High‐resolution DNA melt‐curve analysis for cost‐effective mass screening of pairwise species interactions

2013 ◽  
Vol 13 (5) ◽  
pp. 908-917 ◽  
Author(s):  
James K. McCarthy ◽  
Raphael K. Didham ◽  
Eckehard G. Brockerhoff ◽  
Katherine A. Bysterveldt ◽  
Arvind Varsani
Keyword(s):  
High Resolution ◽  
Effective Mass ◽  
Mass Screening ◽  
Species Interactions ◽  
Cost Effective ◽  
Curve Analysis ◽  
Melt Curve Analysis

scholarly journals NESTROFT—A Cost-Effective Mass Screening Tool for the Detection of β-Thalassemia Carrier Status in Anemic Pregnant Women: A Step Toward Reducing the National Disease Burden

2021 ◽  
Author(s):  
Manasi Gosavi ◽  
Ramesh Chavan ◽  
M. B. Bellad
Keyword(s):  
Pregnant Women ◽  
Effective Mass ◽  
Mass Screening ◽  
Predictive Value ◽  
Screening Tool ◽  
Hematological Parameters ◽  
False Negative ◽  
Cost Effective ◽  
Mean Corpuscular Hemoglobin ◽  
Carrier Status

Abstract Introduction β-Thalassemias are inherited hemoglobinopathies commonly encountered in practice. With chances of a promising cure being rare, the prevention of births with this disorder should assume priority, especially in low-resource countries. This can be achieved by the implementation of a mass screening program that is reliable and, at the same time, cost-effective. Objectives This study focuses on the utility of Naked Eye Single Tube Red Cell Osmotic Fragility Test (NESTROFT) as a mass screening tool to detect thalassemia carriers. Hematological parameters that may predict carrier status were also evaluated. Materials and Methods Hemoglobin estimation was performed on all consented pregnant women. If the patient was found to have hemoglobin < 11 g/dL, the blood sample was subjected to other routine hematological tests along with peripheral smear examination. NESTROFT was performed using 0.36% saline solution. Confirmation was done using high-performance liquid chromatography (HPLC). Statistical Analysis Data obtained were tabulated using version 21 of the Statistical Package for Social Sciences. Means, standard deviations, and percentages were used to describe the sample. Chi-square test and Students’ “t” test were used to identify differences between the groups. Results Of 441 pregnant women enrolled, 206 were found to be anemic. Nineteen (9.2%) of the anemic pregnant women were detected to be carriers of hemoglobinopathies. Among the hematological parameters, mean red blood cell count and reticulocyte count were higher, while mean corpuscular hemoglobin concentration was lower in carriers. Also, carriers were more likely to present with microcytic hypochromic anemia. NESTROFT showed a sensitivity of 84.21%, specificity of 96.25%, a positive predictive value of 69.56%, and a negative predictive value of 98.36%. A false-positive result was seen in 3.74% of the tests, while a false negative result was seen in 15.78% of the tests. Conclusions NESTROFT (0.36%) can be used as a simple and cost-effective mass screening tool for the detection of carrier status. This should be followed by confirmation using HPLC or hemoglobin electrophoresis.

scholarly journals Rapid Differentiation of Mycobacterium tuberculosis and M. bovis by High-Resolution Melt Curve Analysis

2010 ◽  
Vol 48 (11) ◽  
pp. 4269-4272 ◽  
Author(s):  
S. Ereqat ◽  
G. K. Bar-Gal ◽  
A. Nasereddin ◽  
K. Azmi ◽  
S. E. Qaddomi ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
High Resolution ◽  
Curve Analysis ◽  
High Resolution Melt ◽  
Melt Curve Analysis

scholarly journals Direct Detection of Rifampin and Isoniazid Resistance in Sputum Samples from Tuberculosis Patients by High-Resolution Melt Curve Analysis

2017 ◽  
Vol 55 (6) ◽  
pp. 1755-1766 ◽  
Author(s):  
Divya Anthwal ◽  
Rakesh Kumar Gupta ◽  
Manpreet Bhalla ◽  
Shinjini Bhatnagar ◽  
Jaya Sivaswami Tyagi ◽  
...  
Keyword(s):  
High Resolution ◽  
Direct Detection ◽  
Melting Curve ◽  
Curve Analysis ◽  
High Resolution Melt ◽  
Melt Curve Analysis ◽  
Content Type ◽  
Plasmid Library ◽  
Mdr Tb ◽  
Smear Negative

ABSTRACT Drug-resistant tuberculosis (TB) is a major threat to TB control worldwide. Globally, only 40% of the 340,000 notified TB patients estimated to have multidrug-resistant-TB (MDR-TB) were detected in 2015. This study was carried out to evaluate the utility of high-resolution melt curve analysis (HRM) for the rapid and direct detection of MDR-TB in Mycobacterium tuberculosis in sputum samples. A reference plasmid library was first generated of the most frequently observed mutations in the resistance-determining regions of rpoB , katG , and an inhA promoter and used as positive controls in HRM. The assay was first validated in 25 MDR M. tuberculosis clinical isolates. The assay was evaluated on DNA isolated from 99 M. tuberculosis culture-positive sputum samples that included 84 smear-negative sputum samples, using DNA sequencing as gold standard. Mutants were discriminated from the wild type by comparing melting-curve patterns with those of control plasmids using HRM software. Rifampin (RIF) and isoniazid (INH) monoresistance were detected in 11 and 21 specimens, respectively, by HRM. Six samples were classified as MDR-TB by sequencing, one of which was missed by HRM. The HRM-RIF, INH- katG , and INH- inhA assays had 89% (95% confidence interval [CI], 52, 100%), 85% (95% CI, 62, 97%), and 100% (95% CI, 74, 100%) sensitivity, respectively, in smear-negative samples, while all assays had 100% sensitivity in smear-positive samples. All assays had 100% specificity. Concordance of 97% to 100% (κ value, 0.9 to 1) was noted between sequencing and HRM. Heteroresistance was observed in 5 of 99 samples by sequencing. In conclusion, the HRM assay was a cost-effective (Indian rupee [INR]400/US$6), rapid, and closed-tube method for the direct detection of MDR-TB in sputum, especially for direct smear-negative cases.

scholarly journals Evaluation of PCR and high-resolution melt curve analysis for differentiation of Salmonella isolates

2017 ◽  
Vol 46 (3) ◽  
pp. 319-331 ◽  
Author(s):  
Mohammad Sadegh Saeidabadi ◽  
Hassan Nili ◽  
Habibollah Dadras ◽  
Hassan Sharifiyazdi ◽  
Joanne Connolly ◽  
...  
Keyword(s):  
High Resolution ◽  
Curve Analysis ◽  
High Resolution Melt ◽  
Melt Curve Analysis

scholarly journals Evaluation of high-resolution melt curve analysis for rapid differentiation of Campylobacter hepaticus from other species in birds

PLoS ONE ◽  
2021 ◽  
Vol 16 (5) ◽  
pp. e0251328
Author(s):  
Petrina Young ◽  
Pol Tarce ◽  
Sadhana Adhikary ◽  
Joanne Connolly ◽  
Tim Crawshaw ◽  
...  
Keyword(s):  
High Resolution ◽  
Dna Sequence ◽  
Egg Production ◽  
Curve Analysis ◽  
Bacterial Disease ◽  
Campylobacter Coli ◽  
High Resolution Melt ◽  
Melt Curve Analysis ◽  
Clinical Specimens ◽  
Bile Samples

Spotty liver disease (SLD) is a bacterial disease of chicken, causing mortalities and reduction in egg production, hence, contributing to economic loss in the poultry industry. The causative agent of SLD has only recently been identified as a novel Campylobacter species, Campylobacter hepaticus. Specific primers were designed from the hsp60 gene of Campylobacter hepaticus and PCR followed by high-resolution melt curve analysis was optimised to detect and differentiate three species of Campylobacter (Campylobacter coli, Campylobacter jejuni and Campylobacter hepaticus). The three Campylobacter species produced a distinct curve profile and was differentiated using HRM curve analysis. The potential of the PCR-HRM curve analysis was shown in the genotyping of 37 Campylobacter isolates from clinical specimens from poultry farms. PCR-HRM curve analysis of DNA extracts from bile samples or cultures from bile samples, were identified as Campylobacter hepaticus and confirmed by DNA sequencing. The DNA sequence analysis of selected samples from each of the three HRM distinctive curves patterns showed that each DNA sequence was associated with a unique melt profile. The potential of the PCR-HRM curve analysis in genotyping of Campylobacter species was also evaluated using faecal specimens from 100 wild birds. The results presented in this study indicate that PCR followed by HRM curve analysis provides a rapid and robust technique for genotyping of Campylobacter species using either bacterial cultures or clinical specimens.

scholarly journals Assessing high-resolution melt curve analysis for accurate detection of gene variants in complex DNA fragments

2009 ◽  
Vol 30 (6) ◽  
pp. 876-883 ◽  
Author(s):  
Elizabeth A. Tindall ◽  
Desiree C. Petersen ◽  
Paula Woodbridge ◽  
Katharina Schipany ◽  
Vanessa M. Hayes
Keyword(s):  
High Resolution ◽  
Curve Analysis ◽  
Gene Variants ◽  
Dna Fragments ◽  
High Resolution Melt ◽  
Melt Curve Analysis ◽  
Accurate Detection
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