample collection for potential DNA testing, especially at the crime scene and the autopsy. The presentation highlights the diversity of case and evidence types in which DNA testing played a valuable role in the successful prosecution of the case. Conclusions By highlighting homicides where DNA testing contributed to the successful adjudication of cases, police, death investigators, forensic pathologists, and attorneys will be better informed regarding the types of evidence and situations where such testing is of potential value.

  13. DNA damage in plant herbarium tissue.

    NARCIS (Netherlands)

    Staats, M.; Cuenca, A.; Richardson, J.E.; Ginkel, R.V.; Petersen, G.; Seberg, O.; Bakker, F.T.

    2011-01-01

    Dried plant herbarium specimens are potentially a valuable source of DNA. Efforts to obtain genetic information from this source are often hindered by an inability to obtain amplifiable DNA as herbarium DNA is typically highly degraded. DNA post-mortem damage may not only reduce the number of

  14. Alternative end-joining of DNA breaks

    NARCIS (Netherlands)

    Schendel, Robin van

    2016-01-01

    DNA is arguably the most important molecule found in any organism, as it contains all information to perform cellular functions and enables continuity of species. It is continuously exposed to DNA-damaging agents both from endogenous and exogenous sources. To protect DNA against these sources of DNA

  15. Chemically modified DNA : methods and applications

    NARCIS (Netherlands)

    Gjonaj, Lorina

    2013-01-01

    Over het toepassen van DNA in de chemie DNA is een fascinerend biopolymeer dat gemakkelijk gemanipuleerd kan worden. Doel van het onderzoek van Lorina Gjonaj was het ontwikkelen van nieuwe methodologie om DNA chemisch te modificeren. Verschillende functionaliteiten zijn aan DNA gekoppeld. Deze

  16. DNA Fingerprinting in a Forensic Teaching Experiment

    Science.gov (United States)

    Wagoner, Stacy A.; Carlson, Kimberly A.

    2008-01-01

    This article presents an experiment designed to provide students, in a classroom laboratory setting, a hands-on demonstration of the steps used in DNA forensic analysis by performing DNA extraction, DNA fingerprinting, and statistical analysis of the data. This experiment demonstrates how DNA fingerprinting is performed and how long it takes. It…

  17. Fetal DNA: strategies for optimal recovery

    NARCIS (Netherlands)

    Legler, Tobias J.; Heermann, Klaus-Hinrich; Liu, Zhong; Soussan, Aicha Ait; van der Schoot, C. Ellen

    2008-01-01

    For fetal DNA extraction, in principle each DNA extraction method can be used; however, because most methods have been optimized for genomic DNA from leucocytes, we describe here the methods that have been optimized for the extraction of fetal DNA from maternal plasma and validated for this purpose

  18. Extracellular DNA metabolism in Haloferax volcanii

    Directory of Open Access Journals (Sweden)

    Scott eChimileski

    2014-02-01

    Full Text Available Extracellular DNA is found in all environments and is a dynamic component of the micro-bial ecosystem. Microbial cells produce and interact with extracellular DNA through many endogenous mechanisms. Extracellular DNA is processed and internalized for use as genetic information and as a major source of macronutrients, and plays several key roles within prokaryotic biofilms. Hypersaline sites contain some of the highest extracellular DNA con-centrations measured in nature–a potential rich source of carbon, nitrogen and phosphorus for halophilic microorganisms. We conducted DNA growth studies for the halophilic archaeon Haloferax volcanii DS2 and show that this model Halobacteriales strain is capable of using exogenous double-stranded DNA as a nutrient. Further experiments with varying medium composition, DNA concentration and DNA types revealed that DNA is utilized primarily as a phosphorus source, that growth on DNA is concentration-dependent and that DNA isolated from different sources is metabolized selectively, with a bias against highly divergent methylated DNA sources. Additionally, fluorescence microscopy experiments showed that labeled DNA colocalized with Haloferax volcanii cells. The gene Hvo_1477 was also identified using a comparative genomic approach as a factor likely to be involved in extracellular DNA processing at the cell surface, and deletion of Hvo_1477 created an H. volcanii strain deficient in its ability to grow on extracellular DNA. Widespread distribution of Hvo_1477 homologs in archaea suggests metabolism of extracellular DNA may be of broad ecological and physiological relevance in this domain of life.

  19. DNA polymerase beta participates in mitochondrial DNA repair

    DEFF Research Database (Denmark)

    Sykora, P; Kanno, S; Akbari, M

    2017-01-01

    We have detected DNA polymerase beta (Polβ), known as a key nuclear base excision repair (BER) protein, in mitochondrial protein extracts derived from mammalian tissue and cells. Manipulation of the N-terminal sequence affected the amount of Polβ in the mitochondria. Using Polβ fragments, mitocho......We have detected DNA polymerase beta (Polβ), known as a key nuclear base excision repair (BER) protein, in mitochondrial protein extracts derived from mammalian tissue and cells. Manipulation of the N-terminal sequence affected the amount of Polβ in the mitochondria. Using Polβ fragments......, mitochondrial-specific protein partners were identified, with the interactors mainly functioning in DNA maintenance and mitochondrial import. Of particular interest was the identification of the proteins TWINKLE, SSBP1 and TFAM, all of which are mitochondria specific DNA effectors and are known to function...... in the nucleoid. Polβ directly interacted with, and influenced the activity of, the mitochondrial helicase TWINKLE. Human kidney cells with Polβ knock-out (KO) had higher endogenous mtDNA damage. Mitochondrial extracts derived from heterozygous Polβ mouse tissue and KO cells had lower nucleotide incorporation...

  20. Radiation damage to DNA in DNA-protein complexes.

    Science.gov (United States)

    Spotheim-Maurizot, M; Davídková, M

    2011-06-03

    The most aggressive product of water radiolysis, the hydroxyl (OH) radical, is responsible for the indirect effect of ionizing radiations on DNA in solution and aerobic conditions. According to radiolytic footprinting experiments, the resulting strand breaks and base modifications are inhomogeneously distributed along the DNA molecule irradiated free or bound to ligands (polyamines, thiols, proteins). A Monte-Carlo based model of simulation of the reaction of OH radicals with the macromolecules, called RADACK, allows calculating the relative probability of damage of each nucleotide of DNA irradiated alone or in complexes with proteins. RADACK calculations require the knowledge of the three dimensional structure of DNA and its complexes (determined by X-ray crystallography, NMR spectroscopy or molecular modeling). The confrontation of the calculated values with the results of the radiolytic footprinting experiments together with molecular modeling calculations show that: (1) the extent and location of the lesions are strongly dependent on the structure of DNA, which in turns is modulated by the base sequence and by the binding of proteins and (2) the regions in contact with the protein can be protected against the attack by the hydroxyl radicals via masking of the binding site and by scavenging of the radicals. 2011 Elsevier B.V. All rights reserved.