scholarly journals Detection of LytA Genes in Streptococcus pneumoniae Isolated from sputum pneumonia patients

Biomedika ◽  
2020 ◽  
Vol 13 (1) ◽  
pp. 23-30
Author(s):  
Mustika Sari Hutabarat ◽  
Firdaus Hamid ◽  
Irawaty Djaharuddin ◽  
Alfian Zainuddin ◽  
Rossana Agus ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
False Negative ◽  
Pneumococcal Infection ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Clinical Samples ◽  
Biochemical Tests ◽  
Negative Results ◽  
False Negative Results ◽  
Polymerase Chain

Streptococcus pneumoniae (pneumococcus) is a Gram-positive facultative anaerobic bacterium that is a major cause of morbidity and mortality worldwide. But the lack of reporting of disease by this bacterium in Indonesia, one of the causes is because the diagnosis of pneumococcal infection is often clinically not typical and conventional methods which are still the standard gold method often give false-negative results. So the purpose of this study was to evaluate the performance of culture and molecular diagnostic methods using the Polymerase Chain Reaction (PCR) technique in detecting Streptococcus pneumoniae in sputum clinical samples using the Autolysin (LytA) gene which is a virulence factor of this bacterium. 57 isolates from 60 samples were confirmed as Streptococcus sp through microscopic identification, culture, and biochemical tests. Then the sensitivity test with an optochin test of 9 (9%) compared the results descriptively with the PCR technique using the Autolysin A (LytA) gene which was obtained more sensitive by 15 (25%).

scholarly journals UTILITY OF ANTIGEN DETECTION TEST AND POLYMERASE CHAIN REACTION IN THE DIFFERENTIATION OF TUBERCULOUS AND NON-TUBERCULOUS MYCOBACTERIA

2017 ◽  
Vol 10 (6) ◽  
pp. 289
Author(s):  
Yogita Singh ◽  
Raji Vasanth ◽  
Shrikala Baliga ◽  
Dhanashree B
Keyword(s):  
Polymerase Chain Reaction ◽  
False Negative ◽  
Diagnostic Methods ◽  
Clinical Samples ◽  
Chain Reaction ◽  
College Hospital ◽  
Medical College ◽  
Negative Results ◽  
Polymerase Chain ◽  
Mycobacterium Spp

Objectives: Cultivation and identification of mycobacteria to species level remains difficult and time-consuming. Hence, easy and rapid diagnostic methods are necessary for the differentiation of Mycobacterium tuberculosis (MTB) from non-tuberculous mycobacteria (NTM). The present study aims to detect and differentiate MTB from NTM isolated from clinical samples by immunochromatographic test (ICT) and polymerase chain reaction (PCR). Methods: Over a period of 1 year, clinical samples (n=496) received from suspected cases of TB, at the Department of Microbiology, Kasturba Medical College Hospital, Mangalore were cultured to isolate Mycobacterium spp. Identification of all the isolates was done by conventional biochemical technique, ICT, and PCR. Results: Among the 496 samples processed, 49 (9.87%) were acid-fast bacilli smear positive and 59 (11.89%) samples showed the growth of Mycobacterium spp. Among these, 10 were rapid growers, 49 were slow-growing mycobacteria, out of which 30 were MTB as identified by conventional biochemical reaction. Out of 59 Mycobacterial isolates subjected to ICT for the detection of MPT 64 antigen, only 28 were identified as MTB. However, all the 30 isolates were correctly identified as MTB by PCR. Conclusion: Hence, PCR is essential for rapid differentiation of non-tuberculous Mycobacterium from MTB. False negative results seen with immunochromatographic MPT 64 antigen assay could be due to mutations within the mpt64 gene. Further studies are necessary to characterize these PCR-positive and immunochromatographic assay negative MTB isolates.

scholarly journals The Use of a Mimic to Detect Polymerase Chain Reaction– Inhibitory Factors in Feces Examined for the Presence of Lawsonia Intracellularis

2003 ◽  
Vol 15 (3) ◽  
pp. 268-273 ◽  
Author(s):  
Magdalena Jacobson ◽  
Stina Englund ◽  
András Ballagi-Pordány
Keyword(s):  
Polymerase Chain Reaction ◽  
False Negative ◽  
Clinical Samples ◽  
Lawsonia Intracellularis ◽  
Chain Reaction ◽  
Fecal Samples ◽  
Wild Boars ◽  
Negative Results ◽  
False Negative Results ◽  
Polymerase Chain

Lawsonia intracellularis is an intracellular organism that causes proliferative enteritis in pigs. This bacterium is difficult to culture, and antemortem demonstration of the microbe is therefore often performed on fecal samples by polymerase chain reaction (PCR). Polymerase chain reaction is sensitive and specific, but inhibitory factors in feces might cause false-negative results. This article describes the construction and use of an internal standard, a mimic. The mimic is amplified by the same primers as those used for L. intracellularis DNA and thus could indicate false-negative results in clinical samples. The amplicon was clearly visible when as few as 10 mimic molecules were added per amplification reaction and when no inhibitors were present. When fecal samples were spiked with the mimic, the detection limit was 102 molecules per PCR. Sixty clinical samples, 20 from wild boars, 20 from growing pigs with diarrhea, and 20 from pigs without diarrhea, were prepared by a boiling procedure and subjected to PCR together with 103 mimic molecules. Nine samples were positive, of which 7 originated from pigs with diarrhea and 2 from pigs without diarrhea. In 14 samples from wild boars, in 8 samples from pigs without diarrhea, and in 3 samples from pigs with diarrhea, neither the mimic nor the target DNA was visible. This indicated the presence of inhibitors in these samples. It is concluded that the mimic can be used as an internal control in the diagnosis of L. intracellularis to indicate inhibition of PCR.

scholarly journals Impact of false negative results of RT-PCR test for SARS-CoV-2 in COVID-19 case count as applied to Mexico

2020 ◽  
Author(s):  
Isaac J. Núñez ◽  
Pablo F. Belaunzarán-Zamudio ◽  
Yanink Caro-Vega
Keyword(s):  
False Negative ◽  
Open Data ◽  
Clinical Samples ◽  
Rt Pcr ◽  
Negative Results ◽  
Case Count ◽  
False Negative Results ◽  
True Number ◽  
High Prevalence ◽  
Polymerase Chain

Underestimation of the number of cases during the COVID-19 pandemic has been a constant concern worldwide. Case confirmation is based on identification of SARS-CoV-2 RNA using real time polymerase chain reaction (RT-PCR) in clinical samples. However, these tests have suboptimal sensitivity, especially during the early and late course of infection. Using open data, we estimated that among 1 343 730 people tested in Mexico since February 27th, there were 838 377 (95% CL 734 605 - 1 057 164) cases, compared with 604 376 considering only positive tests. ICU admissions and deaths were around 16% and 9% higher than reported. Thus, we show that accounting for the sensitivity of SARS-Cov-2 RT-PCR diagnostic tests is a simple way to improve estimations for the true number of COVID-19 cases in tested people, particularly in high-prevalence populations. This could aid to better inform public health measures and reopening policies.

scholarly journals Diagnosis of COVID-19 by Serology in Admitted Patients with Negative RT-PCR Assay

2021 ◽  
Author(s):  
Mitra Rezaei ◽  
Parvaneh Baghaei ◽  
Makan Sadr ◽  
Afshin Moniri ◽  
Abdolreza Babamahmoodi ◽  
...  
Keyword(s):  
Diagnostic Yield ◽  
Serological Test ◽  
False Negative ◽  
Diagnostic Methods ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Serological Tests ◽  
Negative Results ◽  
False Negative Results ◽  
Polymerase Chain

Considering the increasing prevalence and burden of coronavirus disease 2019 (COVID-19) disease and false-negative results in routine reverse transcription-polymerase chain reaction (RT-PCR) tests, additional diagnostic methods are needed to diagnose active cases of this disease. This prospective study was conducted on patients, in whom clinical and radiological symptoms/signs were in favor of COVID-19 while their first PCR test was negative. Later on, a second RT-PCR was performed and serological evaluation was carried out and results were compared with each other. Out of 707 patients who had been referred to the hospital and were clinically and radiologically suspicious of disease, 137 patients with negative RT-PCR tests entered the study. RT-PCR assay became positive for the second time in 45 (32.8%). Anti-COVID-19 IgM and IgG antibodies were positive in 83 (60.6%) and 86 (62.8%) patients, respectively. Finally, it was determined that serological test was diagnostic in 73% of patients and the diagnostic yield of serology was significantly higher after the first week of illness (54.8% in the first week and 88% after that). Taking advantage of both serological tests and RT-PCR helps in diagnosing 83.9% of cases. Based on the present study, the serology may be useful as a complementary test and in parallel to RT-PCR assay for diagnosis of COVID-19 among admitted symptomatic cases.

Detection of Streptococcus pneumoniae in Sputum Samples by Real- Time PCR.

2020 ◽  
Vol 18 ◽  
Author(s):  
Pegah Shakib ◽  
Mohammad Reza Zolfaghari
Keyword(s):  
Streptococcus Pneumoniae ◽  
Real Time ◽  
Real Time Pcr ◽  
False Negative ◽  
Cross Sectional Study ◽  
Laboratory Culture ◽  
Cross Sectional ◽  
Negative Results ◽  
False Negative Results ◽  
Pcr Method

Background: Conventional laboratory culture-based methods for diagnosis of Streptococcus pneumoniae are time-consuming and yield false negative results. Molecular methods including real-time (RT)-PCR rapid methods and conventional PCR due to higher sensitivity and accuracy have been replaced instead traditional culture assay. The aim of the current study was to evaluate lytA gene for detection of Streptococcus pneumoniae in the cerebrospinal fluid of human patients with meningitis using real-time PCR assay. Material and Methods: In this cross-sectional study, a total of 30 clinical specimens were collected from patients in a period from September to December 2018. In order to evaluate the presence of lytA gene, conventional and real-time PCR methods were used without culture. Results: From 30 sputum samples five (16.66%) isolates were identified as S. pneumoniae by lytA PCR and sequencing. Discussion: In this research, an accurate and rapid real-time PCR method was used, which is based on lytA gene for diagnosis of bacteria so that it can be diagnosed. Based on the sequencing results, the sensitivity for detection of lytA gene was 100% (5/5).

scholarly journals Most but not all laboratories can detect the recently emerged Neisseria gonorrhoeae porA mutants – results from the QCMD 2013 N. gonorrhoeae external quality assessment programme

2014 ◽  
Vol 19 (8) ◽  
Author(s):  
D Luijt ◽  
C Di Lorenzo ◽  
A M van Loon ◽  
M Unemo
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Quality Assessment ◽  
External Quality Assessment ◽  
False Negative ◽  
Molecular Diagnostic ◽  
External Quality ◽  
Diagnostic Assays ◽  
Assessment Programme ◽  
Negative Results ◽  
False Negative Results

We describe the results of the Quality Control for Molecular Diagnostics 2013 Neisseria gonorrhoeae external quality assessment programme that included an N. gonorrhoeae strain harbouring an N. meningitidis porA gene which causes false-negative results in molecular diagnostic assays targeting the gonococcal porA pseudogene. Enhanced awareness of the international transmission of such gonococcal strains is needed to avoid false-negative results in both in-house and commercial molecular diagnostic assays used in laboratories worldwide, but particularly in Europe.

Prevalence of porA pseudogene deletion among Neisseria gonorrhoeae isolates referred to the UK’s Gonococcal Resistance to Antimicrobials Surveillance Program

Sexual Health ◽  
2017 ◽  
Vol 14 (4) ◽  
pp. 392 ◽  
Author(s):  
Martina Toby ◽  
Pamela Saunders ◽  
Michelle Cole ◽  
Vlad Grigorjev ◽  
Sarah Alexander ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
False Negative ◽  
Public Health Problem ◽  
Surveillance Program ◽  
Major Public Health Problem ◽  
Negative Results ◽  
False Negative Results ◽  
Confirmatory Tests ◽  
Polymerase Chain ◽  
Low Prevalence

porA pseudogene-negative Neisseria gonorrhoeae isolates produce false-negative results when examined by polymerase chain reaction (PCR) with porA pseudogene targets. In the present study, 533 representative gonococcal isolates received in 2011 via the Gonococcal Resistance to Antimicrobials Surveillance Program were examined to determine the prevalence of porA-negative isolates. Less than 0.4% (2/533) of isolates were found to be reproducibly negative with the porA real-time PCR but were confirmed as N. gonorrhoeae with molecular, biochemical and immunological confirmatory tests. Sequencing revealed both isolates contained the Neisseria meningitidis porA gene. Low prevalence indicates that although these isolates do not present a major public health problem, microbiologists should remain vigilant.

scholarly journals Reducing False Negative PCR Test for COVID-19

2020 ◽  
Vol 9 (3) ◽  
pp. 408-410
Author(s):  
Fatemeh Bahreini ◽  
Rezvan Najafi ◽  
Razieh Amini ◽  
Salman Khazaei ◽  
Saeid Bashirian
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
False Negative ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Creative Commons ◽  
Negative Results ◽  
False Negative Results ◽  
Polymerase Chain ◽  
Pcr Test

As the SARS-CoV-2 (COVID-19) pandemic spreads rapidly, there is need for a diagnostic test with high accuracy to detect infected individuals especially those without symptoms. Real-time polymerase chain reaction (RT-PCR) is a common molecular test for diagnosing SARS-CoV-2. If some factors are not taken into consideration when performing this test, it can have a relatively large number of false negative results. In this article, we discuss important considerations that could lead to false negative test reduction. Key words: • SARS-CoV-2 • COVID-19 • Real time polymerase chain reaction • RT-PCR test • Diagnosis • False negatives • Genetics • Emerging disease   Copyright © 2020 Bahreini et al. Published by Global Health and Education Projects, Inc. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0)which permits unrestricted use, distribution, and reproduction in any medium, provided the original work, first published in this journal, is properly cited.

scholarly journals SARS-CoV-2 RNA detection using pooling of self-collected samples: Simple protocol may foster asymptomatic surveillance

2020 ◽  
Author(s):  
Giselle Ibette Lopez-Lopes ◽  
Rita de Cassia Compagnoli Carmona ◽  
Valéria Oliveira Silva ◽  
Cintia Mayumi Ahagon ◽  
Lincoln Spinazola do Prado ◽  
...  
Keyword(s):  
False Negative ◽  
Clinical Samples ◽  
Rna Detection ◽  
Negative Results ◽  
False Negative Results ◽  
Throat Wash ◽  
Asymptomatic Volunteers ◽  
Simple Protocol ◽  
Antigen Tests ◽  
Public Health Institute

1AbstractBackgroundSurveillance of COVID infection and isolation of infected individuals is one of the available tools to control the spread of SAR-CoV-2. Asymptomatic and pre symptomatic are responsible for substantial transmission. RNA or antigen tests are necessary to identify non-symptomatic individuals. We tested the feasibility of using samples pooling offering different collection alternatives (swab/throat wash/saliva) to volunteers of a public health institute.MethodsWe evaluated pool samples from frozen material from previously tested samples and a prospective collection from asymptomatic volunteers. Some collections were paired for comparison. Pools and some individual samples were extracted with QIAamp Viral RNA Mini Kit (Qiagen, USA) and/or Lucigen Quick Extract DNA extraction solution (BioSearch, USA) and submitted to rtPCR (Allplex, Seegene, Korea).ResultsA total of 240 samples from 130 new collections and 37 samples with known result were evaluated. Pool CT was generally higher than individual samples. Lucigen extraction showed higher CT, including false negative results for samples with high CT at Qiagen extraction. Paired Swab and TW samples showed comparable results. No volunteer from negative pools reported any symptom in the 2-3 days after collection.ConclusionsClinical samples pooling to detect SARS-CoV-2 RNA is feasible and an economical way to test for COVID-19, especially in surveillance strategies targeting more infectiousness, higher viremia individuals. The use of Lucigen reagents show lower sensibility that may lead to false negative results with lower viremia samples. Combining throat wash with saliva may provide and interesting self-collection alternative, but more comparative work is needed.

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