Limited-Resource Preparable Chitosan Magnetic Particles for Extracting Amplification-Ready Nucleic Acid from Complex Biofluid

The Analyst ◽  
2021 ◽  
Author(s):  
Sayantan Tripathy ◽  
Ashish Chalana ◽  
Arunansu Talukdar ◽  
P.V. Rajesh ◽  
Abhijit Saha ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Pathogen Detection ◽  
Magnetic Particles ◽  
Genetic Diseases ◽  
Limited Resource ◽  
Nucleic Acid Amplification Test ◽  
Nucleic Acid Amplification

Extraction and concentration of pure nucleic acid from complex biofluids is the prerequisite for nucleic acid amplification test (NAAT) applications in pathogen detection, biowarfare prevention, and genetic diseases. However, conventional...

scholarly journals Limited-Resource Preparable Chitosan Magnetic Particles for Extracting Amplification-Ready Zeptomole-Range Nucleic Acid from Complex Biofluid

2021 ◽  
Author(s):  
Sayantan Tripathy ◽  
Arunansu Talukdar ◽  
Goutam Pramanik ◽  
P. V. Rajesh ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Magnetic Particles ◽  
Limited Resource ◽  
Nucleic Acid Amplification ◽  
Acid Extraction ◽  
Analytical Sensitivity ◽  
Nucleic Acid Extraction ◽  
Spin Column

<b>Layman Summary: </b>Nucleic acid extraction is a key prerequisite for any nucleic acid amplification test (NAAT) or isothermal NAAT (iNAAT) based molecular diagnosis assays.<b> </b>Existing methods utilizes spin column system for nucleic acid extraction which are unsuitable for limited resource settings. Our work explores two methods for chitosan coated magnetic particle preparation that can be executed within 6 h from commonly available chemicals with nothing but a magnetic stirrer and water bath and doable by a minimally trained person. We will also investigated the compatibility of the extracted nucleic acid with downstream NAATs such as real time LAMP, colorimetric LAMP, and real time PCR. In the process, we established the analytical sensitivity of the overall method.<div><br><div><b>Characterization methods</b>: SEM, XRD, EDX, FT-IR</div><div><br></div><div><b>Bioanalytical methods:</b> Real time LAMP, Colorimetric LAMP, Real time PCR</div></div>

scholarly journals Limited-Resource Preparable Chitosan Magnetic Particles for Extracting Amplification-Ready Zeptomole-Range Nucleic Acid from Complex Biofluid

2021 ◽  
Author(s):  
Sayantan Tripathy ◽  
Arunansu Talukdar ◽  
Goutam Pramanik ◽  
P. V. Rajesh ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Magnetic Particles ◽  
Limited Resource ◽  
Nucleic Acid Amplification ◽  
Acid Extraction ◽  
Analytical Sensitivity ◽  
Nucleic Acid Extraction ◽  
Spin Column

<b>Layman Summary: </b>Nucleic acid extraction is a key prerequisite for any nucleic acid amplification test (NAAT) or isothermal NAAT (iNAAT) based molecular diagnosis assays.<b> </b>Existing methods utilizes spin column system for nucleic acid extraction which are unsuitable for limited resource settings. Our work explores two methods for chitosan coated magnetic particle preparation that can be executed within 6 h from commonly available chemicals with nothing but a magnetic stirrer and water bath and doable by a minimally trained person. We will also investigated the compatibility of the extracted nucleic acid with downstream NAATs such as real time LAMP, colorimetric LAMP, and real time PCR. In the process, we established the analytical sensitivity of the overall method.<div><br><div><b>Characterization methods</b>: SEM, XRD, EDX, FT-IR</div><div><br></div><div><b>Bioanalytical methods:</b> Real time LAMP, Colorimetric LAMP, Real time PCR</div></div>

scholarly journals A Retrospective Cross-Sectional Study of the Utility of Cartridge-Based Nucleic Acid Amplification Test in Diagnosis of Pulmonary and Extrapulmonary Tuberculosis in People Living with HIV in Second Highest HIV Prevalent State in India

2021 ◽  
Author(s):  
Tade Bagbi ◽  
Ningthoukhongjam Reema ◽  
S. Bhagyabati Devi ◽  
Thangjam Gautam Singh ◽  
Mohammad Jaleel ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Cross Sectional Study ◽  
Nucleic Acid Amplification Test ◽  
Nucleic Acid Amplification ◽  
Rifampicin Resistance ◽  
Sputum Smear ◽  
Smear Microscopy ◽  
Sectional Study ◽  
Cross Sectional ◽  
Smear Negative

Abstract Introduction Tuberculosis (TB) in people living with human immunodeficiency virus (PLHIV) is difficult to diagnose due to fewer organisms in sputum and extrapulmonary samples. Sputum culture takes 4 to 8 weeks for growth of the mycobacteria. Delayed treatment for TB in PLHIV leads to increased mortality. This study evaluated cartridge-based nucleic acid amplification test (CBNAAT) as a diagnostic tool for diagnosis of pulmonary TB (PTB) and extrapulmonary TB (EPTB) in PLHIV in the second most HIV prevalent state in India and for comparing its efficacy between Ziehl–Neelsen (ZN) staining sputum smear–positive and sputum smear–negative TB. Methods This cross-sectional study was conducted in RIMS, Imphal, with 167 PLHIV patients, age 15 years or older, having signs and symptoms of TB. Appropriate samples for sputum microscopy and CBNAAT were sent. Conclusion The overall sensitivity of sputum smear for acid-fast bacillus (AFB) was found to be 30.71% and that of CBNAAT was 38.57%. Sensitivity of CBNAAT for sputum smear–positive and sputum smear–negative TB was 100 and 11.3%, respectively. Sensitivity of ZN smear for AFB of EPTB sample was 48.1% and that of CBNAAT was 59.25%. In both PTB and EPTB, CBNAAT showed an increase in diagnosis of microbiologically confirmed PTB cases by 7.8 and 11.1%, respectively, over and above the cases diagnosed by ZN smear microscopy. Rifampicin resistance was detected in five patients. We conclude that CBNAAT is a rapid test with better sensitivity in diagnosis of PTB and EPTB in PLHIV, compared with ZN smear microscopy. It detects rifampicin resistance for multidrug-resistant TB and helps in early treatment intervention.

scholarly journals Nucleic Acid Amplification Test Quantitation as Predictor of Toxin Presence in Clostridium difficile Infection

2017 ◽  
Vol 56 (3) ◽  
Author(s):  
M. J. T. Crobach ◽  
N. Duszenko ◽  
E. M. Terveer ◽  
C. M. Verduin ◽  
E. J. Kuijper
Keyword(s):  
Nucleic Acid ◽  
Clostridium Difficile ◽  
Characteristic Curve ◽  
Nucleic Acid Amplification Test ◽  
Nucleic Acid Amplification ◽  
Toxin A ◽  
Toxin B ◽  
Content Type ◽  
Quantitative Results ◽  
Testing Algorithm

ABSTRACT Multistep algorithmic testing in which a sensitive nucleic acid amplification test (NAAT) is followed by a specific toxin A and toxin B enzyme immunoassay (EIA) is among the most accurate methods for Clostridium difficile infection (CDI) diagnosis. The obvious shortcoming of this approach is that multiple tests must be performed to establish a CDI diagnosis, which may delay treatment. Therefore, we sought to determine whether a preliminary diagnosis could be made on the basis of the quantitative results of the first test in algorithmic testing, which provide a measure of organism burden. To do so, we retrospectively analyzed two large collections of samples ( n = 2,669 and n = 1,718) that were submitted to the laboratories of two Dutch hospitals for CDI testing. Both hospitals apply a two-step testing algorithm in which a NAAT is followed by a toxin A/B EIA. Of all samples, 208 and 113 samples, respectively, tested positive by NAAT. Among these NAAT-positive samples, significantly lower mean quantification cycle ( C q ) values were found for patients whose stool eventually tested positive for toxin, compared with patients who tested negative for toxin (mean C q values of 24.4 versus 30.4 and 26.8 versus 32.2; P < 0.001 for both cohorts). Receiver operating characteristic curve analysis was performed to investigate the ability of C q values to predict toxin status and yielded areas under the curve of 0.826 and 0.854. Using the optimal C q cutoff values, prediction of the eventual toxin A/B EIA results was accurate for 78.9% and 80.5% of samples, respectively. In conclusion, C q values can serve as predictors of toxin status but, due to the suboptimal correlation between the two tests, additional toxin testing is still needed.

scholarly journals Nucleic acid amplification-based pathogen detection in the blood of severe sepsis patients

Critical Care ◽  
2008 ◽  
Vol 12 (Suppl 5) ◽  
pp. P43 ◽  
Author(s):  
Frank Bloos ◽  
Svea Sachse ◽  
Karl-Herrmann Schmidt ◽  
Mark Lehmann ◽  
Roland Schmitz ◽  
...  
Keyword(s):  
Severe Sepsis ◽  
Nucleic Acid ◽  
Pathogen Detection ◽  
Nucleic Acid Amplification

scholarly journals Prospective study to find out the role of gastric aspirate examination by Ziehl-Neelsen staining (ZN staining) and cartridge based nucleic acid amplification test (CB-NAAT) as a diagnostic method in childhood tuberculosis

2018 ◽  
Vol 5 (4) ◽  
pp. 1609
Author(s):  
Akansha Arora ◽  
Anil Jain ◽  
B. S. Karnawat ◽  
Rakesh Kumawat
Keyword(s):  
Nucleic Acid ◽  
Predictive Value ◽  
Nucleic Acid Amplification Test ◽  
Nucleic Acid Amplification ◽  
Mantoux Test ◽  
P Value ◽  
Specific Method ◽  
Gastric Aspirate ◽  
Tuberculosis In Children

Background: Tuberculosis in children has been relatively neglected mainly because clinical diagnosis has low specificity, radiological interpretation is subject to inter-observer variability and the tuberculin skin test is a marker of exposure, not disease. The recent introduction of Cartridge based nucleic acid amplification test has significantly transformed the diagnostics of tuberculosis in adults but its application for Paediatric Tuberculosis is under evaluation. Therefore, authors conducted a study on role of gastric aspirate examination by ZN stain and Cartridge based nucleic acid amplification test in the diagnosis of childhood Tuberculosis.Methods: Authors did a prospective hospital-based study from Nov 2016 to Nov 2017 consisting of 100 randomly selected patients suspected of tuberculosis who had their gastric aspirate tested for CBNAAT and ZN stain for acid fast bacilli (AFB) along with Mantoux test and other routine investigations. Chi square test was used.Results: Culture positive tuberculosis was found in 21 out of 100 children. The sensitivity, specificity, positive predictive value and negative predictive value for CBNAAT were 76.1%, 98.7%, 94.1% and 93.9% and for ZN stain were 47.6%, 98.7%, 90.9% and 87.6% respectively. Positive history of contact (p value 0.0217), reactive Mantoux test (p value < 0.001) and low socioeconomic status were independently associated with a positive CBNAAT result.Conclusions: Analysis of gastric aspirate samples with CBNAAT is a sensitive and specific method for rapid diagnosis of pulmonary tuberculosis in children who cannot produce sputum. Compared with microscopy, CBNAAT offers better sensitivity and its scale up will improve access to tuberculosis diagnostics in children.

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