scholarly journals Challenges in laboratory diagnosis of Malaria in a low resource country among cases of acute febrile illness at tertiary care hospital in eastern Nepal: Comparative study on Conventional Vs Molecular approach

2020 ◽  
Author(s):  
Pragyan Dahal ◽  
Basudha Khanal ◽  
Keshav Rai ◽  
Vivek Kattel ◽  
Satish Yadav ◽  
...  
Keyword(s):  
Real Time ◽  
Sensitivity And Specificity ◽  
Malaria Elimination ◽  
Real Time Pcr ◽  
Gold Standard ◽  
Febrile Illness ◽  
Acute Febrile Illness ◽  
Low Resource ◽  
Elimination Programme ◽  
Low Resource Country

Abstract Background For ongoing malaria elimination programme, available methods like microscopy and rapid diagnostic test (RDT) are not able to detect all the cases of malaria in acute febrile illness. These methods are entirely dependent on the course of infection, parasite load and skilled technical resources thus the study was carried out to compare performance of microscopy and RDT which are commonly used in a low resource country along with reference to real-time PCR. Methods Altogether 52 blood samples collected from patients with acute febrile illness were tested by microscopy, RDT and real-time PCR. The results were compared in terms of sensitivity and specificity. Results The test results were as follows: 5.8% positivity by Microscopy, 13.5% positivity by RDT and 27% by real time PCR. Taking into consideration of PCR as a gold standard method microscopy showed 21.4% sensitivity and 100% specificity. Likewise, RDT results revealed 28.6% sensitivity and 92.1% specificity. Conclusion Despite of various diagnostic tools available, microscopy stills remains the gold standard for the diagnosis and RDT is the user friendly to execute the test under the tree, but our preliminary results emphasized the need to implement the test with the higher sensitivity and specificity in context of malaria elimination programme which could be another important opportunity to understand the parasite circulation in case of low endemic region. However, these results should be further verified with the large study cohort in order to document the submicroscopic infection.

scholarly journals Challenges in Laboratory Diagnosis of Malaria in a Low-Resource Country at Tertiary Care in Eastern Nepal: A Comparative Study of Conventional vs. Molecular Methodologies

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Pragyan Dahal ◽  
Basudha Khanal ◽  
Keshav Rai ◽  
Vivek Kattel ◽  
Satish Yadav ◽  
...  
Keyword(s):  
Real Time ◽  
Sensitivity And Specificity ◽  
Malaria Elimination ◽  
Real Time Pcr ◽  
Febrile Illness ◽  
Diagnostic Tools ◽  
Acute Febrile Illness ◽  
Gold Standard Method ◽  
Blood Samples ◽  
Ct Value

For ongoing malaria elimination programmes, available methods such as microscopy and rapid diagnostic tests (RDTs) cannot detect all malaria cases in acute febrile illness. These methods are entirely dependent on the course of infection, parasite load, and skilled technical resources. Our study objectives were to estimate the performance of light microscopy and a RDT as well as real-time PCR for the detection of the Plasmodium parasite. Altogether, 52 blood samples collected from patients with acute febrile illness were tested by microscopy, RDT, and real-time PCR. The results were compared in terms of sensitivity and specificity. Microscopy detected the malaria parasite in 5.8% of the blood samples whereas 13.5% were detected by the RDT and 27% by real-time PCR. Considering real-time PCR as the gold standard method, microscopy had a sensitivity of 21.4% and a specificity of 100%, and the RDT had a sensitivity of 28.6% and a specificity of 92.1%. Microscopy together with the RDT successfully detected malaria positive cases in blood samples of Ct value below 20, but both were unable to detect malaria cases between 26–40 Ct value ranges amplified by real-time PCR. Despite various diagnostic tools being available, microscopy still remains the method of choice for diagnosis, while the RDT is user-friendly when applied at the point of care. However, our preliminary results emphasize the need to implement the test with higher sensitivity and specificity in the context of a malaria elimination programme. Such programmes can be a crucial opportunity to understand the species prevalent in a low-endemic region. However, these results should be further verified with a large cohort study to document the submicroscopic infection.

scholarly journals Development of a TaqMan Array Card for Acute-Febrile-Illness Outbreak Investigation and Surveillance of Emerging Pathogens, Including Ebola Virus

2015 ◽  
Vol 54 (1) ◽  
pp. 49-58 ◽  
Author(s):  
Jie Liu ◽  
Caroline Ochieng ◽  
Steve Wiersma ◽  
Ute Ströher ◽  
Jonathan S. Towner ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Salmonella Enterica ◽  
Ebola Virus ◽  
Febrile Illness ◽  
Fever Virus ◽  
Outbreak Investigation ◽  
Acute Febrile Illness ◽  
Content Type ◽  
Taqman Array

Acute febrile illness (AFI) is associated with substantial morbidity and mortality worldwide, yet an etiologic agent is often not identified. Convalescent-phase serology is impractical, blood culture is slow, and many pathogens are fastidious or impossible to cultivate. We developed a real-time PCR-based TaqMan array card (TAC) that can test six to eight samples within 2.5 h from sample to results and can simultaneously detect 26 AFI-associated organisms, including 15 viruses (chikungunya, Crimean-Congo hemorrhagic fever [CCHF] virus, dengue, Ebola virus, Bundibugyo virus, Sudan virus, hantaviruses [Hantaan and Seoul], hepatitis E, Marburg, Nipah virus, o'nyong-nyong virus, Rift Valley fever virus, West Nile virus, and yellow fever virus), 8 bacteria (Bartonellaspp.,Brucellaspp.,Coxiella burnetii,Leptospiraspp.,Rickettsiaspp.,Salmonella entericaandSalmonella entericaserovar Typhi, andYersinia pestis), and 3 protozoa (Leishmaniaspp.,Plasmodiumspp., andTrypanosoma brucei). Two extrinsic controls (phocine herpesvirus 1 and bacteriophage MS2) were included to ensure extraction and amplification efficiency. Analytical validation was performed on spiked specimens for linearity, intra-assay precision, interassay precision, limit of detection, and specificity. The performance of the card on clinical specimens was evaluated with 1,050 blood samples by comparison to the individual real-time PCR assays, and the TAC exhibited an overall 88% (278/315; 95% confidence interval [CI], 84% to 92%) sensitivity and a 99% (5,261/5,326, 98% to 99%) specificity. This TaqMan array card can be used in field settings as a rapid screen for outbreak investigation or for the surveillance of pathogens, including Ebola virus.

scholarly journals Diagnosing SARS-CoV-2 with Antigen Testing, Transcription-Mediated Amplification and Real-Time PCR

2021 ◽  
Vol 10 (11) ◽  
pp. 2404
Author(s):  
Sascha Dierks ◽  
Oliver Bader ◽  
Julian Schwanbeck ◽  
Uwe Groß ◽  
Michael Weig ◽  
...  
Keyword(s):  
Real Time ◽  
Sensitivity And Specificity ◽  
Real Time Pcr ◽  
Gold Standard ◽  
Latent Class ◽  
Antigen Assay ◽  
Low Prevalence ◽  
Antigen Testing ◽  
Positive Results ◽  
Higher Sensitivity

This study was performed as a head-to-head comparison of the performance characteristics of (1) two SARS-CoV-2-specific rapid antigen assays with real-time PCR as gold standard as well as (2) a fully automated high-throughput transcription-mediated amplification (TMA) assay and real-time PCR in a latent class analysis-based test comparison without a gold standard with several hundred samples in a low prevalence “real world” setting. Recorded sensitivity and specificity of the NADAL and the LumiraDx antigen assays and the Hologic Aptima SARS-CoV-2 TMA assay were 0.1429 (0.0194, 0.5835), 0.7644 (0.7016, 0.8174), and 0.7157 (0, 1) as well as 0.4545 (0.2022, 0.7326), 0.9954 (0.9817, 0.9988), and 0.9997 (not estimable), respectively. Agreement kappa between the positive results of the two antigen-based assays was 0.060 (0.002, 0.167) and 0.659 (0.492, 0.825) for TMA and real-time PCR. Samples with low viral load as indicated by cycle threshold (Ct) values > 30 were generally missed by both antigen assays, while 1:10 pooling suggested higher sensitivity of TMA compared to real-time PCR. In conclusion, both sensitivity and specificity speak in favor of the use of the LumiraDx rather than the NADAL antigen assay, while TMA results are comparably as accurate as PCR, when applied in a low prevalence setting.

scholarly journals Comparative Assessment of Sensitivity and Specificity of Rose Bengal Test and Modified In-house ELISA by using IS711 TaqMan Real Time PCR Assay as a Gold Standard for the Diagnosis of Bovine Brucellosis

2018 ◽  
Vol 11 (2) ◽  
pp. 951-957
Author(s):  
Amira M. Zakaria
Keyword(s):  
Positive Predictive Value ◽  
Real Time ◽  
Sensitivity And Specificity ◽  
Rose Bengal ◽  
Real Time Pcr ◽  
Predictive Value ◽  
Gold Standard ◽  
Bovine Brucellosis ◽  
Rt Pcr ◽  
Rose Bengal Test

Serological approaches such as Rose Bengal Test (RBT) and enzyme linked immunosorbent assay (ELISA) are common tests, however they are generally not sensitive or specific enough for diagnosis of Brucella because of cross-reactivity with different bacterial antigens. The work objected to evaluate the sensitivity and specificity of rose Bengal and modified in-house ELISA using IS711 real time PCR as a gold test to detect Brucella in calves sera. Two hundred and thirty (n=230) blood samples were collected from (2-3) years asymptomatic male calves in two Egyptian abattoirs. Rose Bengal test (RBT) and modified in-house ELISA were applied to determine the seroprevalence of brucellosis in abattoirs animals while quantitative Taqman real-time PCRs (RT-PCR) were implemented for the identification of Brucella genus. The overall prevalence of brucellosis was (53.9 %), (75.2 %) and (79.1 %) as determined by ELISA,RBT and RT- PCR assays respectively. Regarding statistical analysis of the reported data and considering real time PCR the gold standard, the RBT recorded a sensitivity of (79.12%) and a specificity of (39.58 %) with an accuracy of (70.87%). While (83.24%) was reported as positive predictive value and (33.33 %) as a negative predictive value. The sensitivity and specificity of ELISA were (55.49%) and (52.08 %) respectively while the accuracy was (54.78%). Positive predictive value and negative predictive value for ELISA were determined as (81.45%) and (23.58 %) respectively. RBT can be routinely used for diagnosis of Brucella as cost effective , more sensitive and accurate test than ELISA However, real time PCR is highly recommend as gold test for identification and differentiation of bovine brucellosis.

scholarly journals Detection of Streptococcus pneumoniae from culture-negative dried blood spots by real-time PCR in Nigerian children with acute febrile illness

2018 ◽  
Vol 11 (1) ◽  
Author(s):  
Pui-Ying Iroh Tam ◽  
Nelmary Hernandez-Alvarado ◽  
Mark R. Schleiss ◽  
Amy J. Yi ◽  
Fatimah Hassan-Hanga ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Real Time ◽  
Real Time Pcr ◽  
Febrile Illness ◽  
Dried Blood Spots ◽  
Acute Febrile Illness ◽  
Blood Spots ◽  
Nigerian Children ◽  
Culture Negative

scholarly journals Development of a Recombinase Polymerase Amplification Assay for Schistosomiasis Japonica Diagnosis in the Experimental Mice and Domestic Goats

2021 ◽  
Vol 11 ◽  
Author(s):  
Qinghong Guo ◽  
Kerou Zhou ◽  
Cheng Chen ◽  
Yongcheng Yue ◽  
Zheng Shang ◽  
...  
Keyword(s):  
Real Time ◽  
Sensitivity And Specificity ◽  
Real Time Pcr ◽  
Genomic Dna ◽  
Cross Reactivity ◽  
Diagnostic Methods ◽  
Schistosomiasis Japonica ◽  
Plasma Samples ◽  
Low Resource ◽  
Amplification Assay

Although the prevalence of schistosomiasis japonica has declined gradually in China, more accurate and sensitive diagnostic methods are urgently needed for the prevention and control of this disease. Molecular diagnostic methods are advantageous in terms of sensitivity and specificity, but they are time-consuming and require expensive instruments and skilled personnel, which limits their application in low-resource settings. In this study, an isothermal DNA amplification assay and recombinase polymerase amplification (RPA) combined with lateral flow dipstick (LFD) were set up. It was used to detect S. japonicum infections in experimental mice and domestic goats by amplifying a specific DNA fragment of S. japonicum. The lower limit of detection for the LFD-RPA assay was evaluated using dilutions of plasmid containing the target sequence. Cross-reactivity was evaluated using genomic DNA from eight other parasites. The effectiveness of the LFD-RPA assay was verified by assessing 36 positive plasma samples and 36 negative plasma samples from mice. The LFD-RPA assay and real-time PCR were also used to assess 48 schistosomiasis japonica-positive plasma samples and 53 negative plasma samples from goats. The LFD-RPA assay could detect 2.6 femtogram (fg) of S. japonicum target DNA (~39 fg genomic DNA of S. japonicum), only 10-fold less sensitive than real-time PCR assay. There was no cross-reactivity with DNA from the other eight parasites, such as Haemonchus contortus and Spirometra. The whole amplification process could be completed within 15 min at 39°C, and the results can be observed easily using the LFD. The sensitivity and specificity of the LFD-RPA assay were 97.22% (35/36, 95% CI, 85.47%–99.93%) and 100% (36/36, 95% CI, 90.26%–100%) in mice, and 93.75% (45/48, 95% CI, 82.80%–98.69%) and 100% (53/53, 95% CI, 93.28%–100%) in goats. By comparison, the sensitivity and specificity of real-time PCR were 100% (36/36, 95% CI, 90.26%–100%) and 100% (36/36, 95% CI, 90.26%–100%) for mice, and 97.92% (47/48, 95% CI, 88.93%–99.95%) and 100% (53/53, 95% CI, 93.28%–100%) for goats. The LFD-RPA assay exhibits high sensitivity and specificity for the diagnosis of schistosomiasis japonica, and it is an alternative method for diagnosis schistosomiasis japonica in low resource setting.

scholarly journals Evaluation of the New BD Max GC Real-Time PCR Assay, Analytically and Clinically as a Supplementary Test for the BD ProbeTec GC Qx Amplified DNA Assay, for Molecular Detection of Neisseria gonorrhoeae

2015 ◽  
Vol 53 (12) ◽  
pp. 3935-3937 ◽  
Author(s):  
Daniel Golparian ◽  
Stina Boräng ◽  
Martin Sundqvist ◽  
Magnus Unemo
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Real Time ◽  
Sensitivity And Specificity ◽  
Real Time Pcr ◽  
Molecular Detection ◽  
Analytical Sensitivity ◽  
Pcr Assay ◽  
Content Type ◽  
Dna Assay ◽  
Supplementary Test

The new BD Max GC real-time PCR assay showed high clinical and analytical sensitivity and specificity. It can be an effective and accurate supplementary test for the BD ProbeTec GC Qx amplified DNA assay, which had suboptimal specificity, and might also be used for initial detection ofNeisseria gonorrhoeae.

scholarly journals Comparison of the sensitivity and specificity of real-time PCR and in situ hybridization in HPV16 and 18 detection in archival cervical cancer specimens

2012 ◽  
Vol 50 (2) ◽  
pp. 239-247 ◽  
Author(s):  
Beata Biesaga ◽  
Sława Szostek ◽  
Małgorzata Klimek ◽  
Jerzy Jakubowicz ◽  
Joanna Wysocka
Keyword(s):  
Cervical Cancer ◽  
Real Time ◽  
Sensitivity And Specificity ◽  
Real Time Pcr

scholarly journals Diagnostic evaluation of IgM ELISA and IgM Immunofluorescence assay for the diagnosis of Acute Scrub Typhus in central Nepal

2020 ◽  
Author(s):  
Rajendra Gautam ◽  
Keshab Parajuli ◽  
Tshokey Tshokey ◽  
John Stenos ◽  
Jeevan Bahadur Sherchand
Keyword(s):  
Likelihood Ratio ◽  
Predictive Value ◽  
Gold Standard ◽  
Scrub Typhus ◽  
Febrile Illness ◽  
Immunofluorescence Assay ◽  
Acute Febrile Illness ◽  
Central Nepal ◽  
Igm Elisa ◽  
Typhus Fever

Abstract Introduction Scrub typhus is an acute febrile illness caused by the obligate intracellular bacterium,Orientia tsutsugamushi. Given their affordability and ease of use, antibody based diagnostic assays can be important diagnostic tools for early detection of scrub typhus fever in resource poor countries like Nepal. Immunochromatography (ICT) and IgM ELISA are two of the routinely employed antibody based assays for diagnosis of Scrub typhus fever in Nepal, although the recommended gold standard diagnostic test is IgM Immunofluorescence assay (IFA). This study evaluated the InBios Scrub Typhus Detect™ Immunoglobulin M (IgM) ELISA and IgM Immunofluorescence assays in single serum sample at the time of admission. Methodology Study participants (1585 suspected cases), were enrolled based on acute febrile illness with suspected scrub typhus cases in central Nepal from April 2017 to March 2018. Blood sample was collected from the suspected patients of scrub typhus, presenting with acute febrile illness. IgM antibody to Orientia tsusugamushi was detected by using Scrub Typhus Detect™ Kit (In Bios International, USA) and an in-house IgM IFA (Australian Rickettsial Reference Laboratory, Geelong, Australia. The IFA assay was performed with the Gilliam, Karp, Kato strains and O. chuto antigens following the ARRL protocol. Result Statistical analysis of ELISA IgM results when compared to reference test, IgM IFA results demonstrated the following characteristics, sensitivity 84.0% (95%CI: 79.73%-87.68%), specificity 94.82% (95% CI: 93.43%-95.99%), positive likelihood ratio 16.21% (95% CI: 12.71%-20.67%), negative likelihood ratio 0.17% (95% CI: 0.13-0.21%), disease prevalence 22.08% (95% CI: 20.06%-24.21%), positive predictive value 82.12% (95% CI: 78.28-85.42%) and negative predictive value 95.44% (95% CI: 94.27%-96.38%) respectively. Conclusion The study indicated that the IgM ELISA has the sensitivity 84.0% (95% CI: 79.73%-87.68%) and specificity 94.82% (95% CI: 93.43%-95.99%). Although IgM IFA is considered the gold standard test for the diagnosis of scrub typhus cases, it is relatively expensive, requires trained personal and a microscope with fluorescence filters. Scrub typhus IgM ELISA with appropriate OD cut–off values may be the best alternative test and possible viable option for resource limited endemic countries like Nepal.

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