The sensitivity and the specifity of rapid antigen test in streptococcal upper respiratory tract infections.

PubMed

Gurol, Yesim; Akan, Hulya; Izbirak, Guldal; Tekkanat, Zuhal Tazegun; Gunduz, Tehlile Silem; Hayran, Osman; Yilmaz, Gulden

2010-06-01

It is aimed to detect the sensitivity and specificity of rapid antigen detection of group A beta hemolytic streptococci from throat specimen compared with throat culture. The other goal of the study is to help in giving clinical decisions in upper respiratory tract infections according to the age group, by detection of sensitivity and positive predictive values of the rapid tests and throat cultures. Rapid antigen detection and throat culture results for group A beta hemolytic streptococci from outpatients attending to our university hospital between the first of November 2005 and 31st of December 2008 were evaluated retrospectively. Throat samples were obtained by swabs from the throat and transported in the Stuart medium and Quickvue Strep A [Quidel, San Diego, USA] cassette test was applied and for culture, specimen was inoculated on 5% blood sheep agar and identified according to bacitracin and trimethoprim-sulphametaxazole susceptibility from beta hemolytic colonies. During the dates between the first of November 2005 and 31st of December 2008, from 453 patients both rapid antigen detection and throat culture were evaluated. Rapid antigen detection sensitivity and specificity were found to be 64.6% and 96.79%, respectively. The positive predictive value was 80.95% whereas negative predictive value was 92.82%. Kappa index was 0.91. When the results were evaluated according to the age groups, the sensitivity and the positive predictive value of rapid antigen detection in children were 70%, 90.3% and in adults 59.4%, 70.4%. When bacterial infection is concerned to prevent unnecessary antibiotic use, rapid streptococcal antigen test (RSAT) is a reliable method to begin immediate treatment. To get the maximum sensitivity of RSAT, the specimen collection technique used and education of the health care workers is important. While giving clinical decision, it must be taken into consideration that the sensitivity and the positive predictive value of the RSAT is quite lower in adult age group than in pediatric age group. Copyright (c) 2010 Elsevier Ireland Ltd. All rights reserved.

Nested PCR Assay for Eight Pathogens: A Rapid Tool for Diagnosis of Bacterial Meningitis.

PubMed

Bhagchandani, Sharda P; Kubade, Sushant; Nikhare, Priyanka P; Manke, Sonali; Chandak, Nitin H; Kabra, Dinesh; Baheti, Neeraj N; Agrawal, Vijay S; Sarda, Pankaj; Mahajan, Parikshit; Ganjre, Ashish; Purohit, Hemant J; Singh, Lokendra; Taori, Girdhar M; Daginawala, Hatim F; Kashyap, Rajpal S

2016-02-01

Bacterial meningitis is a dreadful infectious disease with a high mortality and morbidity if remained undiagnosed. Traditional diagnostic methods for bacterial meningitis pose a challenge in accurate identification of pathogen, making prognosis difficult. The present study is therefore aimed to design and evaluate a specific and sensitive nested 16S rDNA genus-based polymerase chain reaction (PCR) assay using clinical cerebrospinal fluid (CSF) for rapid diagnosis of eight pathogens causing the disease. The present work was dedicated to development of an in-house genus specific 16S rDNA nested PCR covering pathogens of eight genera responsible for causing bacterial meningitis using newly designed as well as literature based primers for respective genus. A total 150 suspected meningitis CSF obtained from the patients admitted to Central India Institute of Medical Sciences (CIIMS), India during the period from August 2011 to May 2014, were used to evaluate clinical sensitivity and clinical specificity of optimized PCR assays. The analytical sensitivity and specificity of our newly designed genus-specific 16S rDNA PCR were found to be ≥92%. With such a high sensitivity and specificity, our in-house nested PCR was able to give 100% sensitivity in clinically confirmed positive cases and 100% specificity in clinically confirmed negative cases indicating its applicability in clinical diagnosis. Our in-house nested PCR system therefore can diagnose the accurate pathogen causing bacterial meningitis and therefore be useful in selecting a specific treatment line to minimize morbidity. Results are obtained within 24 h and high sensitivity makes this nested PCR assay a rapid and accurate diagnostic tool compared to traditional culture-based methods.

Challenges in Routine Implementation and Quality Control of Rapid Diagnostic Tests for Malaria–Rufiji District, Tanzania

PubMed Central

McMorrow, Meredith L.; Masanja, M. Irene; Abdulla, Salim M. K.; Kahigwa, Elizeus; Kachur, S. Patrick

2018-01-01

Rapid diagnostic tests (RDTs) represent an alternative to microscopy for malaria diagnosis and have shown high sensitivity and specificity in a variety of study settings. Current World Health Organization (WHO) guidelines for quality control of RDTs provide detailed instructions on pre-field testing, but offer little guidance for quality assurance once RDTs are deployed in health facilities. From September 2006 to April 2007, we introduced a histidine-rich protein II (HRP2)-based RDT (Paracheck) for suspected malaria cases five years of age and older in nine health facilities in Rufiji District, Tanzania, to assess sensitivity and specificity of RDTs in routine use at rural health facilities. Thick blood smears were collected for all patients tested with RDTs and stained and read by laboratory personnel in each facility. Thick smears were subsequently reviewed by a reference microscopist to determine RDT sensitivity and specificity. In all nine health facilities, there were significant problems with the quality of staining and microscopy. Sensitivity and specificity of RDTs were difficult to assess given the poor quality of routine blood smear staining. Mean operational sensitivity of RDTs based on reference microscopy was 64.8%, but varied greatly by health facility, range 18.8–85.9%. Sensitivity of RDTs increased with increasing parasite density. Specificity remained high at 87.8% despite relatively poor slide quality. Institution of quality control of RDTs based on poor quality blood smear staining may impede reliable measurement of sensitivity and specificity and undermine confidence in the new diagnostic. There is an urgent need for the development of alternative quality control procedures for rapid diagnostic tests that can be performed at the facility level. PMID:18784230

Performance of a HRP-2/pLDH based rapid diagnostic test at the Bangladesh-India-Myanmar border areas for diagnosis of clinical malaria.

PubMed

Elahi, Rubayet; Mohon, Abu Naser; Khan, Wasif A; Haque, Rashidul; Alam, Mohammad Shafiul

2013-10-30

The rapid diagnostic test (RDT) has been adopted in contemporary malaria control and management programmes around the world as it represents a fast and apt alternative for malaria diagnosis in a resource-limited setting. This study assessed the performance of a HRP-2/pLDH based RDT (Parascreen® Pan/Pf) in a laboratory setting utilizing clinical samples obtained from the field. Whole blood samples were obtained from febrile patients referred for malaria diagnosis by clinicians from two different Upazila Health Complexes (UHCs) located near the Bangladesh-India and Bangladesh-Myanmar border where malaria is endemic. RDT was performed on archived samples and sensitivity and specificity evaluated with expert microscopy (EM) and quantitative PCR (qPCR). A total of 327 clinical samples were made available for the study, of which 153 were Plasmodium falciparum-positive and 54 were Plasmodium vivax-positive. In comparison with EM, for P. falciparum malaria, the RDT had sensitivity: 96.0% (95% CI, 91.2-98.3) and specificity: 98.2% (95% CI, 94.6-99.5) and for P. vivax, sensitivity: 90.7% (95% CI, 78.9-96.5) and specificity: 98.9% (95% CI, 96.5-99.7). Comparison with qPCR showed, for P. falciparum malaria, sensitivity: 95.4% (95% CI, 90.5-98.0) and specificity: 98.8% (95% CI, 95.4-99.7) and for P. vivax malaria, sensitivity: 89.0% (95% CI,77.0-95.4) and specificity: 98.8% (95% CI, 96.5-99.7). Sensitivity varied according to different parasitaemia for falciparum and vivax malaria diagnosis. Parascreen® Pan/Pf Rapid test for malaria showed acceptable sensitivity and specificity in border belt endemic areas of Bangladesh when compared with EM and qPCR.

An evaluation of a rapid real time polymerase chain reaction assay for detection of group B streptococcus as part of a neonatal group B streptococcus prevention strategy.

PubMed

Money, Deborah; Dobson, Simon; Cole, Lesley; Karacabeyli, Eda; Blondel-Hill, Edith; Milner, Ruth; Thomas, Eva

2008-09-01

To evaluate the sensitivity, specificity, and feasibility of a rapid real-time polymerase chain reaction (PCR) test for group B streptococcus (GBS) completed during labour, compared with the standard culture test performed at 35 to 37 weeks' gestation. Women presenting to the maternity unit for term vaginal delivery had two vaginal/rectal samples collected. One swab was tested using a rapid PCR method (IDI-Strep B, Infectio Diagnostic [IDI] Inc., Sainte-Foy QC ), and the other was cultured after enrichment (intrapartum culture). Comparisons were made between these results and those of a culture-based screen at 35 to 37 weeks' gestation. Of the 190 women enrolled, 85% had results of the standard screen at 35 to 37 weeks available for comparison. The sensitivity and specificity of the standard 35- to 37-week screen were 84.3% (95% confidence interval [CI], 71.4-93.0) and 93.2% (95% CI 86.5-97.2) respectively, whereas the sensitivity and specificity of the rapid PCR were 90.7% (95% CI 79.7-96.9) and 97.6% (95% CI 93.1-99.5), respectively. The median reporting time for the rapid PCR test was 99 minutes (range 50-255). Results were available more than four hours before delivery in 81% of cases. In this Canadian centre, a rapid PCR test done at the time of labour (IDI-Strep B) demonstrated high sensitivity and specificity, comparable to the 35- to 37-week screen. The time to reporting results was acceptably short, allowing for timely administration of intrapartum prophylactic antibiotics.

Development of a highly sensitive lateral immunochromatographic assay for rapid detection of Vibrio parahaemolyticus.

PubMed

Liu, Xinfeng; Guan, Yuyao; Cheng, Shiliang; Huang, Yidan; Yan, Qin; Zhang, Jun; Huang, Guanjun; Zheng, Jian; Liu, Tianqiang

2016-12-01

Vibrio parahaemolyticus is widely present in brackish water all over the world, causing infections in certain aquatic animals. It is also a foodborne pathogen that causes diarrhea in humans. The aim of this study is to develop an immunochromatographic lateral flow assay (LFA) for rapid detection of V. parahaemolyticus in both aquatic products and human feces of diarrheal patients. Two monoclonal antibody (MAb) pairs, GA1a-IC9 and IC9-KB4c, were developed and proven to be highly specific and sensitive to V. parahaemolyticus. Based on the two MAb pairs, two types of LFA strips were prepared. Their testing limits for V. parahaemolyticus culture were both 1.2×10 3 CFU/ml. The diagnostic sensitivities and specificities were both 100% for the 32 tested microbial species, including 6 Vibrio species. Subsequently, the LFA strips were used to test Whiteleg shrimps and human feces. The type II strip showed a higher diagnostic sensitivity. Its sensitivity and specificity for hepatopancreas and fecal samples from 13 Whiteleg shrimps and fecal samples from 146 human diarrheal patients were all 100%. In conclusion, our homemade type II LFA is a very promising testing device for rapid and convenient detection of V. parahaemolyticus infection not only in aquatic animals, but also in human diarrheal patients. This sensitive immunochromtographic LFA allows rapid detection of V. parahaemolyticus without requirement of culture enrichment. Copyright © 2016. Published by Elsevier B.V.

Correlation between the immunological condition and the results of immunoenzymatic tests in diagnosing infectious mononucleosis.

PubMed

Tamaro, Giorgio; Donato, Michela; Princi, Tanja; Parco, Sergio

2009-04-01

A symptom-based diagnosis of infectious mononucleosis is not sufficiently accurate, since some clinical symptoms of infectious mononucleosis are also detected in other virally induced diseases. Moreover, not all patients suffering from infectious mononucleosis show circulating atypical lymphocytes, which are considered characteristic of this disease. Therefore, when this disorder is suspected, serum analyses are carried out to detect the presence of certain immunoglobulins associated with infectious mononucleosis in the patient's blood. The aim of this study was to evaluate the sensitivity and the specificity of a rapid test detecting heterophil antibodies in diagnosing infectious mononucleosis in a paediatric population. We considered 163 paediatric patients with suspected infectious mononucleosis and we tested their serums to detect heterophil antibodies (using an inexpensive and rapid test) and specific immunoglobulins directed against Epstein-Barr virus (EBV) (these assays are known to be characterized by high sensitivity and specificity, but are more expensive and time-consuming). By comparing the results of the rapid test with those of the other assays, we found that the sensitivity of the first test was 61.8%, whereas its specificity was sufficiently high (about 90%). We show that, in paediatric patients, the detection of heterophil antibodies is not a very sensitive test, therefore the determination of immunoglobulins against specific antigens of EBV is recommended.

Rapid and sensitive detection of human astrovirus in water samples by loop-mediated isothermal amplification with hydroxynaphthol blue dye

PubMed Central

2014-01-01

Background The aim of this paper was to develop a reverse transcription loop-mediated isothermal amplification (RT-LAMP) method for rapid, sensitive and inexpensive detection of astrovirus. Results The detection limit of LAMP using in vitro RNA transcripts was 3.6×10 copies·μL-1, which is as sensitive as the presently used PCR assays. However, the LAMP products could be identified as different colors with the naked eye following staining with hydroxynaphthol blue dye (HNB). No cross-reactivity with other gastroenteric viruses (rotavirus and norovirus) was observed, indicating the relatively high specificity of LAMP. The RT-LAMP method with HNB was used to effectively detect astrovirus in reclaimed water samples. Conclusions The LAMP technique described in this study is a cheap, sensitive, specific and rapid method for the detection of astrovirus. The RT-LAMP method can be simply applied for the specific detection of astrovirus and has the potential to be utilized in the field as a screening test. PMID:24524254

Rapid and sensitive detection of Yersinia pestis using amplification of plague diagnostic bacteriophages monitored by real-time PCR.

PubMed

Sergueev, Kirill V; He, Yunxiu; Borschel, Richard H; Nikolich, Mikeljon P; Filippov, Andrey A

2010-06-28

Yersinia pestis, the agent of plague, has caused many millions of human deaths and still poses a serious threat to global public health. Timely and reliable detection of such a dangerous pathogen is of critical importance. Lysis by specific bacteriophages remains an essential method of Y. pestis detection and plague diagnostics. The objective of this work was to develop an alternative to conventional phage lysis tests--a rapid and highly sensitive method of indirect detection of live Y. pestis cells based on quantitative real-time PCR (qPCR) monitoring of amplification of reporter Y. pestis-specific bacteriophages. Plague diagnostic phages phiA1122 and L-413C were shown to be highly effective diagnostic tools for the detection and identification of Y. pestis by using qPCR with primers specific for phage DNA. The template DNA extraction step that usually precedes qPCR was omitted. phiA1122-specific qPCR enabled the detection of an initial bacterial concentration of 10(3) CFU/ml (equivalent to as few as one Y. pestis cell per 1-microl sample) in four hours. L-413C-mediated detection of Y. pestis was less sensitive (up to 100 bacteria per sample) but more specific, and thus we propose parallel qPCR for the two phages as a rapid and reliable method of Y. pestis identification. Importantly, phiA1122 propagated in simulated clinical blood specimens containing EDTA and its titer rise was detected by both a standard plating test and qPCR. Thus, we developed a novel assay for detection and identification of Y. pestis using amplification of specific phages monitored by qPCR. The method is simple, rapid, highly sensitive, and specific and allows the detection of only live bacteria.

Development of one-step quantitative reverse transcription PCR for the rapid detection of flaviviruses.

PubMed

Patel, Pranav; Landt, Olfert; Kaiser, Marco; Faye, Oumar; Koppe, Tanja; Lass, Ulrich; Sall, Amadou A; Niedrig, Matthias

2013-02-14

The genus Flavivirus includes several pathogenic agents that cause severe illness in humans. Re-emergence of West Nile virus in Europe and continuous spread of certain flaviviruses such as dengue, yellow fever and Japanese encephalitis viruses represent a global danger to public health. Therefore, a rapid and accurate molecular method is required for diagnostics and epidemiological surveillance of flaviviruses. A Pan-Flavi quantitative RT-PCR assay using a Locked-Nucleic Acid probe targeting the flavivirus NS5 gene was developed and optimized to detect a wide range of flaviviruses simultaneously. The specificity and sensitivity of the Pan-Flavi assay were tested using RNA of different flaviviruses and non-flaviviruses. Furthermore, the assay was compared directly to flavivirus species-specific assays for the ability to detect flaviviruses sensitively. Two degenerate primers and one Locked-Nucleic Acids probe were designed to amplify most of the flaviviruses. To increase the specificity and fluorescence signal of the Pan-Flavi assay for detection of yellow fever virus and dengue virus 4, additional primers and probes were included. Viral RNA of thirty different flaviviruses was detected, verifying the broad range specificity. The testing of this assay was successful, using standard plasmid and RNA dilutions of yellow fever virus vaccine strain, dengue virus 1 and tick-borne encephalitis virus, with a sensitivity limit of 10-100 genome copies/reaction. Also comparatively good results were achieved for detecting different flaviviruses by the Pan-Flavi assay when compared to the flavivirus species-specific assays. The assay is rapid, broad-range flavivirus-specific and highly sensitive making it a valuable tool for rapid detection of flaviviruses in livestock samples, epidemiological studies or as useful complement to single flavivirus-specific assays for clinical diagnosis.

Combining magnetic nanoparticle with biotinylated nanobodies for rapid and sensitive detection of influenza H3N2

PubMed Central

2014-01-01

Our objective is to develop a rapid and sensitive assay based on magnetic beads to detect the concentration of influenza H3N2. The possibility of using variable domain heavy-chain antibodies (nanobody) as diagnostic tools for influenza H3N2 was investigated. A healthy camel was immunized with inactivated influenza H3N2. A nanobody library of 8 × 108 clones was constructed and phage displayed. After three successive biopanning steps, H3N2-specific nanobodies were successfully isolated, expressed in Escherichia coli, and purified. Sequence analysis of the nanobodies revealed that we possessed four classes of nanobodies against H3N2. Two nanobodies were further used to prepare our rapid diagnostic kit. Biotinylated nanobody was effectively immobilized onto the surface of streptavidin magnetic beads. The modified magnetic beads with nanobody capture specifically influenza H3N2 and can still be recognized by nanobodies conjugated to horseradish peroxidase (HRP) conjugates. Under optimized conditions, the present immunoassay exhibited a relatively high sensitive detection with a limit of 50 ng/mL. In conclusion, by combining magnetic beads with specific nanobodies, this assay provides a promising influenza detection assay to develop a potential rapid, sensitive, and low-cost diagnostic tool to screen for influenza infections. PMID:25328501

Combining magnetic nanoparticle with biotinylated nanobodies for rapid and sensitive detection of influenza H3N2

NASA Astrophysics Data System (ADS)

Zhu, Min; Hu, Yonghong; Li, Guirong; Ou, Weijun; Mao, Panyong; Xin, Shaojie; Wan, Yakun

2014-09-01

Our objective is to develop a rapid and sensitive assay based on magnetic beads to detect the concentration of influenza H3N2. The possibility of using variable domain heavy-chain antibodies (nanobody) as diagnostic tools for influenza H3N2 was investigated. A healthy camel was immunized with inactivated influenza H3N2. A nanobody library of 8 × 108 clones was constructed and phage displayed. After three successive biopanning steps, H3N2-specific nanobodies were successfully isolated, expressed in Escherichia coli, and purified. Sequence analysis of the nanobodies revealed that we possessed four classes of nanobodies against H3N2. Two nanobodies were further used to prepare our rapid diagnostic kit. Biotinylated nanobody was effectively immobilized onto the surface of streptavidin magnetic beads. The modified magnetic beads with nanobody capture specifically influenza H3N2 and can still be recognized by nanobodies conjugated to horseradish peroxidase (HRP) conjugates. Under optimized conditions, the present immunoassay exhibited a relatively high sensitive detection with a limit of 50 ng/mL. In conclusion, by combining magnetic beads with specific nanobodies, this assay provides a promising influenza detection assay to develop a potential rapid, sensitive, and low-cost diagnostic tool to screen for influenza infections.

Multiplexed lateral flow microarray assay for detection of citrus pathogens Xylella fastidiosa and Xanthomonas axonopodis pv citri

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cary,; Bruce, R; Stubben, Christopher J

The invention provides highly sensitive and specific assays for the major citrus pathogens Xylella fastidiosa and Xanthomonas axonopodis, including a field deployable multiplexed assay capable of rapidly assaying for both pathogens simultaneously. The assays are directed at particular gene targets derived from pathogenic strains that specifically cause the major citrus diseases of citrus variegated chlorosis (Xylella fastidiosa 9a5c) and citrus canker (Xanthomonas axonopodis pv citri). The citrus pathogen assays of the invention offer femtomole sensitivity, excellent linear dynamic range, and rapid and specific detection.

Aptamer-Immobilized Surface Plasmon Resonance Biosensor for Rapid and Sensitive Determination of Virulence Determinant.

PubMed

Song, Myeong-Sub; Sekhon, Simranjeet Singh; Shin, Woo-Ri; Rhee, Sung-Keun; Ko, Jung Ho; Kim, Sang Yong; Min, Jiho; Ahn, Ji-Young; Kim, Yang-Hoon

2018-05-01

Shigella sonnei isolate invasion plasmid antigen protein, IpaH, was successfully expressed in recombinant overexpression bacterial system. The soluble expression IpaH was enhanced with molecular chaperon co-expressed environment. Specific aptamer IpaH17 was isolated through the SELEX process and showed fM binding affinity. IpaH17-SPR biosensor platform was involved to verify the binding sensitivity and specificity. The IpaH concentration dependent IpaH17-SPR sensor response was highly linear with a linear regression constant of 99.4% in the range between 0 and 100 ng/mL. In addition, S. sonnei revealed the specific RU value and detected in a real-time manner within 1 hour. Our study indicated that IpaH17-SPR sensor can allow for rapid, sensitive and specific determination of Shigella sonnei virulent factor.

Lipopolysaccharide Specific Immunochromatography Based Lateral Flow Assay for Serogroup Specific Diagnosis of Leptospirosis in India

PubMed Central

Vanithamani, Shanmugam; Shanmughapriya, Santhanam; Narayanan, Ramasamy; Raja, Veerapandian; Kanagavel, Murugesan; Sivasankari, Karikalacholan; Natarajaseenivasan, Kalimuthusamy

2015-01-01

Background Leptospirosis is a re-emerging infectious disease that is under-recognized due to low-sensitivity and cumbersome serological tests. MAT is the gold standard test and it is the only serogroup specific test used till date. Rapid reliable alternative serogroup specific tests are needed for surveillance studies to identify locally circulating serogroups in the study area. Methods/Principal Findings In the present investigation the serological specificity of leptospiral lipopolysaccharides (LPS) was evaluated by enzyme linked immunosorbent assay (ELISA), dot blot assay and rapid immunochromatography based lateral flow assay (ICG-LFA). Sera samples from 120 MAT positive cases, 174 cases with febrile illness other than leptospirosis, and 121 seronegative healthy controls were evaluated for the diagnostic sensitivity and specificity of the developed assays. LPS was extracted from five locally predominant circulating serogroups including: Australis (27.5%), Autumnalis (11.7%), Ballum (25.8%), Grippotyphosa (12.5%), Pomona (10%) and were used as antigens in the diagnostics to detect IgM antibodies in patients’ sera. The sensitivity observed by IgM ELISA and dot blot assay using various leptospiral LPS was >90% for homologous sera. Except for Ballum LPS, no other LPS showed cross-reactivity to heterologous sera. An attempt was made to develop LPS based ICG-LFA for rapid and sensitive serogroup specific diagnostics of leptospirosis. The developed ICG-LFA showed sensitivity in the range between 93 and 100% for homologous sera. The Wilcoxon analysis showed LPS based ICG-LFA did not differ significantly from the gold standard MAT (P>0.05). Conclusion The application of single array of LPS for serogroup specific diagnosis is first of its kind. The developed assay could potentially be evaluated and employed for as MAT alternative. PMID:26340095

Loop-mediated isothermal amplification for detection of Staphylococcus aureus in dairy cow suffering from mastitis.

PubMed

Tie, Zhang; Chunguang, Wang; Xiaoyuan, Wei; Xinghua, Zhao; Xiuhui, Zhong

2012-01-01

To develop a rapid detection method of Staphylococcus aureus using loop-mediated isothermal amplification (LAMP), four specific primers were designed according to six distinct sequences of the nuc gene. In addition, the specificity and sensitivity of LAMP were verified and compared with those of PCR. Results showed that the LAMP reaction was completed within 45 min at 62.5°C, and ladder bands were appeared in LAMP products analyzed by gel electrophoresis. After adding 1x SYBR Green l, the positive reaction tube showed green color and the negative reaction tube remained orange, indicating that the LAMP has high specificity. The minimal detectable concentration of LAMP was 1 × 10² CFU/mL and that of PCR was 1 × 10⁴ CFU/mL, indicating that the LAMP was 100 times more sensitive than the PCR. The LAMP method for detection of Staphylococcus aureus has many advantages, such as simple operation, high sensitivity, high specificity, and rapid analysis. Therefore, this method is more suitable for the rapid on-site detection of Staphylococcus aureus.

Laboratory evaluation on the sensitivity and specificity of a novel and rapid detection method for malaria diagnosis based on magneto-optical technology (MOT).

PubMed

Mens, Petra F; Matelon, Raphael J; Nour, Bakri Y M; Newman, Dave M; Schallig, Henk D F H

2010-07-19

This study describes the laboratory evaluation of a novel diagnostic platform for malaria. The Magneto Optical Test (MOT) is based on the bio-physical detection of haemozoin in clinical samples. Having an assay time of around one minute, it offers the potential of high throughput screening. Blood samples of confirmed malaria patients from different regions of Africa, patients with other diseases and healthy non-endemic controls were used in the present study. The samples were analysed with two reference tests, i.e. an histidine rich protein-2 based rapid diagnostic test (RDT) and a conventional Pan-Plasmodium PCR, and the MOT as index test. Data were entered in 2 x 2 tables and analysed for sensitivity and specificity. The agreement between microscopy, RDT and PCR and the MOT assay was determined by calculating Kappa values with a 95% confidence interval. The observed sensitivity/specificity of the MOT test in comparison with clinical description, RDT or PCR ranged from 77.2 - 78.8% (sensitivity) and from 72.5 - 74.6% (specificity). In general, the agreement between MOT and the other assays is around 0.5 indicating a moderate agreement between the reference and the index test. However, when RDT and PCR are compared to each other, an almost perfect agreement can be observed (k = 0.97) with a sensitivity and specificity of >95%. Although MOT sensitivity and specificity are currently not yet at a competing level compared to other diagnostic test, such as PCR and RDTs, it has a potential to rapidly screen patients for malaria in endemic as well as non-endemic countries.

Comparative Evaluation of Four Phenotypic Tests for Detection of Carbapenemase-Producing Gram-Negative Bacteria

PubMed Central

Noël, Audrey; Berhin, Catherine; Hoebeke, Martin; Bouchahrouf, Warda; Yunus, Sami; Bogaerts, Pierre; Glupczynski, Youri

2016-01-01

ABSTRACT Four screening assays aimed for rapid detection of carbapenemase production from Gram-negative bacterial isolates, i.e., the Neo-Rapid Carb kit (Rosco Diagnostica A/S), the Rapidec Carba NP test (bioMérieux SA), the β Carba test (Bio-Rad Laboratories N.V.), and a homemade electrochemical assay (BYG Carba test) were evaluated against a panel comprising 328 clinical isolates (Enterobacteriaceae [n = 198] and nonfermentative Gram-negative bacilli [n = 130]) with previously characterized resistance mechanisms to carbapenems. Among Enterobacteriaceae isolates, the BYG Carba test and the β Carba test showed excellent sensitivities (respectively, 100% and 97.3%) and specificities (respectively, 98.9% and 97.7%). The two other assays yielded poorer performances with sensitivity and specificity of 91.9% and 83.9% for the Rapidec Carba NP test and of 89.2% and 89.7% for the Neo-Rapid Carb kit, respectively. Among Pseudomonas spp., sensitivities and specificities ranged, respectively, from 87.3% to 92.7% and from 88.2% to 94.1%. Finally, all tests performed poorly against Acinetobacter spp., with sensitivities and specificities, respectively, ranging from 27.3% to 75.8% and from 75 to 100%. Among commercially available assays, the β Carba test appeared to be the most convenient for routine use and showed the best overall performances, especially against OXA-48-like producers. The excellent performance of the BYG Carba test against Enterobacteriaceae was confirmed (100% sensitivity and 98.9% specificity). PMID:27927915

Study and evaluation of Wondfo rapid diagnostic kit based on nano-gold immunochromatography assay for diagnosis of Plasmodium falciparum.

PubMed

Peng, Yunping; Wu, Junlin; Wang, Jihua; Li, Wenmei; Yu, Shujuan

2012-04-01

Malaria has been recognized as a human disease for thousands of years and remains one of the most common diseases affecting humans worldwide. Therefore, a method for rapidly detecting Plasmodium falciparum is necessary and useful. We have developed Wondfo rapid diagnostic kit based on nano-gold immunochromatography assay for the detection of P. falciparum in patient specimen. In the present study, we demonstrated the sensitivity and specificity of the rapid diagnostic kit in which nano-gold labeling techniques and the monoclonal antibodies against histidine-rich protein-2 of P. falciparum were used to establish two-antibody sandwich immunochromatographic assay for detecting P. falciparum. By using microscopic examination of blood smears as control, the sensitivity, specificity, and feasibility of Wondfo rapid diagnostic kit was determined in the prompt and accurate diagnosis of malaria. In this study, 1,558 blood samples were collected from outpatient clinics in China and detected by both Wondfo kit and microscopic examination. The Wondfo kit did not show cross-reaction with microfilaria, Toxoplasma gondii, and other parasites in the blood. The patient samples positive for rheumatoid factor, HIV, tuberculosis, and syphilis did not show false positivity when testing with Wondfo kit. The detection sensitivity and specificity of Wondfo rapid diagnostic kit were 95.49% and 99.53%, respectively. These results indicate that our rapid diagnostic assay may be useful for detecting P. falciparum in patient specimen.

A multiplex PCR assay for the rapid and sensitive detection of methicillin-resistant Staphylococcus aureus and simultaneous discrimination of Staphylococcus aureus from coagulase-negative staphylococci.

PubMed

Xu, Benjin; Liu, Ling; Liu, Li; Li, Xinping; Li, Xiaofang; Wang, Xin

2012-11-01

Methicillin-resistant Staphylococcus aureus (MRSA) is a global health concern, which had been detected in food and food production animals. Conventional testing for detection of MRSA takes 3 to 5 d to yield complete information of the organism and its antibiotic sensitivity pattern. So, a rapid method is needed to diagnose and treat the MRSA infections. The present study focused on the development of a multiplex PCR assay for the rapid and sensitive detection of MRSA. The assay simultaneously detected 4 genes, namely, 16S rRNA of the Staphylococcus genus, femA of S. aureus, mecA that encodes methicillin resistance, and one internal control. It was rapid and yielded results within 4 h. The analytical sensitivity and specificity of the multiplex PCR assay was evaluated by comparing it with the conventional method. The analytical sensitivity of the multiplex PCR assay at the DNA level was 10 ng DNA. The analytical specificity was evaluated with 10 reference staphylococci strains and was 100%. The diagnostic evaluation of MRSA was carried out using 360 foodborne staphylococci isolates, and showed 99.1% of specificity, 96.4% of sensitivity, 97.5% of positive predictive value, and 97.3% of negative predictive value compared to the conventional method. The inclusion of an internal control in the multiplex PCR assay is important to exclude false-negative cases. This test can be used as an effective diagnostic and surveillance tool to investigate the spread and emergence of MRSA. © 2012 Institute of Food Technologists®

Study on validity of a rapid diagnostic test kit versus light microscopy for malaria diagnosis in Ahmedabad city, India.

PubMed

Vyas, S; Puwar, B; Patel, V; Bhatt, G; Kulkarni, S; Fancy, M

2014-05-01

Light microscopy of blood smears for diagnosis of malaria in the field has several limitations, notably delays in diagnosis. This study in Ahmedabad in Gujarat State, India, evaluated the diagnostic performance of a rapid diagnostic test for malaria (SD Bioline Malaria Ag P.f/Pan) versus blood smear examination as the gold standard. All fever cases presenting at 13 urban health centres were subjected to rapid diagnostic testing and thick and thin blood smears. A total of 677 cases with fever were examined; 135 (20.0%) tested positive by rapid diagnostic test and 86 (12.7%) by blood smear. The sensitivity of the rapid diagnostic test for malaria was 98.8%, specificity was 91.5%, positive predictive value 63.0% and negative predictive value 99.8%. For detection of Plasmodium falciparum the sensitivity of rapid diagnostic test was 100% and specificity was 97.3%. The results show the acceptability of the rapid test as an alternative to light microscopy in the field setting.

Rapid screening for human-pathogenic Mucorales using rolling circle amplification.

PubMed

Dolatabadi, S; Najafzadeh, M J; de Hoog, G S

2014-12-01

Mucormycosis has emerged as a relatively common severe mycosis in patients with haematological and allogeneic stem cell transplantation. Source of transmission is from unidentified sources in the environment. Early diagnosis of infection and its source of contamination are paramount for rapid and appropriate therapy. In this study, rolling circle amplification (RCA) is introduced as a sensitive, specific and reproducible isothermal DNA amplification technique for rapid molecular identification of six of the most virulent species (Rhizopus microsporus, R. arrhizus var. arrhizus, R. arrhizus var. delemar, Mucor irregularis, Mucor circinelloides, Lichtheimia ramosa, Lichtheimia corymbifera). DNAs of target species were successfully amplified, with no cross reactivity between species. RCA can be considered as a rapid detection method with high specificity and sensitivity, suitable for large screening. © 2014 Blackwell Verlag GmbH.

The sensitivity and specificity of touch preparation for rapid diagnosis of invasive fungal sinusitis: A pilot study.

PubMed

Schuman, Theodore A; Nguyen, Josephine H; Yelverton, Joshua C; Almenara, Jorge A; Powers, Celeste N

2018-01-01

Invasive fungal sinusitis is a morbid pathology that typically affects immunocompromised patients and may quickly progress to fulminant disease. The purpose of this study was to measure the sensitivity and specificity of touch preparation of nasal debridement specimens as a rapid diagnostic tool for invasive fungal sinusitis. A retrospective chart review was performed of 22 patients undergoing nasal debridement due to suspicion for invasive fungal sinusitis over a 10-year period. Thirteen patients had touch preparation of nasal specimens followed by routine histologic processing; 2 of these patients underwent two and 1 patient had three separate debridements, for a total of 17 touch preparations performed. The sensitivity and specificity of touch preparation were calculated by correlating the initial results with the presence of fungal invasion on final pathologic analysis. The sensitivity of touch preparation was 56% (95% confidence interval [CI]: 0.23 to 0.85), specificity was 100% (95% CI: 0.60 to 1.00), positive predictive value was 100% (95% CI: 0.46 to 1.00), and negative predictive value was 67% (95% CI: 0.35 to 0.89). This procedure may be a useful adjunct in situations requiring rapid diagnosis of invasive fungal sinusitis but should not be used as the sole criterion for determining the need for surgical intervention.

Highly sensitive chemiluminescent aptasensor for detecting HBV infection based on rapid magnetic separation and double-functionalized gold nanoparticles.

PubMed

Xi, Zhijiang; Gong, Quan; Wang, Chao; Zheng, Bing

2018-06-21

Hepatitis B virus (HBV) infection is a major global public health problem and one of the leading causes of chronic liver disease. HBsAg is the first serological marker to appear in the blood and is the most important marker of HBV infection. Detection of HBsAg in serum samples is commonly carried out using an immunoassay such as an enzyme-linked immunosorbent assay (ELISA), which is complex to perform, time-consuming, and unsatisfactory for testing sensitivity. Therefore, new methods for highly sensitive detection of HBV infection are urgently needed. Aptamers are specific recognition molecules with high affinity and specificity toward their targets. Biosensors that employ aptamers as biorecognition elements are known as aptasensors. In this study, we select an HBsAg-specific aptamer and use it to develop a new chemiluminescent aptasensor based on rapid magnetic separation and double-functionalized gold nanoparticles. This sensor enables rapid magnetic separation and highly sensitive detection of HBsAg in HBV-positive serum. The detection limit of this HBsAg-detecting chemiluminescent aptasensor is as low as 0.05 ng/mL, which is much lower than the 0.5 ng/mL limit of a typical ELISA used in hospitals. Furthermore, this aptasensor works well and is highly specific to HBV infection.

Evaluation of a rapid immunochromatographic ODK-0901 test for detection of pneumococcal antigen in middle ear fluids and nasopharyngeal secretions.

PubMed

Hotomi, Muneki; Togawa, Akihisa; Takei, Shin; Sugita, Gen; Sugita, Rinya; Kono, Masamitsu; Fujimaki, Yutaka; Kamide, Yosuke; Uchizono, Akihiro; Kanesada, Keiko; Sawada, Shoichi; Okitsu, Naohiro; Tanaka, Yumi; Saijo, Yoko; Yamanaka, Noboru

2012-01-01

Since the incidence of penicillin-resistant Streptococcus pneumoniae has been increasing at an astonishing rate throughout the world, the need for accurate and rapid identification of pneumococci has become increasingly important to determine the appropriate antimicrobial treatment. We have evaluated an immunochromatographic test (ODK-0901) that detects pneumococcal antigens using 264 middle ear fluids (MEFs) and 268 nasopharyngeal secretions (NPSs). A sample was defined to contain S. pneumoniae when optochin and bile sensitive alpha hemolytic streptococcal colonies were isolated by culture. The sensitivity and specificity of the ODK-0901 test were 81.4% and 80.5%, respectively, for MEFs from patients with acute otitis media (AOM). In addition, the sensitivity and specificity were 75.2% and 88.8%, respectively, for NPSs from patients with acute rhinosinusitis. The ODK-0901 test may provide a rapid and highly sensitive evaluation of the presence of S. pneumoniae and thus may be a promising method of identifying pneumococci in MEFs and NPSs.

Evaluation of a Rapid Immunochromatographic ODK-0901 Test for Detection of Pneumococcal Antigen in Middle Ear Fluids and Nasopharyngeal Secretions

PubMed Central

Hotomi, Muneki; Togawa, Akihisa; Takei, Shin; Sugita, Gen; Sugita, Rinya; Kono, Masamitsu; Fujimaki, Yutaka; Kamide, Yosuke; Uchizono, Akihiro; Kanesada, Keiko; Sawada, Shoichi; Okitsu, Naohiro; Tanaka, Yumi; Saijo, Yoko; Yamanaka, Noboru

2012-01-01

Since the incidence of penicillin-resistant Streptococcus pneumoniae has been increasing at an astonishing rate throughout the world, the need for accurate and rapid identification of pneumococci has become increasingly important to determine the appropriate antimicrobial treatment. We have evaluated an immunochromatographic test (ODK-0901) that detects pneumococcal antigens using 264 middle ear fluids (MEFs) and 268 nasopharyngeal secretions (NPSs). A sample was defined to contain S. pneumoniae when optochin and bile sensitive alpha hemolytic streptococcal colonies were isolated by culture. The sensitivity and specificity of the ODK-0901 test were 81.4% and 80.5%, respectively, for MEFs from patients with acute otitis media (AOM). In addition, the sensitivity and specificity were 75.2% and 88.8%, respectively, for NPSs from patients with acute rhinosinusitis. The ODK-0901 test may provide a rapid and highly sensitive evaluation of the presence of S. pneumoniae and thus may be a promising method of identifying pneumococci in MEFs and NPSs. PMID:22448257

Utility of a stressed-SNP real-time PCR assay for the rapid identification of measles vaccine strain in patient samples.

PubMed

Tran, Thomas; Kostecki, Renata; Catton, Michael; Druce, Julian

2018-05-09

Rapid differentiation of wild-type measles virus from measles vaccine strains is crucial during a measles outbreak and in a measles elimination setting. A real-time RT-PCR for the rapid detection of measles vaccine strains was developed with high specificity and greater sensitivity than when compared to traditional measles genotyping methods. The "stressed" minor grove binder TaqMan probe design approach achieves specificity to vaccine strains only, without compromising sensitivity. This assay has proven to be extremely useful in outbreak settings, without requiring sequence genotyping, for over 4 years at the Regional Measles Reference Laboratory for the Western Pacific Region. Copyright © 2018 Tran et al.

Sensitivity for Diagnosing Group A Streptococcal Pharyngitis from Manufacturers is 10% Higher than Reported in Peer-Reviewed Publications.

PubMed

Vachhani, Raj; Patel, Toral; Centor, Robert M; Estrada, Carlos A

2017-01-01

Meta-analyses based on peer-reviewed publications report a sensitivity of approximately 85% for rapid antigen streptococcus tests to diagnose group A streptococcal (GAS) pharyngitis. Because these meta-analyses excluded package inserts, we examined the test characteristics of rapid antigen streptococcal tests and molecular methods that manufacturers report in their package inserts. We included tests available in the US market (Food and Drug Administration, period searched 1993-2015) and used package insert data to calculate pooled sensitivity and specificity. To examine quality, we used the Quality Assessment of Diagnostic Accuracy Studies-2. We excluded 26 tests having different trade names but identical methods and data. The study design was prospective in 41.7% (10 of 24). The pooled sensitivity of the most commonly used method, lateral flow/immunochromatographic, was 95% (95% confidence interval [CI] 94-96) and the pooled specificity was 98% (96-98); 7108 patients. The pooled sensitivity of the polymerase chain reaction or molecular methods was 98% (95% CI 96-98) and the pooled specificity was 96% (95% CI 95-97); 5685 patients. Package inserts include sponsored studies that overestimate the sensitivity of rapid tests to diagnose GAS pharyngitis by approximately 10%. Physicians should understand that package inserts overestimate diagnostic test utility; a negative test cannot be used to exclude GAS pharyngitis.

Field-based performance of three pre-market rapid hepatitis C virus antibody assays in STAHR (Study to Assess Hepatitis C Risk) among young adults who inject drugs in San Diego, CA.

PubMed

Jewett, A; Smith, B D; Garfein, R S; Cuevas-Mota, J; Teshale, E H; Weinbaum, C M

2012-07-01

Approximately 4.1 million Americans are estimated to have been infected with hepatitis C virus (HCV), 45-85% of whom are unaware of their infection. Persons who inject drugs (PWID) account for 55.8% of all persons with HCV antibody (anti-HCV) in the U.S. PWID have limited access to healthcare and are infrequently tested for anti-HCV using conventional laboratory assays. To evaluate performance characteristics (sensitivity and specificity) of three, pre-market rapid point-of-care tests (one oral fluid and two finger-stick assays) from two manufacturers (Chembio and MedMira) in settings providing services to young adult PWID in San Diego, CA. Behavioral risk assessment surveys and testing for HCV were conducted among persons who reported injection drug use (IDU) within the past 6 months as part of the Study to Assess Hepatitis C Risk (STAHR) among PWID aged 18-40 years in 2009-2010. Sensitivity and specificity of the rapid anti-HCV assays were evaluated among STAHR participants, using two commonly used testing algorithms. Variability in sensitivity (76.6-97.1%) and specificity (99.0-100.0%) was found across assays. The highest sensitivity achieved for the Chembio finger-stick blood, Chembio oral fluid and MedMira finger-stick blood tests was 97.1%, 85.4% and 80.0% respectively; the highest specificity was 99.0%, 100.0% and 100.0%, respectively. In multivariate analysis false negative anti-HCV results were associated with female sex for the MedMira blood assay. Sensitive anti-HCV rapid assays are appropriate and feasible for high-prevalence, high-risk populations such as young PWID. Copyright © 2012 Elsevier B.V. All rights reserved.

Application of loop-mediated isothermal amplification assay for the sensitive and rapid diagnosis of visceral leishmaniasis and post-kala-azar dermal leishmaniasis.

PubMed

Verma, Sandeep; Avishek, Kumar; Sharma, Vanila; Negi, Narendra Singh; Ramesh, Venkatesh; Salotra, Poonam

2013-04-01

Loop-mediated isothermal amplification (LAMP) is at the forefront in the search for innovative diagnostics for rapid and specific amplification of target DNA under isothermal conditions. We have applied LAMP assay using SYBR Green for clear-cut naked eye detection of Leishmania (Leishmania) donovani in 200 clinical samples of visceral leishmaniasis (VL) and post-kala-azar dermal leishmaniasis (PKDL). The assay was positive in 53/55 VL blood samples (sensitivity, 96.4%; 95% confidence interval [CI], 87.7-99%), 15/15 VL bone marrow aspirate samples (sensitivity, 100%; 95% CI, 79.6-100%), 60/62 PKDL tissue biopsy samples (sensitivity, 96.8%; 95% CI, 88.9-99.1%), and 1/68 control samples (specificity, 98.5%; 95% CI, 92.1-99.7%). The assay was specific for L. (L.) donovani, the causative species for VL and negative for L. (L.) infantum, L. (L.) tropica, and L. (L.) major. This is the first comprehensive clinical study demonstrating the applicability of the LAMP assay for a rapid and reliable molecular diagnosis of VL and PKDL. Copyright © 2013 Elsevier Inc. All rights reserved.

Performance of a HRP-2/pLDH based rapid diagnostic test at the Bangladesh-India-Myanmar border areas for diagnosis of clinical malaria

PubMed Central

2013-01-01

Background The rapid diagnostic test (RDT) has been adopted in contemporary malaria control and management programmes around the world as it represents a fast and apt alternative for malaria diagnosis in a resource-limited setting. This study assessed the performance of a HRP-2/pLDH based RDT (Parascreen® Pan/Pf) in a laboratory setting utilizing clinical samples obtained from the field. Methods Whole blood samples were obtained from febrile patients referred for malaria diagnosis by clinicians from two different Upazila Health Complexes (UHCs) located near the Bangladesh-India and Bangladesh-Myanmar border where malaria is endemic. RDT was performed on archived samples and sensitivity and specificity evaluated with expert microscopy (EM) and quantitative PCR (qPCR). Results A total of 327 clinical samples were made available for the study, of which 153 were Plasmodium falciparum-positive and 54 were Plasmodium vivax-positive. In comparison with EM, for P. falciparum malaria, the RDT had sensitivity: 96.0% (95% CI, 91.2-98.3) and specificity: 98.2% (95% CI, 94.6-99.5) and for P. vivax, sensitivity: 90.7% (95% CI, 78.9-96.5) and specificity: 98.9% (95% CI, 96.5-99.7). Comparison with qPCR showed, for P. falciparum malaria, sensitivity: 95.4% (95% CI, 90.5-98.0) and specificity: 98.8% (95% CI, 95.4-99.7) and for P. vivax malaria, sensitivity: 89.0% (95% CI,77.0-95.4) and specificity: 98.8% (95% CI, 96.5-99.7). Sensitivity varied according to different parasitaemia for falciparum and vivax malaria diagnosis. Conclusion Parascreen® Pan/Pf Rapid test for malaria showed acceptable sensitivity and specificity in border belt endemic areas of Bangladesh when compared with EM and qPCR. PMID:24172045

A loop-mediated isothermal amplification assay for rapid and sensitive detection of bovine papular stomatitis virus.

PubMed

Kurosaki, Yohei; Okada, Sayaka; Nakamae, Sayuri; Yasuda, Jiro

2016-12-01

Bovine papular stomatitis virus (BPSV) causes pustular cutaneous disease in cattle worldwide. This paper describes the development of a specific loop-mediated isothermal amplification (LAMP) assay to detect BPSV which did not cross-react with other parapoxviruses. To assess analytical sensitivity of this LAMP assay, DNA was extracted from serially diluted BPSV from which the infectious titer was determined by a novel assay based on calf kidney epithelial cells. The LAMP assay had equivalent analytical sensitivity to quantitative PCR, and could detect as few as 86 copies of viral DNA per reaction. These results suggest that the assay is a specific and sensitive technique to rapidly diagnose bovine papular stomatitis in domestic animals. Copyright © 2016 Elsevier B.V. All rights reserved.

Effectiveness of a Rapid Lumbar Spine MRI Protocol Using 3D T2-Weighted SPACE Imaging Versus a Standard Protocol for Evaluation of Degenerative Changes of the Lumbar Spine.

PubMed

Sayah, Anousheh; Jay, Ann K; Toaff, Jacob S; Makariou, Erini V; Berkowitz, Frank

2016-09-01

Reducing lumbar spine MRI scanning time while retaining diagnostic accuracy can benefit patients and reduce health care costs. This study compares the effectiveness of a rapid lumbar MRI protocol using 3D T2-weighted sampling perfection with application-optimized contrast with different flip-angle evolutions (SPACE) sequences with a standard MRI protocol for evaluation of lumbar spondylosis. Two hundred fifty consecutive unenhanced lumbar MRI examinations performed at 1.5 T were retrospectively reviewed. Full, rapid, and complete versions of each examination were interpreted for spondylotic changes at each lumbar level, including herniations and neural compromise. The full examination consisted of sagittal T1-weighted, T2-weighted turbo spin-echo (TSE), and STIR sequences; and axial T1- and T2-weighted TSE sequences (time, 18 minutes 40 seconds). The rapid examination consisted of sagittal T1- and T2-weighted SPACE sequences, with axial SPACE reformations (time, 8 minutes 46 seconds). The complete examination consisted of the full examination plus the T2-weighted SPACE sequence. Sensitivities and specificities of the full and rapid examinations were calculated using the complete study as the reference standard. The rapid and full studies had sensitivities of 76.0% and 69.3%, with specificities of 97.2% and 97.9%, respectively, for all degenerative processes. Rapid and full sensitivities were 68.7% and 66.3% for disk herniation, 85.2% and 81.5% for canal compromise, 82.9% and 69.1% for lateral recess compromise, and 76.9% and 69.7% for foraminal compromise, respectively. Isotropic SPACE T2-weighted imaging provides high-quality imaging of lumbar spondylosis, with multiplanar reformatting capability. Our SPACE-based rapid protocol had sensitivities and specificities for herniations and neural compromise comparable to those of the protocol without SPACE. This protocol fits within a 15-minute slot, potentially reducing costs and discomfort for a large subgroup of patients.

Development of a loop-mediated isothermal amplification assay for rapid detection of Burkholderia mallei.

PubMed

Mirzai, S; Safi, S; Mossavari, N; Afshar, D; Bolourchian, M

2016-08-31

The present study was conducted to establish a Loop-mediated isothermal amplification (LAMP) technique for the rapid detection of B. mallei the etiologic agent of glanders, a highly contagious disease of equines. A set of six specific primers targeting integrase gene cluster were designed for the LAMP test. The reaction was optimized using different temperatures and time intervals. The specificity of the assay was evaluated using DNA from B.pseudomallei and Pseudomonas aeruginosa. The LAMP products were analyzed both visually and under UV light after electrophoresis. The optimized conditions were found to be at 63ºC for 60 min. The assay showed high specificity and sensitivity. It was concluded that the established LAMP assay is a rapid, sensitive and practical tool for detection of B. mallei and early diagnosis of glanders.

Clinical profiling and use of loop-mediated isothermal amplification assay for rapid detection of Mycobacterium tuberculosis from sputum.

PubMed

Poudel, A; Pandey, B D; Lekhak, B; Rijal, B; Sapkota, B R; Suzuki, Y

2009-01-01

Tuberculosis is a global health problem and the situation is worsening with newer incidences of drug resistance and HIV association. Diagnosis of tuberculosis can be done by many methods and test, culture of sputum being the ideal one. Nucleic acid amplification (NAA) assay are more time efficient one, that amplify and detect specific nucleic acid sequences allows rapid, sensitive and specific detection of M. tuberculosis in sputum samples. The present study intends to compile the clinical presentations of the pulmonary tuberculosis (PTB) patients and to evaluate the efficacy of in-house loop-mediated isothermal amplification (LAMP) in detecting Mycobacterium tuberculosis in sputum samples by comparing with microscopy and culture. Two hundred two sputum samples were collected from 202 patients at National Tuberculosis Center, Bhaktapur, Nepal. Complete clinical profiling, epidemiological data and record on BCG vaccination were noted and the samples were subjected for microscopy, culture and in-house LAMP with six primers specific for 16S RNA gene of Mycobacterium tuberculosis. Of the 176 cases of clinical profiling, productive cough was most common symptom in 147 (83.52%), followed by chest pain 136 (77.27%), fever 133 (75.56%) and haemoptysis 61 (34.66%). There was a statistically significant association between BCG vaccination and development of TB (chi(2)=5.33, P=0.02). Of 202 cases, 115 (56.93%) were chest X-ray positive, 101(50%) were direct smear-positive and 100 (49.51%) were culture positive. LAMP had a sensitivity of 97% and specificity of 94.12% while comparing with culture. In addition, its sensitivity and specificity were 91.09% and 89.11% respectively with reference to microscopy. As in our previous study, overall, the result of present study further confirms that the in-house LAMP is a simple, rapid, sensitive and specific DNA amplification technique for PTB diagnosis. Because of rapidity of microscopy and specificity of culture, in-house LAMP assay can be used as a very powerful and useful supplementary tool with complete clinical profiling of the patients for rapid diagnosis of TB in both AFB-positive and negative cases who are suspected as PTB in disease endemic country like Nepal.

Rapid concentration and sensitive detection of hookworm ova from wastewater matrices using a real-time PCR method.

PubMed

Gyawali, P; Sidhu, J P S; Ahmed, W; Jagals, P; Toze, S

2015-12-01

The risk of human hookworm infections from land application of wastewater matrices could be high in regions with high hookworm prevalence. A rapid, sensitive and specific hookworm detection method from wastewater matrices is required in order to assess human health risks. Currently available methods used to identify hookworm ova to the species level are time consuming and lack accuracy. In this study, a real-time PCR method was developed for the rapid, sensitive and specific detection of canine hookworm (Ancylostoma caninum) ova from wastewater matrices. A. caninum was chosen because of its morphological similarity to the human hookworm (Ancylostoma duodenale and Necator americanus). The newly developed PCR method has high detection sensitivity with the ability to detect less than one A. caninum ova from 1 L of secondary treated wastewater at the mean threshold cycle (CT) values ranging from 30.1 to 34.3. The method is also able to detect four A. caninum ova from 1 L of raw wastewater and from ∼4 g of treated sludge with mean CT values ranging from 35.6 to 39.8 and 39.8 to 39.9, respectively. The better detection sensitivity obtained for secondary treated wastewater compared to raw wastewater and sludge samples could be attributed to sample turbidity. The proposed method appears to be rapid, sensitive and specific compared to traditional methods and has potential to aid in the public health risk assessment associated with land application of wastewater matrices. Furthermore, the method can be adapted to detect other helminth ova of interest from wastewater matrices. Crown Copyright © 2015. Published by Elsevier Inc. All rights reserved.

[Efficacy of a rapid test to diagnose Plasmodium vivax in symptomatic patients of Chiapas, Mexico].

PubMed

González-Cerón, Lilia; Rodríguez, Mario H; Betanzos, Angel F; Abadía, Acatl

2005-01-01

To evaluate, under laboratory conditions, the sensitivity and specificity of a rapid diagnostic test (OptiMAL), based on immunoreactive strips, to detect Plasmodium vivax infection in febrile patients in Southern Chiapas, Mexico. The presence of parasites in blood samples of 893 patients was investigated by Giemsa-stained thick blood smear microscopic examination (gold standard). A blood drop from the same sample was smeared on immunoreactive strips to investigate the presence of the parasite pLDH. Discordant results were resolved by PCR amplification of the parasite's 18S SSU rRNA, to discard infection. OptiMAL had an overall sensitivity of 93.3% and its specificity was 99.5%. Its positive and negative predictive values were 96.5% and 98.9%, respectively. Signal intensity in OptiMAL strips correlated well with the parasitemia density in the blood samples (r = 0.601, p = 0.0001). This rapid test had acceptable sensitivity and specificity to detect P. vivax under laboratory conditions and could be useful for malaria diagnosis in field operations in Mexico.

Establishment and Application of a Loop-Mediated Isothermal Amplification Method for Simple, Specific, Sensitive and Rapid Detection of Toxoplasma gondii

PubMed Central

CAO, Lili; CHENG, Ronghua; YAO, Lin; YUAN, Shuxian; YAO, Xinhua

2013-01-01

ABSTRACT The Loop-mediated isothermal amplification (LAMP) method amplifies DNA with high simply, specificity, sensitivity and rapidity. In this study, A LAMP assay with 6 primers targeting a highly conserved region of the GRA1 gene was developed to diagnose Toxoplasma gondii. The reaction time of the LAMP assay was shortened to 30 min after optimizing the reaction system. The LAMP assay was found to be highly specific and stable. The detection limit of the LAMP assay was 10 copies, the same as that of the conventional PCR. We used the LAMP assay to develop a real-time fluorogenic protocol to quantitate T. gondii DNA and generated a log-linear regression plot by plotting the time-to-threshold values against genomic equivalent copies. Furthermore, the LAMP assay was applied to detect T. gondii DNA in 423 blood samples and 380 lymph node samples from 10 pig farms, and positive results were obtained for 7.8% and 8.2% of samples, respectively. The results showed that the LAMP method is slightly more sensitive than conventional PCR (6.1% and 7.6%). Positive samples obtained from 6 pig farms. The LAMP assay established in this study resulted in simple, specific, sensitive and rapid detection of T. gondii DNA and is expected to play an important role in clinical detection of T. gondii. PMID:23965849

Specific, sensitive, and rapid assay for human immunodeficiency virus type 1 pol mutations associated with resistance to zidovudine and didanosine.

PubMed Central

Frenkel, L M; Wagner, L E; Atwood, S M; Cummins, T J; Dewhurst, S

1995-01-01

The effectiveness of antiretroviral therapy may be limited by the development of human immunodeficiency virus type 1 (HIV-1) resistance. Monitoring for resistance will perhaps allow changes in therapy prior to deterioration in the patient's clinical or immunologic status. Our objective was to develop a rapid, specific, and sensitive genotypic assay for HIV-1 resistance to zidovudine (ZDV) and didanosine (ddI) which is simple to perform. In our assay the DNA of HIV-1 pol was amplified by PCR using two sets of nested oligonucleotide primers. Mutations of reverse transcriptase (RT) encoding amino acids (aa) 74 and 41, 70, and 215 which have been associated with HIV-1 resistance to ddI and ZDV, respectively, were detected with a ligase detection reaction (LDR) and indicated colorimetrically. The RT genotypes of 35 patient specimens (140 codons) blindly assessed for these mutations were in agreement by PCR-LDR and by dideoxynucleotide sequencing. To evaluate the limits of the assay, other specimens with mutations close to the ligation site were evaluated by PCR-LDR. The assay was sensitive and specific for all specimens except when mutations occurred within 2 bases on either side of the ligation site. In summary, this PCR-LDR assay specifically, sensitively, and rapidly detected pol mutations (RT aa 74, 41, 70, and 215) associated with HIV-1 resistance to ddI and ZDV. PMID:7714190

Commercial lateral flow devices for rapid detection of peanut (Arachis hypogaea) and hazelnut (Corylus avellana) cross-contamination in the industrial production of cookies.

PubMed

Röder, Martin; Vieths, Stefan; Holzhauser, Thomas

2009-09-01

Lateral flow devices (LFDs) are qualitative immunochromatographic tests for the rapid and specific detection of target analytes. We investigated commercially available LFDs for their ability to detect potentially allergenic peanut and hazelnut in raw cookie dough and chocolate, two important food matrices in the industrial production of cookies. Each three commercial LFDs for the detection of hazelnut and peanut were performed according to the manufacturers' instructions. All LFDs had comparably satisfactory specificity that was investigated with a variety of characteristic foods and food ingredients used in the production of cookies. In concordance with hazelnut-specific enzyme-linked immunosorbent assays (ELISAs), walnut was the most cross-reactive food for hazelnut-specific LFD. The sensitivity was verified in raw cookie doughs and chocolates that were either spiked with peanut or hazelnut between 1 and 25 mg/kg, respectively. Two hazelnut-specific LFDs detected hazelnut at a level of 3.5 mg/kg in both matrices, whereas the third LFD detected hazelnut at a level of 3.9 mg/kg in dough and 12.5 mg/kg in chocolate. Two peanut-specific LFDs detected peanut at a level of 1 mg/kg in both matrices. The third LFD detected peanut at a level of 14.2 mg/kg in chocolate and 4 mg/kg in dough. In conclusion, specific and sensitive LFD were identified for each hazelnut and peanut, having a level of sensitivity that is comparable to commercial ELISA for the investigated matrices. Such sensitive, specific, and rapid tests are useful analytical tools for allergen screening and sanitation in the industrial manufacture of foods.

Specific and Sensitive Isothermal Electrochemical Biosensor for Plant Pathogen DNA Detection with Colloidal Gold Nanoparticles as Probes

NASA Astrophysics Data System (ADS)

Lau, Han Yih; Wu, Haoqi; Wee, Eugene J. H.; Trau, Matt; Wang, Yuling; Botella, Jose R.

2017-01-01

Developing quick and sensitive molecular diagnostics for plant pathogen detection is challenging. Herein, a nanoparticle based electrochemical biosensor was developed for rapid and sensitive detection of plant pathogen DNA on disposable screen-printed carbon electrodes. This 60 min assay relied on the rapid isothermal amplification of target pathogen DNA sequences by recombinase polymerase amplification (RPA) followed by gold nanoparticle-based electrochemical assessment with differential pulse voltammetry (DPV). Our method was 10,000 times more sensitive than conventional polymerase chain reaction (PCR)/gel electrophoresis and could readily identify P. syringae infected plant samples even before the disease symptoms were visible. On the basis of the speed, sensitivity, simplicity and portability of the approach, we believe the method has potential as a rapid disease management solution for applications in agriculture diagnostics.

Specific and Sensitive Isothermal Electrochemical Biosensor for Plant Pathogen DNA Detection with Colloidal Gold Nanoparticles as Probes.

PubMed

Lau, Han Yih; Wu, Haoqi; Wee, Eugene J H; Trau, Matt; Wang, Yuling; Botella, Jose R

2017-01-17

Developing quick and sensitive molecular diagnostics for plant pathogen detection is challenging. Herein, a nanoparticle based electrochemical biosensor was developed for rapid and sensitive detection of plant pathogen DNA on disposable screen-printed carbon electrodes. This 60 min assay relied on the rapid isothermal amplification of target pathogen DNA sequences by recombinase polymerase amplification (RPA) followed by gold nanoparticle-based electrochemical assessment with differential pulse voltammetry (DPV). Our method was 10,000 times more sensitive than conventional polymerase chain reaction (PCR)/gel electrophoresis and could readily identify P. syringae infected plant samples even before the disease symptoms were visible. On the basis of the speed, sensitivity, simplicity and portability of the approach, we believe the method has potential as a rapid disease management solution for applications in agriculture diagnostics.

Development of a rapid diagnostic assay for the detection of tomato chlorotic dwarf viroid based on isothermal reverse-transcription-recombinase polymerase amplification

USDA-ARS?s Scientific Manuscript database

A molecular diagnostic assay utilizing reverse transcription-recombinase polymerase amplification (RT-RPA) at an isothermal constant temperature of 39 °C and target-specific primers and probe were developed for the rapid, sensitive, and specific detection of tomato chlorotic dwarf viroid (TCDVd) in ...

Rapid and specific detection of Asian- and African-lineage Zika viruses

PubMed Central

Chotiwan, Nunya; Brewster, Connie D.; Magalhaes, Tereza; Weger-Lucarelli, James; Duggal, Nisha K.; Rückert, Claudia; Nguyen, Chilinh; Garcia Luna, Selene M.; Fauver, Joseph R.; Andre, Barb; Gray, Meg; Black, William C.; Kading, Rebekah C.; Ebel, Gregory D.; Kuan, Guillermina; Balmaseda, Angel; Jaenisch, Thomas; Marques, Ernesto T. A.; Brault, Aaron C.; Harris, Eva; Foy, Brian D.; Quackenbush, Sandra L.; Perera, Rushika; Rovnak, Joel

2017-01-01

Understanding the dynamics of Zika virus transmission and formulating rational strategies for its control require precise diagnostic tools that are also appropriate for resource-poor environments. We have developed a rapid and sensitive loop-mediated isothermal amplification (LAMP) assay that distinguishes Zika viruses of Asian and African lineages. The assay does not detect chikungunya virus or flaviviruses such as dengue, yellow fever, or West Nile viruses. The assay conditions allowed direct detection of Zika virus RNA in cultured infected cells; in mosquitoes; in virus-spiked samples of human blood, plasma, saliva, urine, and semen; and in infected patient serum, plasma, and semen samples without the need for RNA isolation or reverse transcription. The assay offers rapid, specific, sensitive, and inexpensive detection of the Asian-lineage Zika virus strain that is currently circulating in the Western hemisphere, and can also detect the African-lineage Zika virus strain using separate, specific primers. PMID:28469032

Novel approach based on one-tube nested PCR and a lateral flow strip for highly sensitive diagnosis of tuberculous meningitis.

PubMed

Sun, Yajuan; Chen, Jiajun; Li, Jia; Xu, Yawei; Jin, Hui; Xu, Na; Yin, Rui; Hu, Guohua

2017-01-01

Rapid and sensitive detection of Mycobacterium tuberculosis (M. Tb) in cerebrospinal fluid is crucial in the diagnosis of tuberculous meningitis (TBM), but conventional diagnostic technologies have limited sensitivity and specificity or are time-consuming. In this work, a novel, highly sensitive molecular diagnostic method, one-tube nested PCR-lateral flow strip test (OTNPCR-LFST), was developed for detecting M. tuberculosis. This one-tube nested PCR maintains the sensitivity of conventional two-step nested PCR and reduces both the chance of cross-contamination and the time required for analysis. The PCR product was detected by a lateral flow strip assay, which provided a basis for migration of the test to a point-of-care (POC) microfluidic format. The developed assay had an improved sensitivity compared with traditional PCR, and the limit of detection was up to 1 fg DNA isolated from M. tuberculosis. The assay was also specific for M. tuberculosis, and no cross-reactions were found in other non-target bacteria. The application of this technique to clinical samples was successfully evaluated, and OTNPCR-LFST showed 89% overall sensitivity and 100% specificity for TBM patients. This one-tube nested PCR-lateral flow strip assay is useful for detecting M. tuberculosis in TBM due to its rapidity, high sensitivity and simple manipulation.

Sensitivity and specificity of HIV rapid tests used for research and voluntary counselling and testing.

PubMed

Anzala, O; Sanders, E J; Kamali, A; Katende, M; Mutua, G N; Ruzagira, E; Stevens, G; Simek, M; Price, M

2008-10-01

HIV rapid tests (RT) are a quick and non-technically demanding means to perform HIV voluntary counselling and testing (VCT) but understanding their limitations is vital to delivering quality VCT. To determine the sensitivity and specificity of HIV rapid tests used for research and voluntary counselling and testing at four sites in East Africa. Cross-sectional study. Masaka District, Uganda; a sugar plantation in Kakira, Uganda; Coastal Villages in the Kilifi District of Kenya; and the Urban slum of Kangemi located West of Nairobi, Kenya. Six thousands two hundred and fifty five consenting volunteers were enrolled into the study, and 675 prevalent HIV infections were identified. The RT sensitivity tended to be high for all assays at all sites (97.63-100%) with the exception of the Uni-Gold assay (90.24% in Kangemi, 96.58% in Kilifi). Twenty four RT results were recorded as 'weak positives', 22 (92%) of which were negative by ELISA. There was a high rate of RT false positives in Uganda (positive predictive values ranging from 45.70% to 86.62%). The sensitivity and specificity of the RT varied significantly across sites. The rate of RT misclassification in Uganda suggests that a multiple test algorithm may be preferable to a single test as screener for HIV VCT.

[Rapid identification of potato cultivars using NIR-excited fluorescence and Raman spectroscopy].

PubMed

Dai, Fen; Bergholt, Mads Sylvest; Benjamin, Arnold Julian Vinoj; Hong, Tian-Sheng; Zhiwei, Huang

2014-03-01

Potato is one of the most important food in the world. Rapid and noninvasive identification of potato cultivars plays a important role in the better use of varieties. In this study, The identification ability of optical spectroscopy techniques, including near-infrared (NIR) Raman spectroscopy and NIR fluorescence spectroscopy, for invasive detection of potato cultivars was evaluated. A rapid NIR Raman spectroscopy system was applied to measure the composite Raman and NIR fluorescence spectroscopy of 3 different species of potatoes (98 samples in total) under 785 nm laser light excitation. Then pure Raman and NIR fluorescence spectroscopy were abstracted from the composite spectroscopy, respectively. At last, the partial least squares-discriminant analysis (PLS-DA) was utilized to analyze and classify Raman spectra of 3 different types of potatoes. All the samples were divided into two sets at random: the calibration set (74samples) and prediction set (24 samples), the model was validated using a leave-one-out, cross-validation method. The results showed that both the NIR-excited fluorescence spectra and pure Raman spectra could be used to identify three cultivars of potatoes. The fluorescence spectrum could distinguish the Favorita variety well (sensitivity: 1, specificity: 0.86 and accuracy: 0.92), but the result for Diamant (sensitivity: 0.75, specificity: 0.75 and accuracy: 0. 75) and Granola (sensitivity: 0.16, specificity: 0.89 and accuracy: 0.71) cultivars identification were a bit poorer. We demonstrated that Raman spectroscopy uncovered the main biochemical compositions contained in potato species, and provided a better classification sensitivity, specificity and accuracy (sensitivity: 1, specificity: 1 and accuracy: 1 for all 3 potato cultivars identification) among the three types of potatoes as compared to fluorescence spectroscopy.

Rapid and sensitive detection of Zika virus by reverse transcription loop-mediated isothermal amplification.

PubMed

Wang, Xuan; Yin, Fenggui; Bi, Yuhai; Cheng, Gong; Li, Jing; Hou, Lidan; Li, Yunlong; Yang, Baozhi; Liu, Wenjun; Yang, Limin

2016-12-01

Zika virus (ZIKV) is an arbovirus that recently emerged and has expanded worldwide, causing a global threat and raising international concerns. Current molecular diagnostics, e.g., real-time PCR and reverse transcription PCR (RT-PCR), are time consuming, expensive, and can only be deployed in a laboratory instead of for field diagnostics. This study aimed to develop a one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) platform showing sensitivity, specificity, and more convenience than previous methods, being easily distributed and implemented. Specific primers were designed and screened to target the entire ZIKV genome. The analytical sensitivity and specificity of the assay were evaluated and compared with traditional PCR and quantitative real-time PCR. Three different simulated clinical sample quick preparation protocols were evaluated to establish a rapid and straightforward treatment procedure for clinical specimens in open field detection. The RT-LAMP assay for detection of ZIKV demonstrated superior specificity and sensitivity compared to traditional PCR at the optimum reaction temperature. For the ZIKV RNA standard, the limit of detection was 20 copies/test. For the simulated ZIKV clinical samples, the limit of detection was 0.02 pfu/test, which was one order of magnitude higher than RT-PCR and similar to real-time PCR. The detection limit of simulated ZIKV specimens prepared using a protease quick processing method was consistent with that of samples prepared using commercial nucleic acid extraction kits, indicating that our ZIKV detection method could be used in point-of-care testing. The RT-LAMP assay had excellent sensitivity and specificity for detecting ZIKV and can be deployed together with a rapid specimen processing method, offering the possibility for ZIKV diagnosis outside of the laboratory. Copyright © 2016 Elsevier B.V. All rights reserved.

Rapid quantification of color vision: the cone contrast test.

PubMed

Rabin, Jeff; Gooch, John; Ivan, Douglas

2011-02-09

To describe the design, specificity, and sensitivity of the cone contrast test (CCT), a computer-based, cone-specific (L, M, S) contrast sensitivity test for diagnosing type and severity of color vision deficiency (CVD). The CCT presents a randomized series of colored letters visible only to L, M or S cones in decreasing steps of cone contrast to determine L, M, and S letter-recognition thresholds. Sensitivity and specificity were determined by retrospective comparison of CCT scores to anomaloscope and pseudoisochromatic plate (PIP) results in 1446 applicants for pilot training. CVD was detected in 49 (3.4%) of 1446 applicants with hereditary red-green (protan or deutan) CVD detected in 47 (3.5%) of 1359 men and blue-yellow (tritan) in 2 of 1446. In agreement with the anomaloscope, the CCT showed 100% sensitivity for detection and categorization of CVD (40 deutan, 7 protan, 2 tritan). PIP testing showed lower sensitivity (80% detected; 20% missed) due in part to the applicant's prior experience and/or pretest preparation. CCT specificity for confirming normal color vision was 100% for L and M cone tests and 99.8% for S cones. The CCT has sensitivity and specificity comparable to anomaloscope testing and exceeds PIP sensitivity in practiced observers. The CCT provides a rapid (6 minutes), clinically expedient, measure of color vision for quantifying normal color performance, diagnosing type and severity of hereditary deficiency, and detection of acquired sensitivity loss due to ocular, neurologic, and/or systemic disease, as well as injury and physiological stressors, such as altitude and fatigue.

Operational feasibility of using whole blood in the rapid HIV testing algorithm of a resource-limited settings like Bangladesh.

PubMed

Munshi, Saif U; Oyewale, Tajudeen O; Begum, Shahnaz; Uddin, Ziya; Tabassum, Shahina

2016-03-01

Serum-based rapid HIV testing algorithm in Bangladesh constitutes operational challenge to scaleup HIV testing and counselling (HTC) in the country. This study explored the operational feasibility of using whole blood as alternative to serum for rapid HIV testing in Bangladesh. Whole blood specimens were collected from two study groups. The groups included HIV-positive patients (n = 200) and HIV-negative individuals (n = 200) presenting at the reference laboratory in Dhaka, Bangladesh. The specimens were subjected to rapid HIV tests using the national algorithm with A1 = Alere Determine (United States), A2 = Uni-Gold (Ireland), and A3 = First Response (India). The sensitivity and specificity of the test results, and the operational cost were compared with current serum-based testing. The sensitivities [95% of confidence interval (CI)] for A1, A2, and A3 tests using whole blood were 100% (CI: 99.1-100%), 100% (CI: 99.1-100%), and 97% (CI: 96.4-98.2%), respectively, and specificities of all test kits were 100% (CI: 99.1-100%). Significant (P < 0.05) reduction in the cost of establishing HTC centre and consumables by 94 and 61%, respectively, were observed. The cost of administration and external quality assurance reduced by 39 and 43%, respectively. Overall, there was a 36% cost reduction in total operational cost of rapid HIV testing with blood when compared with serum. Considering the similar sensitivity and specificity of the two specimens, and significant cost reduction, rapid HIV testing with whole blood is feasible. A review of the national HIV rapid testing algorithm with whole blood will contribute toward improving HTC coverage in Bangladesh.

[Evaluation of ICT malaria immunochromatographic Binax NOW ICT P.f/P.v test for rapid diagnosis of malaria in a Colombian endemic area].

PubMed

Pabón, Adriana; Alvarez, Gonzalo; Yánez, Jorge; Céspedes, Carlos; Rodríguez, Yensa; Restrepo, Angela; Blair, Silvia

2007-06-01

One of the strategies to reduce malarial morbidity and mortality is to make an early diagnosis, using simple rapid tests which are highly sensitive and specific. Furthermore, the tests must be easy to perform and understand by local people in such a way that a suitable and prompt antimalarial treatment is guaranteed. The sensitivity and specificity was determined for the immuno-chromographic malaria dipstick (ICT Pf/Pv) test for the rapid diagnosis of malaria in Turbo, Antioquia. The sample consisted of 171 patients distributed into two groups: the first group was 118 patients with acute febrile syndrome compatible with malaria to which ICT Pf/Pv and thick smears were applied simultaneously; a second group was 53 patients with positive diagnosis by thick smear, with follow-up on the 4th and 7th days after beginning treatment. Sensitivity and specificity of the ICT Pf/Pv test for Plasmodium falciparum infections were 54.2% (95%CI: 52.0-53.4%) and 93.6% (95%CI: 93.1-94.2%), respectively. In addition, for Plasmodium vivax the sensitivity and specificity were 80% (95%CI: 77.9-82.1%) and 100% (95%CI: 99.5-100%); there was a 21.4% loss of sensitivity for P. falciparum 21.4% and a 33% loss for P. vivax malaria with parasitaemias under 500 parasites/ul. For the confirmatory test, ICT Pf/Pv showed a global sensitivity of 71.6% with 20.7% false positive and 5.6% false negative results. During follow-up, ICT showed 36% and 34% false positive results for day 4 and 7, respectively. The ICT Pf/Pv test has a poor sensitivity for P. falciparum malaria and its capacity to detect parasitemias under 500 parasites/ul is minimal. As a confirmatory test, the ICT Pf/Pv has a good sensitivity for P. falciparum. Its use for patient follow-up is not recommended.

Low specificity of 2 tetanus rapid tests in Cambodia.

PubMed

Schlumberger, M; Yvonnet, B; Lesage, G; Tep, B

2015-01-01

Rapid testing for tetanus on serum or blood allows for an immediate evaluation of individual protection against tetanus in developed countries, using a "single step" immunochromatographic technique using tetanus toxoid. The specificity of these tests, compared to the reference method for tetanus, mouse serum neutralization testing, has however never been assessed in these countries, due to the difficulty to perform serum neutralization titration in mice, because of animal testing bioethical regulations. A collection of sera from adult volunteers in Cambodia, living in rural environment, was tested for tetanus antibodies by ELISA in France, and by mouse serum neutralization in Vietnam. This allowed estimating the sensitivity and specificity of 2 rapid tetanus tests, available on the market: TQS™ and Tetanotop™. The sensitivity of these tests was adequate, compared to mice serum neutralization test, for a test threshold of 0.01 IU/mL, (100% for TQS™, 91% for Tetanotop™), but their specificity was very low (1% for TQS™ and 13% for Tetanotop™). The results prove that these rapid tests for the assessment of individual protection against tetanus should not be used in the adult rural Cambodian population. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Rapid detection of Enterovirus and Coxsackievirus A10 by a TaqMan based duplex one-step real time RT-PCR assay.

PubMed

Chen, Jingfang; Zhang, Rusheng; Ou, Xinhua; Yao, Dong; Huang, Zheng; Li, Linzhi; Sun, Biancheng

2017-06-01

A TaqMan based duplex one-step real time RT-PCR (rRT-PCR) assay was developed for the rapid detection of Coxsackievirus A10 (CV-A10) and other enterovirus (EVs) in clinical samples. The assay was fully evaluated and found to be specific and sensitive. When applied in 115 clinical samples, a 100% diagnostic sensitivity in CV-A10 detection and 97.4% diagnostic sensitivity in other EVs were found. Copyright © 2017 Elsevier Ltd. All rights reserved.

Aptamer-Nanoparticle Strip Biosensor for Rapid and Sensitive Detection of Cancer Cells

PubMed Central

Mao, Xun; Phillips, Joseph A.; Xu, Hui; Tan, Weihong; Zeng, Lingwen; Liu, Guodong

2009-01-01

We report an aptamer-nanoparticle strip biosensor (ANSB) for the rapid, specific, sensitive and low-cost detection of circulating cancer cells. Known for their high specificity and affinity, aptamers were first selected from live cells by the cell-SELEX (systematic evolution of ligands by exponential enrichment) process. When next combined with the unique optical properties of gold nanoparticles (Au-NPs), ANSBs were prepared on a lateral flow device. Ramos cells were used as a model target cell to demonstrate proof of principle. Under optimal conditions, the ANSB was capable of detecting a minimum of 4000 Ramos cells without instrumentation (visual judgment) and 800 Ramos cells with a portable strip reader within 15 minutes. Importantly, ANSB has successfully detected Ramos cells in human blood, thus providing a rapid, sensitive and low-cost quantitative tool for the detection of circulating cancer cells. ANSB therefore shows great promise for in-field and point-of-care cancer diagnosis and therapy. PMID:19904989

Evaluation of the OnSite malaria rapid test performance in Miandrivazo, Madagascar.

PubMed

Ravaoarisoa, E; Andriamiandranoro, T; Raherinjafy, R; Jahevitra, M; Razanatsiorimalala, S; Andrianaranjaka, V; Randrianarivelojosia, M

2017-10-01

The performance of the malaria rapid diagnostic test OnSite-for detecting pan specific pLDH and Plasmodium falciparum specific HRP2 - was assessed during the malaria transmission peak period in Miandrivazo, in the southwestern part of Madagascar from April 20 to May 6, 2010. At the laboratory, the quality control OnSite Malaria Rapid Test according to the WHO/TDR/FIND method demonstrated that the test had good sensitivity. Of the 218 OnSite tests performed at the Miandrivazo Primary Health Center on patients with fever or a recent history of fever, four (1.8%, 95% CI: 0.6-4.9%) were invalid. Ninety four (43,1%) cases of malaria were confirmed by microscopy, of which 90 were P. falciparum malaria and 4 Plasmodium vivax malaria. With a Cohen's kappa coefficient of 0.94, the agreement between microscopy and OnSite is excellent. Compared with the rapid test CareStart™ commonly used within the public health structures in Madagascar, the sensitivity and specificity of the OnSite test were 97.9% and 96.8%.

Comparison of two antigen assays for rapid intrapartum detection of vaginal group B streptococcal colonization.

PubMed

Green, M; Dashefsky, B; Wald, E R; Laifer, S; Harger, J; Guthrie, R

1993-01-01

As part of a clinical investigation evaluating the efficacy of intrapartum antigen detection for screening for heavy vaginal colonization with group B streptococci (GBS), we compared the performance of modified Bactigen and Directigen GBS latex particle agglutination (LPA) kits. Paired vaginal swabs obtained from women in labor were rapidly transported to the laboratory and used for culturing (both swabs) and LPA testing (one swab by each method). GBS growth was estimated semiquantitatively and further designated as light or heavy growth. Performance specifications for each method were determined by comparing LPA and culture results from the same swab. A total of 4,251 paired swabs were evaluated during the study period. The performance specifications for detecting GBS growth of any degree for Bactigen and Directigen, respectively, were as follows: sensitivity, 20 and 24%; specificity, 99 and 99%. The performance specifications for heavy colonization for Bactigen and Directigen, respectively, were as follows: sensitivity, 57 and 62%; specificity, 99 and 99%. Neither LPA kit was a sensitive indicator of vaginal colonization with GBS or neonatal infection.

Field evaluation of a rapid, visually-read colloidal dye immunofiltration assay for Schistosoma japonicum for screening in areas of low transmission.

PubMed Central

Xiao, Xiang; Wang, Tianping; Ye, Hongzhuan; Qiang, Guangxiang; Wei, Haiming; Tian, Zhigang

2005-01-01

OBJECTIVE: To determine the validity of a recently developed rapid test--a colloidal dye immunofiltration assay (CDIFA)--used by health workers in field settings to identify villagers infected with Schistosoma japonicum. METHODS: Health workers in the field used CDIFA to test samples from 1553 villagers in two areas of low endemicity and an area where S. japonicum was not endemic in Anhui, China. All the samples were then tested in the laboratory by laboratory staff using a standard parasitological method (Kato-Katz), an indirect haemagglutination assay (IHA), and CDIFA. The results of CDIFA performed by health workers were compared with those obtained by Kato-Katz and IHA. FINDINGS: Concordance between the results of CDIFA performed in field settings and in the laboratory was high (kappa index, 0.95; 95% confidence interval, 0.93-0.97). When Kato-Katz was used as the reference test, the overall sensitivity and specificity of CDIFA were 98.5% and 83.6%, respectively in the two villages in areas of low endemicity, while the specificity was 99.8% in the nonendemic village. Compared with IHA, the overall specificity and sensitivity of CDIFA were greater than 99% and 96%, respectively. With the combination of Kato-Katz and IHA as the reference standard, CDIFA had a sensitivity of 95.8% and a specificity of 99.5%, and an accuracy of 98.6% in the two areas of low endemicity. CONCLUSION: CDIFA is a specific, sensitive, and reliable test that can be used for rapid screening for schistosomiasis by health workers in field settings. PMID:16175827

Multicenter Evaluation of the Solana Group A Streptococcus Assay: Comparison with Culture.

PubMed

Uphoff, Timothy S; Buchan, Blake W; Ledeboer, Nathan A; Granato, Paul A; Daly, Judy A; Marti, Tara N

2016-09-01

We compared group A Streptococcus (GAS) culture with a rapid helicase-dependent amplification (HDA) method using 1,082 throat swab specimens. The HDA method demonstrated 98.2% sensitivity and 97.2% specificity. GAS prevalence by culture was 20.7%, and it was 22.6% using the HDA method. In 35 min, the HDA method provided rapid, sensitive GAS detection, making culture confirmation unnecessary. Copyright © 2016 Uphoff et al.

Comparison of four rapid diagnostic tests, ELISA, microscopy and PCR for the detection of Giardia lamblia, Cryptosporidium spp. and Entamoeba histolytica in feces.

PubMed

Van den Bossche, Dorien; Cnops, Lieselotte; Verschueren, Jacob; Van Esbroeck, Marjan

2015-03-01

Microscopy is the diagnostic reference standard for the detection of parasites, but it is labor-intensive and requires experience. Rapid diagnostic tests (RDTs) can provide an alternative to microscopy. RDTs from four different manufacturers were compared to enzyme-linked immunosorbent assay (ELISA), microscopy and/or parasite-specific real-time PCR: ImmunoCardSTAT!®CGE (Meridian Bioscience Inc., Cincinnati, Ohio, USA) (A), Crypto/Giardia Duo-Strip (Coris Bioconcepts, Gembloux, Belgium) (B), RIDA®QUICK Cryptosporidium/Giardia/Entamoeba Combi (R-BioPharm, Darmstadt, Germany) (C) and Giardia/Cryptosporidium Quik Chek (Techlab Inc., Blacksburg, Virginia, USA) (D). Thirty frozen samples were analyzed retrospectively. For Giardia lamblia (n=12) and Cryptosporidium (n=12) sensitivities ranged from 58% (B), over 83% (A, C) to 100% (D) and from 92% (B) to 100% (A, C, D), respectively. Specificity for both G. lamblia and Cryptosporidium was 100% for all RDT brands. Sensitivity for Entamoeba histolytica (n=5) was 100%, while specificity reached 80% (A) to 88% (C). In a prospective study, fresh samples were tested. For G. lamblia (n=30), sensitivity ranged from 66% (B), over 79% (A) and 83% (C) to 100% (D) and specificity varied between 94% (D) and 100% (A, B, C). For Cryptosporidium (n=3), sensitivity was 100% for all brands except (B) (67%) and specificities were 95% (A, B), 98% (C) and 100% (D). E. histolytica (n=1) was detected by both (A) and (C), while specificity was 81% and 87% respectively. RDTs can be a valuable tool when microscopic expertise is poor and in remote and outbreak settings where other techniques are often not available and rapid diagnosis is required. Copyright © 2015 Elsevier B.V. All rights reserved.

Utility of Gram staining for diagnosis of Malassezia folliculitis.

PubMed

Tu, Wei-Ting; Chin, Szu-Ying; Chou, Chia-Lun; Hsu, Che-Yuan; Chen, Yu-Tsung; Liu, Donald; Lee, Woan-Ruoh; Shih, Yi-Hsien

2018-02-01

Malassezia folliculitis (MalF) mimics acne vulgaris and bacterial folliculitis in clinical presentations. The role of Gram staining in rapid diagnosis of MalF has not been well studied. In our study, 32 patients were included to investigate the utility of Gram staining for MalF diagnosis. The final diagnoses of MalF were determined according to clinical presentation, pathological result and treatment response to antifungal agents. Our results show that the sensitivity and specificity of Gram staining are 84.6% and 100%, respectively. In conclusion, Gram staining is a rapid, non-invasive, sensitive and specific method for MalF diagnosis. © 2017 Japanese Dermatological Association.

Rapid screening of toxigenic vibrio cholerae O1 strains from south Iran by PCR-ELISA.

PubMed

Mousavi, Seyed Latif; Nazarian, Shahram; Amani, Jafar; Rahgerdi, Ahmad Karimi

2008-01-01

The ability to sensitively detect Vibrio cholera with PCR-ELISA method represents a considerable advancement over alternative more time-consuming methods for detection of this pathogen. The aim of this research is to evaluate the suitability of a PCR-enzyme-linked immunosorbent assay for sensitive and rapid detection of V. cholera O1. The 398-bp sequence of a gene that codes for the cholera toxin B subunit was amplified by PCR. The digoxigenin-labeled amplified products were coated on microplates and detected by ELISA. The PCR product was also hybridized with biotin labelled probe and detected by ELISA using streptavidin. The specificity of the PCR was determined using 10 bacterial strains and 50 samples from south Iran. The detection limit was 0.5 pg of the genomic DNA and five bacterial cells. Adaptation of PCR into PCR-ELISA assay format facilitates specific and sensitive detection and diagnosis of human cholera disease. We conclude that this PCR-ELISA is a diagnostic method that specifically detects toxin genes in V. cholera O1 strains. It is more rapid and less cumbersome than other diagnostic methods for detection of toxicity in these strains.

Usefulness of an inexpensive, Paracheck test in detecting asymptomatic infectious reservoir of plasmodium falciparum during dry season in an inaccessible terrain in central India.

PubMed

Singh, N; Saxena, A; Sharma, V P

2002-10-01

The performance of a new indigenous rapid diagnostic test, Paracheck Pf was evaluated in detection of Plasmodium falciparum in asymptomatic children in remote forest villages of Mandla district, central India to determine the lower limits of sensitivity and specificity of rapid test. A finger prick blood sample was collected to prepare blood smear and for testing with the Paracheck test. The blood smears were read by an experienced technician blinded to the Paracheck results. The figures for specificity, sensitivity, accuracy and predictive values were calculated using microscopy as gold standard. The new diagnostic test had a sensitivity of 94% and a specificity of 89%. The positive and negative predictive values were 71% and 98%, respectively. The J -index was 0.83%. The rapid test was found to be very easy to perform and the result could be read reliably by field workers. The field evaluation with this new inexpensive test, ($0.65/test) indicates that it could be used as an epidemiological tool in the management of malaria particularly in areas where microscopy is not operationally feasible to attain the goal of the roll back malaria initiative.

Improved Sensitivity for Molecular Detection of Bacterial and Candida Infections in Blood

PubMed Central

Bacconi, Andrea; Richmond, Gregory S.; Baroldi, Michelle A.; Laffler, Thomas G.; Blyn, Lawrence B.; Carolan, Heather E.; Frinder, Mark R.; Toleno, Donna M.; Metzgar, David; Gutierrez, Jose R.; Massire, Christian; Rounds, Megan; Kennel, Natalie J.; Rothman, Richard E.; Peterson, Stephen; Carroll, Karen C.; Wakefield, Teresa; Ecker, David J.

2014-01-01

The rapid identification of bacteria and fungi directly from the blood of patients with suspected bloodstream infections aids in diagnosis and guides treatment decisions. The development of an automated, rapid, and sensitive molecular technology capable of detecting the diverse agents of such infections at low titers has been challenging, due in part to the high background of genomic DNA in blood. PCR followed by electrospray ionization mass spectrometry (PCR/ESI-MS) allows for the rapid and accurate identification of microorganisms but with a sensitivity of about 50% compared to that of culture when using 1-ml whole-blood specimens. Here, we describe a new integrated specimen preparation technology that substantially improves the sensitivity of PCR/ESI-MS analysis. An efficient lysis method and automated DNA purification system were designed for processing 5 ml of whole blood. In addition, PCR amplification formulations were optimized to tolerate high levels of human DNA. An analysis of 331 specimens collected from patients with suspected bloodstream infections resulted in 35 PCR/ESI-MS-positive specimens (10.6%) compared to 18 positive by culture (5.4%). PCR/ESI-MS was 83% sensitive and 94% specific compared to culture. Replicate PCR/ESI-MS testing from a second aliquot of the PCR/ESI-MS-positive/culture-negative specimens corroborated the initial findings in most cases, resulting in increased sensitivity (91%) and specificity (99%) when confirmed detections were considered true positives. The integrated solution described here has the potential to provide rapid detection and identification of organisms responsible for bloodstream infections. PMID:24951806

A simple and rapid DNA extraction method for Chlamydia trachomatis detection from urogenital swabs.

PubMed

Butzler, Matthew A; Reed, Jennifer L; McFall, Sally M

2017-11-01

A highly sensitive and specific Chlamydia trachomatis (CT) diagnostic test was developed by combining filtration isolation of nucleic acid (FINA) extraction with quantitative polymerase chain reaction including an internal control to identify test inhibition. A pilot study of 40 clinical specimens yielded 100% sensitivity and specificity. Copyright © 2017 Elsevier Inc. All rights reserved.

Sensitive and specific detection of classical scrapie prions in the brain of goats by real-time quaking-induced conversion

USDA-ARS?s Scientific Manuscript database

The real-time quaking-induced conversion (RT-QuIC) is a rapid, specific, and sensitive prion seeding activity detection assay that uses recombinant prion protein (rPrPSen) to detect sub-infectious levels of the abnormal isoforms of the prion protein (PrPSc). Although RT-QuIC has been successfully us...

Novel approach based on one-tube nested PCR and a lateral flow strip for highly sensitive diagnosis of tuberculous meningitis

PubMed Central

Sun, Yajuan; Chen, Jiajun; Li, Jia; Xu, Yawei; Jin, Hui; Xu, Na; Yin, Rui

2017-01-01

Rapid and sensitive detection of Mycobacterium tuberculosis (M. Tb) in cerebrospinal fluid is crucial in the diagnosis of tuberculous meningitis (TBM), but conventional diagnostic technologies have limited sensitivity and specificity or are time-consuming. In this work, a novel, highly sensitive molecular diagnostic method, one-tube nested PCR-lateral flow strip test (OTNPCR-LFST), was developed for detecting M. tuberculosis. This one-tube nested PCR maintains the sensitivity of conventional two-step nested PCR and reduces both the chance of cross-contamination and the time required for analysis. The PCR product was detected by a lateral flow strip assay, which provided a basis for migration of the test to a point-of-care (POC) microfluidic format. The developed assay had an improved sensitivity compared with traditional PCR, and the limit of detection was up to 1 fg DNA isolated from M. tuberculosis. The assay was also specific for M. tuberculosis, and no cross-reactions were found in other non-target bacteria. The application of this technique to clinical samples was successfully evaluated, and OTNPCR-LFST showed 89% overall sensitivity and 100% specificity for TBM patients. This one-tube nested PCR-lateral flow strip assay is useful for detecting M. tuberculosis in TBM due to its rapidity, high sensitivity and simple manipulation. PMID:29084241

Screening for Wilson disease in acute liver failure: a comparison of currently available diagnostic tests.

PubMed

Korman, Jessica D; Volenberg, Irene; Balko, Jody; Webster, Joe; Schiodt, Frank V; Squires, Robert H; Fontana, Robert J; Lee, William M; Schilsky, Michael L

2008-10-01

Acute liver failure (ALF) due to Wilson disease (WD) is invariably fatal without emergency liver transplantation. Therefore, rapid diagnosis of WD should aid prompt transplant listing. To identify the best method for diagnosis of ALF due to WD (ALF-WD), data and serum were collected from 140 ALF patients (16 with WD), 29 with other chronic liver diseases and 17 with treated chronic WD. Ceruloplasmin (Cp) was measured by both oxidase activity and nephelometry and serum copper levels by atomic absorption spectroscopy. In patients with ALF, a serum Cp <20 mg/dL by the oxidase method provided a diagnostic sensitivity of 21% and specificity of 84% while, by nephelometry, a sensitivity of 56% and specificity of 63%. Serum copper levels exceeded 200 microg/dL in all ALF-WD patients measured (13/16), but were also elevated in non-WD ALF. An alkaline phosphatase (AP) to total bilirubin (TB) ratio <4 yielded a sensitivity of 94%, specificity of 96%, and a likelihood ratio of 23 for diagnosing fulminant WD. In addition, an AST:ALT ratio >2.2 yielded a sensitivity of 94%, a specificity of 86%, and a likelihood ratio of 7 for diagnosing fulminant WD. Combining the tests provided a diagnostic sensitivity and specificity of 100%. Conventional WD testing utilizing serum ceruloplasmin and/or serum copper levels are less sensitive and specific in identifying patients with ALF-WD than other available tests. More readily available laboratory tests including alkaline phosphatase, bilirubin and serum aminotransferases by contrast provides the most rapid and accurate method for diagnosis of ALF due to WD.

Rapid detection of Mycobacterium tuberculosis and rifampicin resistance in extrapulmonary tuberculosis and sputum smear-negative pulmonary suspects using Xpert MTB/RIF.

PubMed

Ullah, Irfan; Javaid, Arshad; Masud, Haleema; Ali, Mazhar; Basit, Anila; Ahmad, Waqas; Younis, Faisal; Yasmin, Rehana; Khan, Afsar; Jabbar, Abdul; Husain, Masroor; Butt, Zahid Ahmad

2017-04-01

Tuberculosis (TB) is a serious public health problem in developing countries such as Pakistan. Rapid diagnosis of TB and detection of drug resistance are very important for timely and appropriate management of multidrug-resistant TB (MDR-TB). The purpose of this study was to determine the diagnostic efficacy of the Xpert MTB/RIF assay for rapid diagnosis of TB and detection of rifampicin (RIF) resistance in extrapulmonary and smear-negative pulmonary TB suspects. A total of 98 bronchoalveolar lavage fluid (BALF) and 168 extrapulmonary specimens were processed by Xpert MTB/RIF. Culture results are considered as the gold standard for diagnosis of TB, and drug susceptibility testing for detection of RIF resistance. Diagnostic efficacy was measured in terms of sensitivity, specificity and positive and negative predictive values. The Xpert MTB/RIF assay detected 40 (40.8 %) of 98 BALF of presumptive pulmonary TB and 60 (35.7 %) of 168 extrapulmonary specimens. Sensitivity and specificity of the Xpert MTB/RIF assay for detection of TB was 86 and 88.4 %, respectively. The positive predictive value was 71.5 % while negative predictive value was 95.1 %. The Xpert MTB/RIF assay is a rapid and simple technique with high sensitivity and specificity for diagnosing TB and detecting drug resistance in extrapulmonary and smear-negative TB cases.

[Evaluation of a rapid diagnostic test in the diagnosis of toxoplasmosis in pregnant women in Cotonou (Bénin)].

PubMed

Ogouyèmi-Hounto, A; Agbayahoun-Chokki, F; Sissinto Savi de Tove, Y; Biokou Bankole, B; Adinsi de Souza, V; Assogba, M; Kinde-Gazard, D; Massougbodji, A

2014-05-01

The aim of the study was to evaluate the performance of the ImmunoComb® Toxo IgG and ImmunoComb® Toxo IgMassays (rapid diagnostic test) in the laboratory diagnosis of toxoplasmosis in pregnant women in Cotonou. We interviewed 266 pregnant women, who first answered an epidemiological questionnaire, and collected blood samples for measurement of IgG and IgM anti T. gondii antibodies with the ImmunoComb toxo assays and with the ARCHITECT CIMA method. The sensitivity, specificity, positive predictive values (PPV) and negative predictive values (NPV) were calculated to determine the performance of the rapid test. The seroprevalences of IgG against T. gondii by CIMA technique and rapid test were respectively 48.9% and 48.5%. The prevalence increased with age. Performances for IgG were: sensitivity 97%, specificity 100%, PPV 100%, NPV = 97.10%. For IgM, Sensitivity: 33.3% Specificity: 100%, PPV 100%, NPV = 99.2%. Seroprevalence obtained shows that about half of the study population is not immune against T. gondii and requires regular serological monitoring until delivery. According to these results, and given the needs of toxoplasmosis diagnosis on the field characterized by an important decrease of immunized women, this test may be recommended in the laboratory diagnosis of toxoplasmosis in peripheral levels of the health pyramid.

Rapid screening for inflammatory neuropathies by standardized clinical criteria

PubMed Central

Tramontozzi, Louis A.

2016-01-01

Abstract Background: Delay in recognition and treatment of inflammatory neuropathies increases morbidity and mortality. We have developed and standardized 3 clinical screening criteria that rapidly detect inflammatory neuropathies. Methods: We reviewed all patients with definite large fiber neuropathy in 2 different patient populations: 1 from a private neurology clinic and the other from a tertiary care center. Patients were divided into 2 groups: those with an inflammatory neuropathy and those with a noninflammatory neuropathy. We specifically noted the 3 key neuropathy characteristics: onset, distribution, and associated systemic features (ODS). We studied the sensitivity and specificity of ODS in differentiating between inflammatory and noninflammatory neuropathies. Results: A total of 206 patients were included: 51 from the private clinic and 155 from the tertiary care center. The sensitivity of using ODS in detecting an inflammatory neuropathy was 96% and the specificity was 85%. The positive predictive value of ODS was 0.8 and negative predictive value was 0.97. Conclusions: Rapid screening for inflammatory neuropathies by ODS clinical criteria is highly sensitive and has a high negative predictive value for noninflammatory neuropathies. ODS uses simple clinical criteria to rapidly screen for patients with a potentially treatable form of neuropathy and accelerate their diagnostic evaluation. Classification of evidence: This study provides Class IV evidence that 3 neuropathy characteristics—onset, distribution, and associated systemic features—accurately identify patients with inflammatory neuropathies. PMID:29443273

Rapid Molecular Microbiologic Diagnosis of Prosthetic Joint Infection

PubMed Central

Cazanave, Charles; Greenwood-Quaintance, Kerryl E.; Hanssen, Arlen D.; Karau, Melissa J.; Schmidt, Suzannah M.; Gomez Urena, Eric O.; Mandrekar, Jayawant N.; Osmon, Douglas R.; Lough, Lindsay E.; Pritt, Bobbi S.; Steckelberg, James M.

2013-01-01

We previously showed that culture of samples obtained by prosthesis vortexing and sonication was more sensitive than tissue culture for prosthetic joint infection (PJI) diagnosis. Despite improved sensitivity, culture-negative cases remained; furthermore, culture has a long turnaround time. We designed a genus-/group-specific rapid PCR assay panel targeting PJI bacteria and applied it to samples obtained by vortexing and sonicating explanted hip and knee prostheses, and we compared the results to those with sonicate fluid and periprosthetic tissue culture obtained at revision or resection arthroplasty. We studied 434 subjects with knee (n = 272) or hip (n = 162) prostheses; using a standardized definition, 144 had PJI. Sensitivities of tissue culture, of sonicate fluid culture, and of PCR were 70.1, 72.9, and 77.1%, respectively. Specificities were 97.9, 98.3, and 97.9%, respectively. Sonicate fluid PCR was more sensitive than tissue culture (P = 0.04). PCR of prosthesis sonication samples is more sensitive than tissue culture for the microbiologic diagnosis of prosthetic hip and knee infection and provides same-day PJI diagnosis with definition of microbiology. The high assay specificity suggests that typical PJI bacteria may not cause aseptic implant failure. PMID:23658273

Empirical validation of guidelines for the management of pharyngitis in children and adults.

PubMed

McIsaac, Warren J; Kellner, James D; Aufricht, Peggy; Vanjaka, Anita; Low, Donald E

2004-04-07

Recent guidelines for management of pharyngitis vary in their recommendations concerning empirical antibiotic treatment and the need for laboratory confirmation of group A streptococcus (GAS). To assess the impact of guideline recommendations and alternative approaches on identification and treatment of GAS pharyngitis in children and adults. Throat cultures and rapid antigen tests were performed on 787 children and adults aged 3 to 69 years with acute sore throat attending a family medicine clinic in Calgary, Alberta, from September 1999 to August 2002. Recommendations from 2 guidelines (those of the Infectious Diseases Society of America and of the American College of Physicians-American Society of Internal Medicine/American Academy of Family Physicians/US Centers for Disease Control and Prevention) were compared with rapid testing alone, a clinical prediction rule (ie, the modified Centor score), and a criterion standard of treatment for positive throat culture results only. Sensitivity and specificity of each strategy for identifying GAS pharyngitis, total antibiotics recommended, and unnecessary antibiotic prescriptions. In children, sensitivity for streptococcal infection ranged from 85.8% (133/155; 95% confidence interval [CI], 79.3%-90.0%) for rapid testing to 100% for culturing all. In adults, sensitivity ranged from 76.7% (56/73; 95% CI, 65.4%-85.8%) for rapid testing without culture confirmation of negative results to 100% for culturing all. In children, specificity ranged from 90.3% (270/299; 95% CI, 86.4%-93.4%) for use of modified Centor score and throat culture to 100% for culturing all. In adults, specificity ranged from 43.8% (114/260; 95% CI, 37.7%-50.1%) for empirical treatment based on a modified Centor score of 3 or 4 to 100% for culturing all. Total antibiotic prescriptions were lowest with rapid testing (24.7% [194/787]; 95% CI, 21.7%-27.8%) and highest with empirical treatment of high-risk adults (45.7% [360/787]; 95% CI, 42.2%-49.3%), due to a high rate of unnecessary prescriptions in adults (43.8% [146/333]; 95% CI, 38.4%-49.4%). Guideline recommendations for the selective use of throat cultures but antibiotic treatment based only on positive rapid test or throat culture results can reduce unnecessary use of antibiotics for treatment of pharyngitis. However, empirical treatment of adults having a Centor score of 3 or 4 is associated with a high rate of unnecessary antibiotic use. In children, strategies incorporating throat culture or throat culture confirmation of negative rapid antigen test results are highly sensitive and specific. Throat culture of all adults or those selected on the basis of a clinical prediction rule had the highest sensitivity and specificity.

Multicenter Clinical Evaluation of the Novel Alere i Strep A Isothermal Nucleic Acid Amplification Test.

PubMed

Cohen, Daniel M; Russo, Michael E; Jaggi, Preeti; Kline, Jennifer; Gluckman, William; Parekh, Amisha

2015-07-01

Rapid detection of group A beta-hemolytic streptococcus (GAS) is used routinely to help diagnose and treat pharyngitis. However, available rapid antigen detection tests for GAS have relatively low sensitivity, and backup testing is recommended in children. Newer assays are more sensitive yet require excessive time for practical point-of-care use as well as laboratory personnel. The Alere i strep A test is an isothermal nucleic acid amplification test designed to offer highly sensitive results at the point of care within 8 min when performed by nonlaboratory personnel. The performance of the Alere i strep A test was evaluated in a multicenter prospective trial in a Clinical Laboratory Improvement Amendments (CLIA)-waived setting in comparison to bacterial culture in 481 children and adults. Compared to culture, the Aleri i strep A test had 96.0% sensitivity and 94.6% specificity. Discrepant results were adjudicated by PCR and found the Alere i strep A test to have 98.7% sensitivity and 98.5% specificity. Overall, the Alere i strep A test could provide a one-step, rapid, point-of-care testing method for GAS pharyngitis and obviate backup testing on negative results. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Comparison of a Clinical Prediction Rule and a LAM Antigen-Detection Assay for the Rapid Diagnosis of TBM in a High HIV Prevalence Setting

PubMed Central

Patel, Vinod B.; Singh, Ravesh; Connolly, Cathy; Kasprowicz, Victoria; Zumla, Allimudin; Ndungu, Thumbi; Dheda, Keertan

2010-01-01

Background/Objective The diagnosis of tuberculous meningitis (TBM) in resource poor TB endemic environments is challenging. The accuracy of current tools for the rapid diagnosis of TBM is suboptimal. We sought to develop a clinical-prediction rule for the diagnosis of TBM in a high HIV prevalence setting, and to compare performance outcomes to conventional diagnostic modalities and a novel lipoarabinomannan (LAM) antigen detection test (Clearview-TB®) using cerebrospinal fluid (CSF). Methods Patients with suspected TBM were classified as definite-TBM (CSF culture or PCR positive), probable-TBM and non-TBM. Results Of the 150 patients, 84% were HIV-infected (median [IQR] CD4 count = 132 [54; 241] cells/µl). There were 39, 55 and 54 patients in the definite, probable and non-TBM groups, respectively. The LAM sensitivity and specificity (95%CI) was 31% (17;48) and 94% (85;99), respectively (cut-point ≥0.18). By contrast, smear-microscopy was 100% specific but detected none of the definite-TBM cases. LAM positivity was associated with HIV co-infection and low CD4 T cell count (CD4<200 vs. >200 cells/µl; p = 0.03). The sensitivity and specificity in those with a CD4<100 cells/µl was 50% (27;73) and 95% (74;99), respectively. A clinical-prediction rule ≥6 derived from multivariate analysis had a sensitivity and specificity (95%CI) of 47% (31;64) and 98% (90;100), respectively. When LAM was combined with the clinical-prediction-rule, the sensitivity increased significantly (p<0.001) to 63% (47;68) and specificity remained high at 93% (82;98). Conclusions Despite its modest sensitivity the LAM ELISA is an accurate rapid rule-in test for TBM that has incremental value over smear-microscopy. The rule-in value of LAM can be further increased by combination with a clinical-prediction rule, thus enhancing the rapid diagnosis of TBM in HIV-infected persons with advanced immunosuppression. PMID:21203513

An evaluation of the SD Bioline HIV/syphilis duo test.

PubMed

Holden, Jeffrey; Goheen, Joshua; Jett-Goheen, Mary; Barnes, Mathilda; Hsieh, Yu-Hsiang; Gaydos, Charlotte A

2018-01-01

Many health agencies now recommend routine HIV and syphilis testing for pregnant women and most-at-risk populations such as men who have sex with men. With the increased availability of highly sensitive, low cost rapid point-of-care tests, the ability to meet those recommendations has increased, granting wider access to quick and accurate diagnoses. Using blood specimens collected from a Baltimore City Health Department (BCHD) sexually transmitted infection clinic, we evaluated the SD Bioline HIV/Syphilis Duo, a rapid test that simultaneously detects antibodies to HIV and syphilis and has the potential to further benefit clinics and patients by reducing costs, testing complexity, and patient wait times. SD DUO HIV sensitivity and specificity, when compared to BCHD results, were 91.7 and 99.5%, respectively. SD DUO syphilis sensitivity and specificity, when compared to rapid plasma reagin, were 85.7 and 96.8%, respectively, and 69.7 and 99.7%, respectively, when compared to Treponema pallidum particle agglutination (TPPA). SD DUO syphilis sensitivity and specificity, when compared to a traditional screening algorithm, improved to 92.3 and 100%, respectively, and improved to 72.9 and 99.7%, respectively, when compared to a reverse screening algorithm. The HIV component of the SD DUO performed moderately well. However, results for the SD DUO syphilis component, when compared to TPPA, support the need for further testing and assessment.

Multiplex qRT-PCR for the Detection of Western Equine Encephalomyelitis, St. Louis Encephalitis, and West Nile Viral RNA in Mosquito Pools (Diptera: Culicidae)

PubMed Central

Brault, Aaron C.; Fang, Ying; Reisen, William K.

2015-01-01

Following the introduction of West Nile virus into California during the summer of 2003, public health and vector control programs expanded surveillance efforts and were in need of diagnostics capable of rapid, sensitive, and specific detection of arbovirus infections of mosquitoes to inform decision support for intervention. Development of a multiplex TaqMan or real-time semiquantitative reverse transcription polymerase chain reaction (RT-PCR) assay in which three virus specific primer–probe sets were used in the same reaction is described herein for the detection of western equine encephalomyelitis, St. Louis encephalitis and West Nile viral RNA. Laboratory validation and field data from 10 transmission seasons are reported. The comparative sensitivity and specificity of this multiplex assay to singleplex RT-PCR as well as an antigen detection (rapid analyte measurement platform) and standard plaque assays indicate this assay to be rapid and useful in providing mosquito infection data to estimate outbreak risk. PMID:26334826

Rapid and specific detection of Asian- and African-lineage Zika viruses.

PubMed

Chotiwan, Nunya; Brewster, Connie D; Magalhaes, Tereza; Weger-Lucarelli, James; Duggal, Nisha K; Rückert, Claudia; Nguyen, Chilinh; Garcia Luna, Selene M; Fauver, Joseph R; Andre, Barb; Gray, Meg; Black, William C; Kading, Rebekah C; Ebel, Gregory D; Kuan, Guillermina; Balmaseda, Angel; Jaenisch, Thomas; Marques, Ernesto T A; Brault, Aaron C; Harris, Eva; Foy, Brian D; Quackenbush, Sandra L; Perera, Rushika; Rovnak, Joel

2017-05-03

Understanding the dynamics of Zika virus transmission and formulating rational strategies for its control require precise diagnostic tools that are also appropriate for resource-poor environments. We have developed a rapid and sensitive loop-mediated isothermal amplification (LAMP) assay that distinguishes Zika viruses of Asian and African lineages. The assay does not detect chikungunya virus or flaviviruses such as dengue, yellow fever, or West Nile viruses. The assay conditions allowed direct detection of Zika virus RNA in cultured infected cells; in mosquitoes; in virus-spiked samples of human blood, plasma, saliva, urine, and semen; and in infected patient serum, plasma, and semen samples without the need for RNA isolation or reverse transcription. The assay offers rapid, specific, sensitive, and inexpensive detection of the Asian-lineage Zika virus strain that is currently circulating in the Western hemisphere, and can also detect the African-lineage Zika virus strain using separate, specific primers. Copyright © 2017, American Association for the Advancement of Science.

Comparative evaluation of two rapid Salmonella-IgM tests and blood culture in the diagnosis of enteric fever.

PubMed

Prasad, K J; Oberoi, J K; Goel, N; Wattal, C

2015-01-01

Enteric fever is a major public health problem in developing countries like India. An early and accurate diagnosis is necessary for a prompt and effective treatment. We have evaluated the diagnostic accuracy of two Rapid Salmonella-IgM tests (Typhidot-IgM and Enteroscreen-IgM) as compared to blood culture in rapid and early diagnosis of enteric fever. A total of 2,699 patients' serum samples were tested by Rapid Salmonella-IgM tests and blood culture. Patients were divided into two groups. Test group - patients with enteric fever and blood culture positives for Salmonella Typhi; and three types of Controls, i.e. patients with non-enteric fever illnesses, normal healthy controls and patients positive for S. Paratyphi- A. In addition to this we have also evaluated the significance of positive Salmonella-IgM tests among blood culture-negative cases. The overall sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of the Typhidot-IgM test and Enteroscreen-IgM test considering blood culture as gold standard were 97.29% and 88.13%, 97.40% and 87.83%, 98.18% and 92.03%, 96.15% and 82.27%, respectively. Typhidot-IgM test was found to be significantly more sensitive and specific as compared to Enteroscreen-IgM. Among blood culture-negative patients, Rapid Salmonella-IgM tests detected 72.25% additional cases of enteric fever. Although the Rapid Salmonella-IgM tests are meant to diagnose S. Typhi only, but these tests detect S. Paratyphi- A also. Thirty-eight patients who were blood culture-positive for S. Paratyphi- A were also positive by Rapid Salmonella-IgM tests. Rapid Salmonella-IgM tests offer an advantage of increased sensitivity, rapidity, early diagnosis and simplicity over blood culture.

Comparative study of efficacy, rapidity of detection, and cost-effectiveness of potassium hydroxide, calcofluor white, and Chicago sky blue stains in the diagnosis of dermatophytoses.

PubMed

Prakash, R; Prashanth, Hosakere Veerappa; Ragunatha, Shivanna; Kapoor, Meenakshi; Anitha, T K; Krishnamurthy, Veena

2016-04-01

The diagnosis of superficial mycosis such as dermatophytosis is often done clinically. However, in difficult cases, a rapid test with high sensitivity and specificity helps in the immediate confirmation and administration of treatment. The efficacy, rapidity of detection, and cost-effectiveness of KOH preparation, calcofluor white (CW) stain, and Chicago sky blue (CSB) stain in the identification of fungal elements were assessed in patients with dermatophytoses attending the dermatology clinic of a tertiary care hospital. All three tests were performed on each sample collected from 73 patients according to standard procedure. The slides were examined after 5 and 30 minutes in × 10 and × 40 magnifications. The sensitivity and specificity of CW and CSB at 5 and 30 minutes were calculated using KOH preparation as the standard test. CSB stain showed highest positivity (94.5%) within 5 minutes when compared to KOH (75.3%) and CW (83.5%). After 30 minutes, positivity increased in KOH (84.9%) and CW stains (89%), but it remained the same in CSB stain. Both CW and CSB stains when compared to 10% KOH are equally sensitive (100%), but CW was more specific (72.7%), particularly at 30 minutes. When cost of performing tests on 100 specimens is considered, KOH, CW, and CSB stains cost Rs 5, 100, and 15, respectively. CSB stain is a better stain for rapid diagnosis of dermatophytoses because of ease of performance, rapidity of detection, better appreciation of morphology of fungal elements, and cost effectiveness. © 2016 The International Society of Dermatology.

Accuracy of commercially available c-reactive protein rapid tests in the context of undifferentiated fevers in rural Laos.

PubMed

Phommasone, Koukeo; Althaus, Thomas; Souvanthong, Phonesavanh; Phakhounthong, Khansoudaphone; Soyvienvong, Laxoy; Malapheth, Phatthaphone; Mayxay, Mayfong; Pavlicek, Rebecca L; Paris, Daniel H; Dance, David; Newton, Paul; Lubell, Yoel

2016-02-04

C-Reactive Protein (CRP) has been shown to be an accurate biomarker for discriminating bacterial from viral infections in febrile patients in Southeast Asia. Here we investigate the accuracy of existing rapid qualitative and semi-quantitative tests as compared with a quantitative reference test to assess their potential for use in remote tropical settings. Blood samples were obtained from consecutive patients recruited to a prospective fever study at three sites in rural Laos. At each site, one of three rapid qualitative or semi-quantitative tests was performed, as well as a corresponding quantitative NycoCard Reader II as a reference test. We estimate the sensitivity and specificity of the three tests against a threshold of 10 mg/L and kappa values for the agreement of the two semi-quantitative tests with the results of the reference test. All three tests showed high sensitivity, specificity and kappa values as compared with the NycoCard Reader II. With a threshold of 10 mg/L the sensitivity of the tests ranged from 87-98 % and the specificity from 91-98 %. The weighted kappa values for the semi-quantitative tests were 0.7 and 0.8. The use of CRP rapid tests could offer an inexpensive and effective approach to improve the targeting of antibiotics in remote settings where health facilities are basic and laboratories are absent. This study demonstrates that accurate CRP rapid tests are commercially available; evaluations of their clinical impact and cost-effectiveness at point of care is warranted.

Comparison of Conventional PCR, Multiplex PCR, and Loop-Mediated Isothermal Amplification Assays for Rapid Detection of Arcobacter Species

PubMed Central

Wang, Xiaoyu; Seo, Dong Joo; Lee, Min Hwa

2014-01-01

This study aimed to develop a loop-mediated isothermal amplification (LAMP) method for the rapid detection of Arcobacter species. Specific primers targeting the 23S ribosomal RNA gene were used to detect Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii. The specificity of the LAMP primer set was assessed using DNA samples from a panel of Arcobacter and Campylobacter species, and the sensitivity was determined using serial dilutions of Arcobacter species cultures. LAMP showed a 10- to 1,000-fold-higher sensitivity than multiplex PCR, with a detection limit of 2 to 20 CFU per reaction in vitro. Whereas multiplex PCR showed cross-reactivity with Campylobacter species, the LAMP method developed in this study was more sensitive and reliable than conventional PCR or multiplex PCR for the detection of Arcobacter species. PMID:24478488

Evaluation of wondfo rapid diagnostic kit (Pf-HRP2/PAN-pLDH) for diagnosis of malaria by using nano-gold immunochromatographic assay.

PubMed

Wu, Junlin; Peng, Yunping; Liu, Xiaoyun; Li, Wenmei; Tang, Shixing

2014-06-01

Prompt and accurate diagnosis is necessary to start adequate treatment for different affecting species including P. falciparum and P. vivax. Here we described the Wondfo Rapid diagnostic Kit (Pf-HRP2/PAN-pLDH) for the detection of P. falciparum and pan-plasmodium in patient specimen by using a nano-gold immunochromatographic assay. Our rapid assay adapted nano-gold labeling techniques and the monoclonal antibodies (mAbs) against both histidine rich protein-2 (Pf HRP-2) of P. falciparum and pan plasmodium-specific pLDH (pan pLDH). The established two-antibody sandwich immunochromatographic assay could detect P. falciparum and pan-plasmodium. The sensitivity and specificity of Wondfo rapid diagnostic kit were determined by comparing with the "gold standard" of microscopic examination of blood smears. In this study1023 blood samples were collected from outpatient clinics in China and Burma, and detected by both Wondfo kit and microscopic examination. The detection sensitivity and specificity of Wondfo rapid diagnostic kit were 96.46% and 99.67% for P. falciparum (HRP2), 95.03% and 99.24% for pLDH, 96.83% and 99.74% for non-falciparum species, 96.70% and 99.74% for P. vivax, respectively. These results indicate that Wondfo rapid diagnostic assay may be useful for detecting P. falciparum and non-P. falciparum (especially P.v.) in patient specimen.

Assessment of the diagnostic value of a urinary adipsin rapid strip test for pre-eclampsia: A prospective multicenter study.

PubMed

Peng, Bing; Zhang, Li; Yan, Jianying; Qi, Hongbo; Zhang, Weiyuan; Fan, Ling; Hu, Yayi; Lin, Li; Li, Xiaotian; Hu, Rong; Xie, Lan; Zhang, Jianping; Wu, Yanqiao; Li, Li; Zhou, Rong

2017-01-01

The purpose of the present study was to evaluate the clinical value of the rapid strip test of urinary adipsin for the quick diagnosis of pre-eclampsia. In a multicenter diagnostic test study, we studied the diagnostic accuracy of the rapid strip test of urinary adipsin in women presenting with pre-eclampsia. A total of 204 pre-eclampsia patients and 254 healthy pregnant women were recruited for this study, respectively. The rapid strip test of urinary adipsin was used to detect the adipsin in the urine of each patient. The diagnostic value of the rapid strip test of urinary adipsin for pre-eclampsia was demonstrated by its high sensitivity and specificity (95.10% and 97.64%, respectively). The diagnostic accuracy was 96.51%. The consistency analysis showed that the kappa value was 0.93 compared with the gold standard diagnosis of pre-eclampsia. The rapid strip test of urinary adipsin is a non-invasive test for the diagnosis of pre-eclampsia with high sensitivity and specificity. It could help the quick diagnosis of pre-eclampsia in clinical practice greatly. © 2016 Japan Society of Obstetrics and Gynecology.

Rapid detection of duck hepatitis A virus genotype C using reverse transcription loop-mediated isothermal amplification.

PubMed

Li, Chuanfeng; Chen, Zongyan; Meng, Chunchun; Liu, Guangqing

2014-02-01

A one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was used and optimized to develop a rapid and sensitive detection system for duck hepatitis A virus genotype C (DHAV-C) RNA. A set of four specific primers was designed against highly conserved sequences located within the 3D gene from DHAV (strain GX1201). Under optimal reaction conditions, the sensitivity of DHAV-C-specific RT-LAMP was 100-fold higher than that of reverse transcriptase-polymerase chain reaction (RT-PCR), with a detection limit of 0.3pg (6.59×10(4) copies) per reaction. No cross-reactivity was observed from the samples of other duck viruses, which is in good accordance with RT-PCR. Furthermore, a positive reaction can be visually inspected by observing turbidity or color change after the addition of SYBR green I dye. The DHAV-C-specific RT-LAMP assay was applied to the samples and compared with RT-PCR. The positive-sample ratios were 26.7% (12 of 45) by RT-LAMP and 20% (9 of 45) by RT-PCR. Therefore, the newly developed RT-LAMP assay is a rapid, specific, sensitive, and cost-effective method of DHAV-C detection. This assay has potential applications in both clinical diagnosis and field surveillance of DHAV-C infection. Copyright © 2013. Published by Elsevier B.V.

Development of a loop-mediated isothermal amplification (LAMP) assay for rapid and specific detection of common genetically modified organisms (GMOs).

PubMed

Feng, Jiawang; Tang, Shiming; Liu, Lideng; Kuang, Xiaoshan; Wang, Xiaoyu; Hu, Songnan; You, Shuzhu

2015-03-01

Here, we developed a loop-mediated isothermal amplification (LAMP) assay for 11 common transgenic target DNA in GMOs. Six sets of LAMP primer candidates for each target were designed and their specificity, sensitivity, and reproductivity were evaluated. With the optimized LAMP primers, this LAMP assay was simply run within 45-60 min to detect all these targets in GMOs tested. The sensitivity, specificity, and reproductivity of the LAMP assay were further analyzed in comparison with those of Real-Time PCR. In consistent with real-time PCR, detection of 0.5% GMOs in equivalent background DNA was possible using this LAMP assay for all targets. In comparison with real-time PCR, the LAMP assay showed the same results with simple instruments. Hence, the LAMP assay developed can provide a rapid and simple approach for routine screening as well as specific events detection of many GMOs.

Comparison of real-time SYBR green dengue assay with real-time taqman RT-PCR dengue assay and the conventional nested PCR for diagnosis of primary and secondary dengue infection

PubMed Central

Paudel, Damodar; Jarman, Richard; Limkittikul, Kriengsak; Klungthong, Chonticha; Chamnanchanunt, Supat; Nisalak, Ananda; Gibbons, Robert; Chokejindachai, Watcharee

2011-01-01

Background: Dengue fever and dengue hemorrhagic fever are caused by dengue virus. Dengue infection remains a burning problem of many countries. To diagnose acute dengue in the early phase we improve the low cost, rapid SYBR green real time assay and compared the sensitivity and specificity with real time Taqman® assay and conventional nested PCR assay. Aims: To develop low cost, rapid and reliable real time SYBR green diagnostic dengue assay and compare with Taqman real-time assay and conventional nested PCR (modified Lanciotti). Materials and Methods: Eight cultured virus strains were diluted in tenth dilution down to undetectable level by the PCR to optimize the primer, temperature (annealing, and extension and to detect the limit of detection of the assay. Hundred and ninety three ELISA and PCR proved dengue clinical samples were tested with real time SYBR® Green assay, real time Taqman® assay to compare the sensitivity and specificity. Results: Sensitivity and specificity of real time SYBR® green dengue assay (84% and 66%, respectively) was almost comparable to those (81% and 74%) of Taqman real time PCR dengue assay. Real time SYBR® green RT-PCR was equally sensitive in primary and secondary infection while real time Taqman was less sensitive in the secondary infection. Sensitivity of real time Taqman on DENV3 (87%) was equal to SYBR green real time PCR dengue assay. Conclusion: We developed low cost rapid diagnostic SYBR green dengue assay. Further study is needed to make duplex primer assay for the serotyping of dengue virus. PMID:22363089

Biomagnetic separation of Salmonella Typhimurium with high affine and specific ligand peptides isolated by phage display technique

NASA Astrophysics Data System (ADS)

Steingroewer, Juliane; Bley, Thomas; Bergemann, Christian; Boschke, Elke

2007-04-01

Analyses of food-borne pathogens are of great importance in order to minimize the health risk for customers. Thus, very sensitive and rapid detection methods are required. Current conventional culture techniques are very time consuming. Modern immunoassays and biochemical analysis also require pre-enrichment steps resulting in a turnaround time of at least 24 h. Biomagnetic separation (BMS) is a promising more rapid method. In this study we describe the isolation of high affine and specific peptides from a phage-peptide library, which combined with BMS allows the detection of Salmonella spp. with a similar sensitivity as that of immunomagnetic separation using antibodies.

Nucleic Acid-Based Approaches for Detection of Viral Hepatitis

PubMed Central

Behzadi, Payam; Ranjbar, Reza; Alavian, Seyed Moayed

2014-01-01

Context: To determining suitable nucleic acid diagnostics for individual viral hepatitis agent, an extensive search using related keywords was done in major medical library and data were collected, categorized, and summarized in different sections. Results: Various types of molecular biology tools can be used to detect and quantify viral genomic elements and analyze the sequences. These molecular assays are proper technologies for rapidly detecting viral agents with high accuracy, high sensitivity, and high specificity. Nonetheless, the application of each diagnostic method is completely dependent on viral agent. Conclusions: Despite rapidity, automation, accuracy, cost-effectiveness, high sensitivity, and high specificity of molecular techniques, each type of molecular technology has its own advantages and disadvantages. PMID:25789132

EU-approved rapid tests might underestimate bovine spongiform encephalopathy infection in goats.

PubMed

Meloni, Daniela; Bozzetta, Elena; Langeveld, Jan P M; Groschup, Martin H; Goldmann, Wilfred; Andrèoletti, Olivier; Lantier, Isabelle; Van Keulen, Lucien; Bossers, Alex; Pitardi, Danilo; Nonno, Romolo; Sklaviadis, Theodoros; Ingravalle, Francesco; Peletto, Simone; Colussi, Silvia; Acutis, Pier Luigi

2017-03-01

We report the diagnostic sensitivity of 3 EU-approved rapid tests (ELISAs; 1 from IDEXX and 2 from Bio-Rad) for the detection of transmissible spongiform encephalopathy diseases in goats. Ninety-eight goat brainstem samples were tested. All the rapid tests had 100% specificity and ≥80% sensitivity, with the IDEXX test significantly more sensitive than the 2 Bio-Rad tests. All tests detected 100% of samples from goats with clinical scrapie, but missed 8% (IDEXX) to 33% (Bio-Rad SG) of samples from preclinical goats. Importantly, only IDEXX picked up all samples from clinical bovine spongiform encephalopathy (BSE)-infected goats, whereas the other 2 rapid tests missed 15% (Bio-Rad SG) to 25% (Bio-Rad SAP). These results show that a fraction of preclinical scrapie infections are likely missed by EU surveillance, with sensitivity of detection strongly dependent on the choice of the rapid test. Moreover, a significant proportion of clinical BSE infections are underestimated by using either Bio-Rad test. Assuming that the same sensitivity on preclinical goats would also occur in BSE-infected goats, our data suggest that IDEXX is likely the most sensitive test for detecting preclinical field cases of BSE infection in goats, although with an 8% failure rate. These results raise some concerns about the reliability of current EU surveillance figures on BSE infection in goats.

Multicentre evaluations of two new rapid IgG4 tests (WB rapid and panLF rapid) for detection of lymphatic filariasis

PubMed Central

Noordin, Rahmah; Itoh, Makoto; Kimura, Eisaku; Abdul Rahman, Rohana; Ravindran, Balachandran; Mahmud, Rohela; Supali, Taniawati; Weerasooriya, Mirani

2007-01-01

In the global effort to eliminate lymphatic filariasis (LF), rapid field-applicable tests are useful tools that will allow on-site testing to be performed in remote places and the results to be obtained rapidly. Exclusive reliance on the few existing tests may jeopardize the progress of the LF elimination program, thus the introduction of other rapid tests would be useful to address this issue. Two new rapid immunochromatographic IgG4 cassette tests have been produced, namely WB rapid and panLF rapid, for detection of bancroftian filariasis and all three species of lymphatic filaria respectively. WB rapid was developed using BmSXP recombinant antigen, while PanLF rapid was developed using BmR1 and BmSXP recombinant antigens. A total of 165 WB rapid and 276 panLF rapid tests respectively were evaluated at USM and the rest were couriered to another university in Malaysia (98 WB rapid, 129 panLF rapid) and to universities in Indonesia (56 WB rapid, 62 panLF rapid), Japan (152 of each test) and India (18 of each test) where each of the tests underwent independent evaluations in a blinded manner. The average sensitivities of WB rapid and panLF rapid were found to be 97.6% (94%–100%) and 96.5% (94%–100%) respectively; while their average specificities were both 99.6% (99%–100%). Thus this study demonstrated that both the IgG4 rapid tests were highly sensitive and specific, and would be useful additional tests to facilitate the global drive to eliminate this disease. PMID:17961262

Development and evaluation of probe based real time loop mediated isothermal amplification for Salmonella: A new tool for DNA quantification.

PubMed

Mashooq, Mohmad; Kumar, Deepak; Niranjan, Ankush Kiran; Agarwal, Rajesh Kumar; Rathore, Rajesh

2016-07-01

A one step, single tube, accelerated probe based real time loop mediated isothermal amplification (RT LAMP) assay was developed for detecting the invasion gene (InvA) of Salmonella. The probe based RT LAMP is a novel method of gene amplification that amplifies nucleic acid with high specificity and rapidity under isothermal conditions with a set of six primers. The whole procedure is very simple and rapid, and amplification can be obtained in 20min. Detection of gene amplification was accomplished by amplification curve, turbidity and addition of DNA binding dye at the end of the reaction results in colour difference and can be visualized under normal day light and in UV. The sensitivity of developed assay was found 10 fold higher than taqman based qPCR. The specificity of the RT LAMP assay was validated by the absence of any cross reaction with other members of enterobacteriaceae family and other gram negative bacteria. These results indicate that the probe based RT LAMP assay is extremely rapid, cost effective, highly specific and sensitivity and has potential usefulness for rapid Salmonella surveillance. Crown Copyright © 2016. Published by Elsevier B.V. All rights reserved.

Shortened Time to Identify Staphylococcus Species from Blood Cultures and Methicillin Resistance Testing Using CHROMAgar

PubMed Central

Chihara, Shingo; Hayden, Mary K.; Minogue-Corbett, Eileen; Singh, Kamaljit

2009-01-01

The ability to rapidly differentiate coagulase-negative staphylococcus (CoNS) from Staphylococcus aureus and to determine methicillin resistance is important as it affects the decision to treat empiric antibiotic selection. The objective of this study was to evaluate CHROMagar S. aureus and CHROMagar MRSA (Becton Dickinson) for rapid identification of Staphylococcus spp. directly from blood cultures. Consecutive blood culture bottles (BacT Alert 3D SA and SN, bioMérieux) growing gram-positive cocci in clusters were evaluated. An aliquot was plated onto CHROMagar MRSA (C-MRSA) and CHROMagar S. aureus (C-SA) plates, which were read at 12 to 16 hours. C-SA correctly identified 147/147 S. aureus (100% sensitivity); 2 CoNS were misidentified as S. aureus (98% specificity). C-MRSA correctly identified 74/77 MRSA (96% sensitivity). None of the MSSA isolates grew on C-MRSA (100% specificity). In conclusion, CHROMagar is a rapid and sensitive method to distinguish MRSA, MSSA, and coagulase-negative Staphylococcus and may decrease time of reporting positive results. PMID:20016679

Tag Array gene chip rapid diagnosis anti-tuberculosis drug resistance in pulmonary tuberculosis -a feasibility study.

PubMed

Wu, Wenjie; Cheng, Peng; Lyu, Jingtong; Zhang, Zehua; Xu, Jianzhong

2018-05-01

We developed a Tag Array chip for detecting first- and second-line anti tuberculosis drug resistance in pulmonary tuberculosis and compared the analytical performance of the gene chip to that of phenotypic drug susceptibility testing (DST). From November 2011 to April 2016.234 consecutive culture-confirmed, clinically and imaging diagnosed patients with pulmonary tuberculosis from Southwest Hospital, Chongqing were enrolled into the study. Specimens collected during sputum or bronchoalveolar lavage fluid from the pulmonary tuberculosis patients were subjected to M. tuberculosis species identification and drug-resistance detection by the Tag Array gene chip, and evaluate the sensitivity and specificity of chip. A total of 186 patients was diagnosed drug-resistant tuberculosis. The detection of rifampicin (RFP), isoniazid (INH), fluoroquinolones (FQS), streptomycin (SM) resistance genes was highly sensitive and specific: however, for detection of amikacin (AMK), capreomycin (CPM), Kanamycin (KM), specificity was higher, but sensitivity was lower. Sensitivity for the detection of a mutation in the eis promoter region could be improved. The detection sensitivity of the EMB resistance gene was low, therefore it is easy to miss a diagnosis of EMB drug resistance, but its specificity was high. Tag Array chip can achieve rapid, accurate and high-throughput detection of tuberculosis resistance in pulmonary tuberculosis, which has important clinical significance and feasibility. Copyright © 2018. Published by Elsevier Ltd.

Rapid detection of pandemic influenza in the presence of seasonal influenza

PubMed Central

2010-01-01

Background Key to the control of pandemic influenza are surveillance systems that raise alarms rapidly and sensitively. In addition, they must minimise false alarms during a normal influenza season. We develop a method that uses historical syndromic influenza data from the existing surveillance system 'SERVIS' (Scottish Enhanced Respiratory Virus Infection Surveillance) for influenza-like illness (ILI) in Scotland. Methods We develop an algorithm based on the weekly case ratio (WCR) of reported ILI cases to generate an alarm for pandemic influenza. From the seasonal influenza data from 13 Scottish health boards, we estimate the joint probability distribution of the country-level WCR and the number of health boards showing synchronous increases in reported influenza cases over the previous week. Pandemic cases are sampled with various case reporting rates from simulated pandemic influenza infections and overlaid with seasonal SERVIS data from 2001 to 2007. Using this combined time series we test our method for speed of detection, sensitivity and specificity. Also, the 2008-09 SERVIS ILI cases are used for testing detection performances of the three methods with a real pandemic data. Results We compare our method, based on our simulation study, to the moving-average Cumulative Sums (Mov-Avg Cusum) and ILI rate threshold methods and find it to be more sensitive and rapid. For 1% case reporting and detection specificity of 95%, our method is 100% sensitive and has median detection time (MDT) of 4 weeks while the Mov-Avg Cusum and ILI rate threshold methods are, respectively, 97% and 100% sensitive with MDT of 5 weeks. At 99% specificity, our method remains 100% sensitive with MDT of 5 weeks. Although the threshold method maintains its sensitivity of 100% with MDT of 5 weeks, sensitivity of Mov-Avg Cusum declines to 92% with increased MDT of 6 weeks. For a two-fold decrease in the case reporting rate (0.5%) and 99% specificity, the WCR and threshold methods, respectively, have MDT of 5 and 6 weeks with both having sensitivity close to 100% while the Mov-Avg Cusum method can only manage sensitivity of 77% with MDT of 6 weeks. However, the WCR and Mov-Avg Cusum methods outperform the ILI threshold method by 1 week in retrospective detection of the 2009 pandemic in Scotland. Conclusions While computationally and statistically simple to implement, the WCR algorithm is capable of raising alarms, rapidly and sensitively, for influenza pandemics against a background of seasonal influenza. Although the algorithm was developed using the SERVIS data, it has the capacity to be used at other geographic scales and for different disease systems where buying some early extra time is critical. PMID:21106071

The Psychological Essence of the Child Prodigy Phenomenon: Sensitive Periods and Cognitive Experience.

ERIC Educational Resources Information Center

Shavinina, Larisa V.

1999-01-01

Examination of the child prodigy phenomenon suggests it is a result of extremely accelerated mental development during sensitive periods that leads to the rapid growth of a child's cognitive resources and their construction into specific exceptional achievements. (Author/DB)

The development of a loop-mediated isothermal amplification assay for rapid and sensitive detection of abalone herpesvirus DNA.

PubMed

Chen, M H; Kuo, S T; Renault, T; Chang, P H

2014-02-01

A loop-mediated isothermal amplification (LAMP) assay was developed for the detection of abalone herpesvirus DNA. Two pairs of primers were designed, based on the sequence of the DNA polymerase gene of abalone herpesvirus. The reaction temperature and time were optimized to 63°C and 60min, respectively. LAMP amplicons were analyzed by 2% agarose gel electrophoresis or by visual inspection of a colour change emitted by fluorescent dye. The method developed was specific for the detection of abalone herpesvirus, without cross-reactions with other tested herpesviruses including ostreid herpesvirus 1 (OsHV-1), European eel herpesvirus, koi herpesvirus (KHV) and an avian herpesvirus. The LAMP assay was 100 folds more sensitive than a conventional PCR and 10 folds less sensitive than a SYBR Green PCR. These results indicate that the developed LAMP assay is a simple, rapid, sensitive, specific and reliable technique for the detection of abalone herpesvirus. Copyright © 2013 Elsevier B.V. All rights reserved.

Rapid Estrogen Receptor-Mediated Mechanisms Determine the Sexually Dimorphic Sensitivity of Ventricular Myocytes to 17β-Estradiol and the Environmental Endocrine Disruptor Bisphenol A

PubMed Central

Belcher, Scott M.; Chen, Yamei; Yan, Sujuan

2012-01-01

Previously we showed that 17β-estradiol (E2) and/or the xenoestrogen bisphenol A (BPA) alter ventricular myocyte Ca2+ handing, resulting in increased cardiac arrhythmias in a female-specific manner. In the present study, the roles of estrogen receptors (ER) in mediating the rapid contractile and arrhythmogenic effects of estrogens were examined. Contractility was used as an index to assess the impact of E2 or BPA on Ca2+ handling in rodent ventricular myocytes. The concentration-response curve for the stimulatory effects of BPA and E2 on female myocyte was inverted-U shaped. Detectable effects for each compound were observed at 10−12 m, and the most efficacious concentrations for each were at 10−9 m. Sensitivity to E2 and BPA was not observed in male myocytes and was abolished in myocytes from ovariectomized females. Analysis using protein-conjugated E2 suggests that these rapid actions are induced by membrane-associated receptors. Analysis using selective ER agonists and antagonists and a genetic ERβ knockout mouse model showed that ERα and ERβ have opposing actions in myocytes and that the balance between ERβ and ERα signaling is the prime regulator of the sex-specific sensitivity toward estrogens. The response of female myocytes to E2 and BPA is dominated by the stimulatory ERβ-mediated signaling, and the absence of BPA and E2 responsiveness in males is due to a counterbalancing-suppressive action of ERα. We conclude that the sex-specific sensitivity of myocytes to estrogens and the rapid arrhythmogenic effects of BPA and estradiol in the female heart are regulated by the balance between ERα and ERβ signaling. PMID:22166976

Rapid polymerase chain reaction diagnosis of white-nose syndrome in bats

USGS Publications Warehouse

Lorch, J.M.; Gargas, A.; Meteyer, C.U.; Berlowski-Zier, B. M.; Green, D.E.; Shearn-Bochsler, V.; Thomas, N.J.; Blehert, D.S.

2010-01-01

A newly developed polymerase chain reaction (PCR)-based method to rapidly and specifically detect Geomyces destructans on the wings of infected bats from small quantities (1-2 mg) of tissue is described in the current study (methods for culturing and isolating G. destructans from bat skin are also described). The lower limits of detection for PCR were 5 fg of purified fungal DNA or 100 conidia per 2 mg of wing tissue. By using histology as the standard, the PCR had a diagnostic specificity of 100% and a diagnostic sensitivity of 96%, whereas the diagnostic sensitivity of culture techniques was only 54%. The accuracy and fast turnaround time of PCR provides field biologists with valuable information on infection status more rapidly than traditional methods, and the small amount of tissue required for the test would allow diagnosis of white-nose syndrome in live animals.

Molecular Detection and Species-Specific Identification of Medically Important Aspergillus Species by Real-Time PCR in Experimental Invasive Pulmonary Aspergillosis ▿

PubMed Central

Walsh, Thomas J.; Wissel, Mark C.; Grantham, Kevin J.; Petraitiene, Ruta; Petraitis, Vidmantas; Kasai, Miki; Francesconi, Andrea; Cotton, Margaret P.; Hughes, Johanna E.; Greene, Lora; Bacher, John D.; Manna, Pradip; Salomoni, Martin; Kleiboeker, Steven B.; Reddy, Sushruth K.

2011-01-01

Diagnosis of invasive pulmonary aspergillosis (IPA) remains a major challenge to clinical microbiology laboratories. We developed rapid and sensitive quantitative PCR (qPCR) assays for genus- and species-specific identification of Aspergillus infections by use of TaqMan technology. In order to validate these assays and understand their potential diagnostic utility, we then performed a blinded study of bronchoalveolar lavage (BAL) fluid specimens from well-characterized models of IPA with the four medically important species. A set of real-time qPCR primers and probes was developed by utilizing unique ITS1 regions for genus- and species-specific detection of the four most common medically important Aspergillus species (Aspergillus fumigatus, A. flavus, A. niger, and A. terreus). Pan-Aspergillus and species-specific qPCRs with BAL fluid were more sensitive than culture for detection of IPA caused by A. fumigatus in untreated (P < 0.0007) and treated (P ≤ 0.008) animals, respectively. For infections caused by A. terreus and A. niger, culture and PCR amplification from BAL fluid yielded similar sensitivities for untreated and treated animals. Pan-Aspergillus PCR was more sensitive than culture for detection of A. flavus in treated animals (P = 0.002). BAL fluid pan-Aspergillus and species-specific PCRs were comparable in sensitivity to BAL fluid galactomannan (GM) assay. The copy numbers from the qPCR assays correlated with quantitative cultures to determine the pulmonary residual fungal burdens in lung tissue. Pan-Aspergillus and species-specific qPCR assays may improve the rapid and accurate identification of IPA in immunocompromised patients. PMID:21976757

[Accuracy of three methods for the rapid diagnosis of oral candidiasis].

PubMed

Lyu, X; Zhao, C; Yan, Z M; Hua, H

2016-10-09

Objective: To explore a simple, rapid and efficient method for the diagnosis of oral candidiasis in clinical practice. Methods: Totally 124 consecutive patients with suspected oral candidiasis were enrolled from Department of Oral Medicine, Peking University School and Hospital of Stomatology, Beijing, China. Exfoliated cells of oral mucosa and saliva or concentrated oral rinse) obtained from all participants were tested by three rapid smear methods(10% KOH smear, gram-stained smear, Congo red stained smear). The diagnostic efficacy(sensitivity, specificity, Youden's index, likelihood ratio, consistency, predictive value and area under curve(AUC) of each of the above mentioned three methods was assessed by comparing the results with the gold standard(combination of clinical diagnosis, laboratory diagnosis and expert opinion). Results: Gram-stained smear of saliva(or concentrated oral rinse) demonstrated highest sensitivity(82.3%). Test of 10%KOH smear of exfoliated cells showed highest specificity(93.5%). Congo red stained smear of saliva(or concentrated oral rinse) displayed highest diagnostic efficacy(79.0% sensitivity, 80.6% specificity, 0.60 Youden's index, 4.08 positive likelihood ratio, 0.26 negative likelihood ratio, 80% consistency, 80.3% positive predictive value, 79.4% negative predictive value and 0.80 AUC). Conclusions: Test of Congo red stained smear of saliva(or concentrated oral rinse) could be used as a point-of-care tool for the rapid diagnosis of oral candidiasis in clinical practice. Trial registration: Chinese Clinical Trial Registry, ChiCTR-DDD-16008118.

Performance and usefulness of the Hexagon rapid diagnostic test in children with asymptomatic malaria living in the Mount Cameroon region.

PubMed

Wanji, Samuel; Kimbi, Helen K; Eyong, Joan E; Tendongfor, Nicholas; Ndamukong, Judith L

2008-05-22

Rapid and correct diagnosis of malaria is considered an important strategy in the control of the disease. However, it remains to be determined how well these tests can perform in those who harbour the parasite, but are asymptomatic, so that rapid diagnostic tests (RDTs) could be used in rapid mass surveillance in malaria control programmes. Microscopic and immunochromatographic diagnosis of malaria were performed on blood samples from the hyperendemic Mount Cameroon region. Thin and thick blood films were stained with Giemsa and examined under light microscopy for malaria parasites. The RDT was performed on the blood samples for the detection of Plasmodium species. In addition, the performance characteristics of the test were determined using microscopy as gold standard. Results revealed 40.32% to be positive for microscopy and 34.41% to be positive for the RDT. Parasites were detected in a greater proportion of samples as the parasite density increase. Plasmodium falciparum was the predominant Plasmodium species detected in the study population either by microscopy or by the RDT. Overall, the test recorded a sensitivity and specificity of 85.33% and 95.05% respectively, and an accuracy of 91.40%. The sensitivity and specificity of the RDT increased as parasite densities increased. The Hexagon Malaria Combi test showed a high sensitivity and specificity in diagnosing malaria in asymptomatic subjects and so could be suitable for use in mass surveillance programmes for the management and control of malaria.

Use of a Rapid Test for Diagnosis of Dengue during Suspected Dengue Outbreaks in Resource-Limited Regions

PubMed Central

Sharp, Tyler M.; Lalita, Paul; Tikomaidraubuta, Kini; Cardoso, Yolanda Rebello; Naivalu, Taina; Khan, Aalisha Sahu; Marfel, Maria; Hancock, W. Thane; Tomashek, Kay M.; Margolis, Harold S.

2016-01-01

Dengue is major public health problem, globally. Timely verification of suspected dengue outbreaks allows for public health response, leading to the initiation of appropriate clinical care. Because the clinical presentation of dengue is nonspecific, dengue diagnosis would benefit from a sensitive rapid diagnostic test (RDT). We evaluated the diagnostic performance of an RDT that detects dengue virus (DENV) nonstructural protein 1 (NS1) and anti-DENV IgM during suspected acute febrile illness (AFI) outbreaks in four countries. Real-time reverse transcription-PCR and anti-DENV IgM enzyme-linked immunosorbent assay were used to verify RDT results. Anti-DENV IgM RDT sensitivity and specificity ranged from 55.3 to 91.7% and 85.3 to 98.5%, respectively, and NS1 sensitivity and specificity ranged from 49.7 to 92.9% and 22.2 to 89.0%, respectively. Sensitivity varied by timing of specimen collection and DENV serotype. Combined test results moderately improved the sensitivity. The use of RDTs identified dengue as the cause of AFI outbreaks where reference diagnostic testing was limited or unavailable. PMID:27225409

Pathological-Condition-Driven Construction of Supramolecular Nanoassemblies for Bacterial Infection Detection.

PubMed

Li, Li-Li; Ma, Huai-Lei; Qi, Guo-Bin; Zhang, Di; Yu, Faquan; Hu, Zhiyuan; Wang, Hao

2016-01-13

A pyropheophorbide-α-based building block (Ppa-PLGVRG-Van) can be used to construct self-aggregated superstructures in vivo for highly specific and sensitive diagnosis of bacterial infection by noninvasive photoacoustic tomography. This in vivo supramolecular chemistry approach opens a new avenue for efficient, rapid, and early-stage disease diagnosis with high sensitivity and specificity. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Comparison of Sofia Legionella FIA and BinaxNOW® Legionella urinary antigen card in two national reference centers.

PubMed

Beraud, L; Gervasoni, K; Freydiere, A M; Descours, G; Ranc, A G; Vandenesch, F; Lina, G; Gaia, V; Jarraud, S

2015-09-01

The Sofia Legionella Fluorescence Immunoassay (FIA; Quidel) is a recently introduced rapid immunochromatographic diagnostic test for Legionnaires' disease using immunofluorescence technology designed to enhance its sensitivity. The aim of this study was to evaluate its performance for the detection of urinary antigens for Legionella pneumophila serogroup 1 in two National Reference Centers for Legionella. The sensitivity and specificity of the Sofia Legionella FIA test were determined in concentrated and nonconcentrated urine samples, before and after boiling, in comparison with the BinaxNOW® Legionella Urinary Antigen Card (UAC; Alere). Compared with BinaxNOW® Legionella UAC, the sensitivity of the Sofia Legionella test was slightly higher in nonconcentrated urine samples and was identical in concentrated urine samples. The specificity of the Sofia Legionella FIA test was highly reduced by the concentration of urine samples. In nonconcentrated samples, a lack of specificity was observed in 2.3 % of samples, all of them resolved by heat treatment. The Sofia Legionella FIA is a sensitive test for detecting Legionella urinary antigens with no previous urine concentration. However, all positive samples have to be re-tested after boiling to reach a high specificity. The reading is automatized on the Sofia analyzer, which can be connected to laboratory information systems, facilitating accurate and rapid reporting of results.

Combining parasite lactate dehydrogenase-based and histidine-rich protein 2-based rapid tests to improve specificity for diagnosis of malaria Due to Plasmodium knowlesi and other Plasmodium species in Sabah, Malaysia.

PubMed

Grigg, Matthew J; William, Timothy; Barber, Bridget E; Parameswaran, Uma; Bird, Elspeth; Piera, Kim; Aziz, Ammar; Dhanaraj, Prabakaran; Yeo, Tsin W; Anstey, Nicholas M

2014-06-01

Plasmodium knowlesi causes severe and fatal malaria in Malaysia. Microscopic misdiagnosis is common and may delay appropriate treatment. P. knowlesi can cross-react with "species-specific" parasite lactate dehydrogenase (pLDH) monoclonal antibodies used in rapid diagnostic tests (RDTs) to detect P. falciparum and P. vivax. At one tertiary-care hospital and two district hospitals in Sabah, we prospectively evaluated two combination RDTs for malaria diagnosis by using both a pan-Plasmodium-pLDH (pan-pLDH)/P. falciparum-specific-pLDH (Pf-pLDH) RDT (OptiMAL-IT) and a non-P. falciparum VOM-pLDH/Pf-HRP2 RDT (CareStart). Differential cross-reactivity among these combinations was hypothesized to differentiate P. knowlesi from other Plasmodium monoinfections. Among 323 patients with PCR-confirmed P. knowlesi (n = 193), P. falciparum (n = 93), and P. vivax (n = 37) monoinfections, the VOM-pLDH individual component had the highest sensitivity for nonsevere (35%; 95% confidence interval [CI], 27 to 43%) and severe (92%; CI, 81 to 100%) P. knowlesi malaria. CareStart demonstrated a P. knowlesi sensitivity of 42% (CI, 34 to 49%) and specificity of 74% (CI, 65 to 82%), a P. vivax sensitivity of 83% (CI, 66 to 93%) and specificity of 71% (CI, 65 to 76%), and a P. falciparum sensitivity of 97% (CI, 90 to 99%) and specificity of 99% (CI, 97 to 100%). OptiMAL-IT demonstrated a P. knowlesi sensitivity of 32% (CI, 25 to 39%) and specificity of 21% (CI, 15 to 29%), a P. vivax sensitivity of 60% (CI, 42 to 75%) and specificity of 97% (CI, 94 to 99%), and a P. falciparum sensitivity of 82% (CI, 72 to 89%) and specificity of 39% (CI, 33 to 46%). The combination of CareStart plus OptiMAL-IT for P. knowlesi using predefined criteria gave a sensitivity of 25% (CI, 19 to 32%) and specificity of 97% (CI, 92 to 99%). Combining two RDT combinations was highly specific for P. knowlesi malaria diagnosis; however, sensitivity was poor. The specificity of pLDH RDTs was decreased for P. vivax and P. falciparum because of P. knowlesi cross-reactivity and cautions against their use alone in areas where P. knowlesi malaria is endemic. Sensitive P. knowlesi-specific RDTs and/or alternative molecular diagnostic tools are needed in areas where P. knowlesi malaria is endemic. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Accuracy of an Immunochromatographic Diagnostic Test (ICT Malaria Combo Cassette Test) Compared to Microscopy among under Five-Year-Old Children when Diagnosing Malaria in Equatorial Guinea

PubMed Central

Portero, José-Luis; Rubio-Yuste, Maria; Descalzo, Miguel Angel; Raso, Jose; Lwanga, Magdalena; Obono, Jaquelina; Nseng, Gloria; Benito, Agustin; Cano, Jorge

2010-01-01

Conventional malaria diagnosis based on microscopy raises serious difficulties in weak health systems. Cost-effective and sensitive rapid diagnostic tests have been recently proposed as alternatives to microscopy. In Equatorial Guinea, a study was conducted to assess the reliability of a rapid diagnostic test compared to microscopy. The study was designed in accordance with the directives of the Standards for Reporting Diagnostic Accuracy Initiative (STARD). Peripheral thick and thin films for the microscopy diagnosis and a rapid immunochromatographic test (ICT Malaria Combo Cassette Test) were performed on under five-year-old children with malaria suspicion. The ICT test detected Plasmodium spp. infection with a sensitivity of 81.5% and a specificity of 81.9% while P. falciparum diagnosis occurred with a sensitivity of 69.7% and a specificity of 73.7%. The sensitivity of the ICT test increased with higher parasitemias. The general results showed little concordance between the ICT test and microscopy (kappa = 0.28, se: 0.04). In Equatorial Guinea, the ICT Malaria Combo Cassette Test has proven to be an acceptable test to detect high P. falciparum parasitemias. However, the decrease of sensitivity at medium and low parasitemias hampers that ICT can replace properly performed microscopy at present in the diagnosis of malaria in children. PMID:22332024

Time-Resolved Fluorescent Immunochromatography of Aflatoxin B1 in Soybean Sauce: A Rapid and Sensitive Quantitative Analysis.

PubMed

Wang, Du; Zhang, Zhaowei; Li, Peiwu; Zhang, Qi; Zhang, Wen

2016-07-14

Rapid and quantitative sensing of aflatoxin B1 with high sensitivity and specificity has drawn increased attention of studies investigating soybean sauce. A sensitive and rapid quantitative immunochromatographic sensing method was developed for the detection of aflatoxin B1 based on time-resolved fluorescence. It combines the advantages of time-resolved fluorescent sensing and immunochromatography. The dynamic range of a competitive and portable immunoassay was 0.3-10.0 µg·kg(-1), with a limit of detection (LOD) of 0.1 µg·kg(-1) and recoveries of 87.2%-114.3%, within 10 min. The results showed good correlation (R² > 0.99) between time-resolved fluorescent immunochromatographic strip test and high performance liquid chromatography (HPLC). Soybean sauce samples analyzed using time-resolved fluorescent immunochromatographic strip test revealed that 64.2% of samples contained aflatoxin B1 at levels ranging from 0.31 to 12.5 µg·kg(-1). The strip test is a rapid, sensitive, quantitative, and cost-effective on-site screening technique in food safety analysis.

Rapid and Sensitive Detection of Lymphocystis Disease Virus Genotype VII by Loop-Mediated Isothermal Amplification.

PubMed

Valverde, Estefanía J; Cano, Irene; Castro, Dolores; Paley, Richard K; Borrego, Juan J

2017-03-01

Lymphocystis disease virus (LCDV) infections have been described in gilthead seabream (Sparus aurata L.) and Senegalese sole (Solea senegalensis, Kaup), two of the most important marine fish species in the Mediterranean aquaculture. In this study, a rapid, specific, and sensitive detection method for LCDV genotype VII based on loop-mediated isothermal amplification (LAMP) was developed. The LAMP assay, performed using an apparatus with real-time amplification monitoring, was able to specifically detect LCDV genotype VII from clinically positive samples in less than 12 min. In addition, the assay allowed the detection of LCDV in all asymptomatic carrier fish analysed, identified by qPCR, showing an analytical sensitivity of ten copies of viral DNA per reaction. The LCDV LAMP assay has proven to be a promising diagnostic method that can be used easily in fish farms to detect the presence and spread of this iridovirus.

Evaluation of a new rapid diagnostic test for the detection of influenza and RSV.

PubMed

Gómez, Sara; Prieto, Columbiana; Vera, Carmen; R Otero, Joaquín; Folgueira, Lola

2016-05-01

Influenza viruses and respiratory syncytial virus (RSV) can cause an acute respiratory disease that occurs seasonally in epidemic waves. This retrospective study was conducted to evaluate the Sofia(®) Influenza A+B and the Sofia(®) RSV fluorescence immunoassays (FIAs), two novel rapid detection tests (RDTs) for influenza A and B and RSV. Two hundred and nine breath samples were selected from patients with respiratory symptoms determined to be positive/negative for influenza A, influenza B or RSV using one of the reference diagnostic techniques, cell culture and/or RT-PCR (Simplexa™Flu A/B & RSV). The Sofia Influenza A+B FIA was tested on 123 samples (63 from children and 60 from adults) and the Sofia RSV FIA was tested on 86 pediatric samples. Sensitivity and specificity values of both assays were calculated assuming the reference techniques as the gold standard. Sensitivity and specificity values for the Sofia Influenza A+B FIA were 73.1% and 97.8%, respectively. Sensitivity and specificity values for the Sofia RSV FIA were 87.5% and 86.7%, respectively. The sensitivity results obtained for the two assays were considerably higher than those reported for other RDTs. In conclusion, the Sofia Influenza A+B and the Sofia RSV FIAs are appropriate tools for the rapid diagnosis of these viruses. Copyright © 2015 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Performance of a histidine-rich protein 2 rapid diagnostic test, Paracheck Pf®, for detection of malaria infections in Ugandan pregnant women.

PubMed

Dhorda, Mehul; Piola, Patrice; Nyehangane, Dan; Tumwebaze, Benon; Nalusaji, Aisha; Nabasumba, Carolyn; Turyakira, Eleanor; McGready, Rose; Ashley, Elizabeth; Guerin, Philippe J; Snounou, Georges

2012-01-01

Improved laboratory diagnosis is critical to reduce the burden of malaria in pregnancy. Peripheral blood smears appear less sensitive than Plasmodium falciparum histidine-rich protein 2-based rapid diagnostic tests (RDTs) for placental malaria infections in studies conducted at delivery. In this study, 81 women in Uganda in the second or third trimester of pregnancy were followed-up until delivery. At each visit, peripheral blood was tested by blood smear, RDT, and nested species-specific polymerase chain reaction (PCR). Sensitivity and specificity of the tests was calculated with PCR, which detected 22 infections of P. falciparum, as the gold standard. The sensitivity and specificity of blood smears were 36.4% (95% confidence interval [CI] = 18.0-59.2%) and 99.6% (95% CI = 97.7-100%), respectively. The corresponding values for RDT were 31.8% (95% CI = 14.7-54.9%) and 100% (95% CI = 98.3-100%). The RDTs could replace blood smears for diagnosis of malaria in pregnancy by virtue of their relative ease of use. Field-based sensitive tests for malaria in pregnancy are urgently needed.

Nucleic acid detection system and method for detecting influenza

DOEpatents

Cai, Hong; Song, Jian

2015-03-17

The invention provides a rapid, sensitive and specific nucleic acid detection system which utilizes isothermal nucleic acid amplification in combination with a lateral flow chromatographic device, or DNA dipstick, for DNA-hybridization detection. The system of the invention requires no complex instrumentation or electronic hardware, and provides a low cost nucleic acid detection system suitable for highly sensitive pathogen detection. Hybridization to single-stranded DNA amplification products using the system of the invention provides a sensitive and specific means by which assays can be multiplexed for the detection of multiple target sequences.

Combining Parasite Lactate Dehydrogenase-Based and Histidine-Rich Protein 2-Based Rapid Tests To Improve Specificity for Diagnosis of Malaria Due to Plasmodium knowlesi and Other Plasmodium Species in Sabah, Malaysia

PubMed Central

William, Timothy; Barber, Bridget E.; Parameswaran, Uma; Bird, Elspeth; Piera, Kim; Aziz, Ammar; Dhanaraj, Prabakaran; Yeo, Tsin W.; Anstey, Nicholas M.

2014-01-01

Plasmodium knowlesi causes severe and fatal malaria in Malaysia. Microscopic misdiagnosis is common and may delay appropriate treatment. P. knowlesi can cross-react with “species-specific” parasite lactate dehydrogenase (pLDH) monoclonal antibodies used in rapid diagnostic tests (RDTs) to detect P. falciparum and P. vivax. At one tertiary-care hospital and two district hospitals in Sabah, we prospectively evaluated two combination RDTs for malaria diagnosis by using both a pan-Plasmodium-pLDH (pan-pLDH)/P. falciparum-specific-pLDH (Pf-pLDH) RDT (OptiMAL-IT) and a non-P. falciparum VOM-pLDH/Pf-HRP2 RDT (CareStart). Differential cross-reactivity among these combinations was hypothesized to differentiate P. knowlesi from other Plasmodium monoinfections. Among 323 patients with PCR-confirmed P. knowlesi (n = 193), P. falciparum (n = 93), and P. vivax (n = 37) monoinfections, the VOM-pLDH individual component had the highest sensitivity for nonsevere (35%; 95% confidence interval [CI], 27 to 43%) and severe (92%; CI, 81 to 100%) P. knowlesi malaria. CareStart demonstrated a P. knowlesi sensitivity of 42% (CI, 34 to 49%) and specificity of 74% (CI, 65 to 82%), a P. vivax sensitivity of 83% (CI, 66 to 93%) and specificity of 71% (CI, 65 to 76%), and a P. falciparum sensitivity of 97% (CI, 90 to 99%) and specificity of 99% (CI, 97 to 100%). OptiMAL-IT demonstrated a P. knowlesi sensitivity of 32% (CI, 25 to 39%) and specificity of 21% (CI, 15 to 29%), a P. vivax sensitivity of 60% (CI, 42 to 75%) and specificity of 97% (CI, 94 to 99%), and a P. falciparum sensitivity of 82% (CI, 72 to 89%) and specificity of 39% (CI, 33 to 46%). The combination of CareStart plus OptiMAL-IT for P. knowlesi using predefined criteria gave a sensitivity of 25% (CI, 19 to 32%) and specificity of 97% (CI, 92 to 99%). Combining two RDT combinations was highly specific for P. knowlesi malaria diagnosis; however, sensitivity was poor. The specificity of pLDH RDTs was decreased for P. vivax and P. falciparum because of P. knowlesi cross-reactivity and cautions against their use alone in areas where P. knowlesi malaria is endemic. Sensitive P. knowlesi-specific RDTs and/or alternative molecular diagnostic tools are needed in areas where P. knowlesi malaria is endemic. PMID:24696029

Evaluation of a new in-clinic test system to detect feline immunodeficiency virus and feline leukemia virus infection.

PubMed

Sand, Christina; Englert, Theresa; Egberink, Herman; Lutz, Hans; Hartmann, Katrin

2010-06-01

Many in-house tests for the diagnosis of feline immunodeficiency virus (FIV) and feline leukemia virus (FeLV) infection are licensed for use in veterinary practice. A new test with unknown performance has recently appeared on the market. The aims of this study were to define the efficacy of a new in-clinic test system, the Anigen Rapid FIV Ab/FeLV Ag Test, and to compare it with the current leading in-clinic test, the SNAP Kombi Plus FeLV Antigen/FIB Antibody Test. Three-hundred serum samples from randomly selected healthy and diseased cats presented to the Clinic of Small Animal Medicine at Ludwig Maximilian University were tested using both the Anigen Rapid Test and the SNAP Kombi Plus Test. Diagnostic sensitivity, specificity, and positive and negative predictive values were calculated for both tests using Western blot as the gold standard for verification of FIV infection and PCR as the gold standard for FeLV infection. The presence of antibodies against FIV was confirmed by Western blot in 9/300 samples (prevalence 3%). FeLV DNA was detected by PCR in 15/300 samples (prevalence 5%). For FIV infection the Anigen Rapid Test had a sensitivity of 88.9%, specificity of 99.7%, positive predictive value of 88.9%, and negative predictive value of 99.7%. For FeLV infection, the Anigen Rapid Test had a sensitivity of 40.0%, specificity of 100%, positive predictive value of 100%, and negative predictive value of 96.9%. Diagnostic accuracy was similar to that of the SNAP Kombi Plus Test. The new Anigen Rapid FIV Ab/FeLV Ag Test performed very well and can be recommended for use in veterinary practice.

Evaluation of a cow-side milk progesterone assay and assessment of the positive predictive value of oestrus diagnosis by dairy farmers in New South Wales.

PubMed

Ingenhoff, L; Hall, E; House, J K

2016-12-01

The three objectives of this study were to determine the positive predictive value (PPV) of oestrus diagnosis (heat detection accuracy) by dairy farmers, calculate the diagnostic sensitivity and specificity of the P4 Rapid milk progesterone assay for detecting a corpus luteum and evaluate the economics of using a cow-side milk progesterone assay designed to aid oestrus diagnosis. Milk samples were collected from 752 cows diagnosed in oestrus by farm personnel on 14 dairy farms. Samples were tested using the P4 Rapid milk progesterone assay to estimate the PPV of oestrus diagnosis at each farm and a crude pooled mean of PPV of oestrus diagnosis across all farms. A further 156 milk samples were collected from cows with luteal tissue status determined by transrectal ultrasound and tested by the P4 Rapid assay to enable calculation of the sensitivity and specificity of the P4 Rapid assay. For pooled farm samples, the PPV was 97.0%, with a range between farms of 88.9-100%. Sensitivity of the P4 Rapid milk progesterone assay for detecting a corpus luteum was 90.1% and specificity was 98.7%. Misclassification of oestrus in cows previously identified as pregnant was the most common cause of false-positive oestrus diagnoses by farm personnel. Routine testing of milk progesterone in all cows diagnosed in oestrus is not economically justified and may even slightly reduce submission rates; conversely, strategic use of cow-side milk progesterone assays can improve herd reproductive performance by facilitating decisions on whether to rebreed cows previously diagnosed as pregnant. © 2016 Australian Veterinary Association.

Development of rapid immunochromatographic test for hemagglutinin antigen of H7 subtype in patients infected with novel avian influenza A (H7N9) virus.

PubMed

Kang, Keren; Chen, Li; Zhao, Xiang; Qin, Chengfeng; Zhan, Zanwu; Wang, Jihua; Li, Wenmei; Dzakah, Emmanuel E; Huang, Weijuang; Shu, Yuelong; Jiang, Tao; Cao, Wuchun; Xie, Mingquan; Luo, Xiaochun; Tang, Shixing

2014-01-01

Since human infection with the novel H7N9 avian influenza virus was identified in China in March 2013, the relatively high mortality rate and possibility of human-to-human transmission have highlighted the urgent need for sensitive and specific assays for diagnosis of H7N9 infection. We developed a rapid diagnostic test for the novel avian influenza A (H7N9) virus using anti-hemagglutinin (HA) monoclonal antibodies specifically targeting H7 in an immunochromatographic assay system. The assay limit of detection was 103.5 pfu/ml or 103TCID50 of H7N9 virus. The assay specifically detected H7N9 viral isolates and recombinant HA proteins of H7 subtypes including H7N7 and H7N9, but did not react with non-H7 subtypes including H1N1, H3N2, H5N1, H5N9, and H9N2. The detection sensitivity was 59.4% (19/32) for H7N9 patients confirmed by RT-PCR. Moreover, the highest sensitivity of 61.5% (16/26) was obtained when testing H7N9 positive sputum samples while 35.7% (5/14) of nasopharyngeal swabs and 20% (2/10) of fecal samples tested positive. No false positive detection was found when testing 180 H7N9 negative samples. Our novel rapid assay can specifically detect H7 HA antigen, facilitating rapid diagnosis for prevention and control of the on-going H7N9 epidemic.

Development of Rapid Immunochromatographic Test for Hemagglutinin Antigen of H7 Subtype in Patients Infected with Novel Avian Influenza A (H7N9) Virus

PubMed Central

Kang, Keren; Chen, Li; Zhao, Xiang; Qin, Chengfeng; Zhan, Zanwu; Wang, Jihua; Li, Wenmei; Dzakah, Emmanuel E.; Huang, Weijuang; Shu, Yuelong; Jiang, Tao; Cao, Wuchun; Xie, Mingquan; Luo, Xiaochun; Tang, Shixing

2014-01-01

Background Since human infection with the novel H7N9 avian influenza virus was identified in China in March 2013, the relatively high mortality rate and possibility of human-to-human transmission have highlighted the urgent need for sensitive and specific assays for diagnosis of H7N9 infection. Methodology/Principal Findings We developed a rapid diagnostic test for the novel avian influenza A (H7N9) virus using anti-hemagglutinin (HA) monoclonal antibodies specifically targeting H7 in an immunochromatographic assay system. The assay limit of detection was 103.5 pfu/ml or 103TCID50 of H7N9 virus. The assay specifically detected H7N9 viral isolates and recombinant HA proteins of H7 subtypes including H7N7 and H7N9, but did not react with non-H7 subtypes including H1N1, H3N2, H5N1, H5N9, and H9N2. The detection sensitivity was 59.4% (19/32) for H7N9 patients confirmed by RT-PCR. Moreover, the highest sensitivity of 61.5% (16/26) was obtained when testing H7N9 positive sputum samples while 35.7% (5/14) of nasopharyngeal swabs and 20% (2/10) of fecal samples tested positive. No false positive detection was found when testing 180 H7N9 negative samples. Conclusions/Significance Our novel rapid assay can specifically detect H7 HA antigen, facilitating rapid diagnosis for prevention and control of the on-going H7N9 epidemic. PMID:24647358

Establishment of the cross-clade antigen detection system for H5 subtype influenza viruses using peptide monoclonal antibodies specific for influenza virus H5 hemagglutinin.

PubMed

Takahashi, Hitoshi; Nagata, Shiho; Odagiri, Takato; Kageyama, Tsutomu

2018-04-15

The H5 subtype of highly pathogenic avian influenza (H5 HPAI) viruses is a threat to both animal and human public health and has the potential to cause a serious future pandemic in humans. Thus, specific and rapid detection of H5 HPAI viruses is required for infection control in humans. To develop a simple and rapid diagnostic system to detect H5 HPAI viruses with high specificity and sensitivity, we attempted to prepare monoclonal antibodies (mAbs) that specifically recognize linear epitopes in hemagglutinin (HA) of H5 subtype viruses. Nine mAb clones were obtained from mice immunized with a synthetic partial peptide of H5 HA molecules conserved among various H5 HPAI viruses. The antigen-capture enzyme-linked immunosorbent assay using the most suitable combination of these mAbs, which bound specifically to lysed H5 HA under an optimized detergent condition, was specific for H5 viruses and could broadly detect H5 viruses in multiple different clades. Taken together, these peptide mAbs, which recognize linear epitopes in a highly conserved region of H5 HA, may be useful for specific and highly sensitive detection of H5 HPAI viruses and can help in the rapid diagnosis of human, avian, and animal H5 virus infections. Copyright © 2018 Elsevier Inc. All rights reserved.

Rapid assessment of antimicrobial resistance prevalence using a Lot Quality Assurance sampling approach.

PubMed

van Leth, Frank; den Heijer, Casper; Beerepoot, Mariëlle; Stobberingh, Ellen; Geerlings, Suzanne; Schultsz, Constance

2017-04-01

Increasing antimicrobial resistance (AMR) requires rapid surveillance tools, such as Lot Quality Assurance Sampling (LQAS). LQAS classifies AMR as high or low based on set parameters. We compared classifications with the underlying true AMR prevalence using data on 1335 Escherichia coli isolates from surveys of community-acquired urinary tract infection in women, by assessing operating curves, sensitivity and specificity. Sensitivity and specificity of any set of LQAS parameters was above 99% and between 79 and 90%, respectively. Operating curves showed high concordance of the LQAS classification with true AMR prevalence estimates. LQAS-based AMR surveillance is a feasible approach that provides timely and locally relevant estimates, and the necessary information to formulate and evaluate guidelines for empirical treatment.

Rapid Identification of Methicillin-Resistant Staphylococcus aureus (MRSA) by the Vitek MS Saramis system.

PubMed

Shan, Weiguang; Li, Jiaping; Fang, Ying; Wang, Xuan; Gu, Danxia; Zhang, Rong

2016-01-01

A rapid, sensitive, and accurate Vitek MS assay was developed to distinguish clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) from clinical isolates of methicillin-sensitive Staphylococcus aureus (MSSA) by developing an in-house knowledgebase of SuperSpectra. Three unique peaks, including peaks at 2305.6 and 3007.3 Da specific to MRSA, and 6816.7 Da specific to MSSA, were selected for differentiating MRSA and MSSA. This assay accurately identified 84 and 91% of clinical MRSA and MSSA strains out of the total 142 clinically acquired S. aureus strains that were tested. This method will greatly improve the efficiency of single clinical sample identification of MRSA, thereby facilitating a reduction in the transmission of MRSA in clinical settings.

Evaluation of two line probe assays for rapid detection of Mycobacterium tuberculosis, tuberculosis (TB) drug resistance, and non-TB Mycobacteria in HIV-infected individuals with suspected TB.

PubMed

Luetkemeyer, Anne F; Kendall, Michelle A; Wu, Xingye; Lourenço, Maria Cristina; Jentsch, Ute; Swindells, Susan; Qasba, Sarojini S; Sanchez, Jorge; Havlir, Diane V; Grinsztejn, Beatriz; Sanne, Ian M; Firnhaber, Cynthia

2014-04-01

Limited performance data from line probe assays (LPAs), nucleic acid tests used for the rapid diagnosis of tuberculosis (TB), nontuberculosis mycobacteria (NTM), and Mycobacterium tuberculosis drug resistance are available for HIV-infected individuals, in whom paucibacillary TB is common. In this study, the strategy of testing sputum with GenoType MTBDRplus (MTBDR-Plus) and GenoType Direct LPA (Direct LPA) was compared to a gold standard of one mycobacterial growth indicator tube (MGIT) liquid culture. HIV-positive (HIV(+)) individuals with suspected TB from southern Africa and South America with <7 days of TB treatment had 1 sputum specimen tested with Direct LPA, MTBDR-Plus LPA, smear microscopy, MGIT, biochemical identification of mycobacterial species, and culture-based drug-susceptibility testing (DST). Of 639 participants, 59.3% were MGIT M. tuberculosis culture positive, of which 276 (72.8%) were acid-fast bacillus (AFB) smear positive. MTBDR-Plus had a sensitivity of 81.0% and a specificity of 100%, with sensitivities of 44.1% in AFB smear-negative versus 94.6% in AFB smear-positive specimens. For specimens that were positive for M. tuberculosis by MTBDR-Plus, the sensitivity and specificity for rifampin resistance were 91.7% and 96.6%, respectively, and for isoniazid (INH) they were 70.6% and 99.1%. The Direct LPA had a sensitivity of 88.4% and a specificity of 94.6% for M. tuberculosis detection, with a sensitivity of 72.5% in smear-negative specimens. Ten of 639 MGIT cultures grew Mycobacterium avium complex or Mycobacterium kansasii, half of which were detected by Direct LPA. Both LPA assays performed well in specimens from HIV-infected individuals, including in AFB smear-negative specimens, with 72.5% sensitivity for M. tuberculosis identification with the Direct LPA and 44.1% sensitivity with MTBDR-Plus. LPAs have a continued role for use in settings where rapid identification of INH resistance and clinically relevant NTM are priorities.

Comparison of Xpert Flu rapid nucleic acid testing with rapid antigen testing for the diagnosis of influenza A and B.

PubMed

DiMaio, Michael A; Sahoo, Malaya K; Waggoner, Jesse; Pinsky, Benjamin A

2012-12-01

Influenza infections are associated with thousands of hospital admissions and deaths each year. Rapid detection of influenza is important for prompt initiation of antiviral therapy and appropriate patient triage. In this study the Cepheid Xpert Flu assay was compared with two rapid antigen tests, BinaxNOW Influenza A & B and BD Directigen EZ Flu A+B, as well as direct fluorescent antibody testing for the rapid detection of influenza A and B. Using real-time, hydrolysis probe-based, reverse transcriptase PCR as the reference method, influenza A sensitivity was 97.3% for Xpert Flu, 95.9% for direct fluorescent antibody testing, 62.2% for BinaxNOW, and 71.6% for BD Directigen. Influenza B sensitivity was 100% for Xpert Flu and direct fluorescent antibody testing, 54.5% for BinaxNOW, and 48.5% for BD Directigen. Specificity for influenza A was 100% for Xpert Flu, BinaxNOW, and BD Directigen, and 99.2% for direct fluorescent antibody testing. All methods demonstrated 100% specificity for influenza B. These findings support the use of the Xpert Flu assay in settings requiring urgent diagnosis of influenza A and B. Copyright © 2012 Elsevier B.V. All rights reserved.

Evaluation of phage assay for rapid phenotypic detection of rifampicin resistance in Mycobacterium tuberculosis

PubMed Central

Yzquierdo, Sergio Luis; Lemus, Dihadenys; Echemendia, Miguel; Montoro, Ernesto; McNerney, Ruth; Martin, Anandi; Palomino, Juan Carlos

2006-01-01

Background Conventional methods for susceptibility testing require several months before results can be reported. However, rapid methods to determine drug susceptibility have been developed recently. Phage assay have been reported as a rapid useful tools for antimicrobial susceptibility testing. The aim of this study was to apply the Phage assay for rapid detection of resistance on Mycobacterium tuberculosis strains in Cuba. Methods Phage D29 assay was performed on 102 M. tuberculosis strains to detect rifampicin resistance. The results were compared with the proportion method (gold standard) to evaluate the sensitivity and specificity of Phage assay. Results Phage assay results were available in 2 days whereas Proportion Methods results were obtain in 42 days. A total of 44 strains were detected as rifampicin resistant by both methods. However, one strains deemed resistant by Proportion Methods was susceptible by Phage assay. The sensitivity and specificity of Phage assay were 97.8 % and 100% respectively. Conclusion Phage assay provides rapid and reliable results for susceptibility testing; it's easy to perform, requires no specialized equipment and is applicable to drug susceptibility testing in low income countries where tuberculosis is a major public health problem. PMID:16630356

Variola Virus-Specific Diagnostic Assays: Characterization, Sensitivity, and Specificity

PubMed Central

Kondas, Ashley V.; Olson, Victoria A.; Li, Yu; Abel, Jason; Laker, Miriam; Rose, Laura; Wilkins, Kimberly; Turner, Jonathan; Kline, Richard

2015-01-01

A public health response relies upon rapid and reliable confirmation of disease by diagnostic assays. Here, we detail the design and validation of two variola virus-specific real-time PCR assays, since previous assays cross-reacted with newly identified cowpox viruses. The assay specificity must continually be reassessed as other closely related viruses are identified. PMID:25673790

Sensitivity and Specificity of Suspected Case Definition Used during West Africa Ebola Epidemic

PubMed Central

Champaloux, Steven W.; Keïta, Sakoba; Martel, Lise; Bilivogui, Pepe; Knust, Barbara; McCollum, Andrea M.

2018-01-01

Rapid early detection and control of Ebola virus disease (EVD) is contingent on accurate case definitions. Using an epidemic surveillance dataset from Guinea, we analyzed an EVD case definition developed by the World Health Organization (WHO) and used in Guinea. We used the surveillance dataset (March–October 2014; n = 2,847 persons) to identify patients who satisfied or did not satisfy case definition criteria. Laboratory confirmation determined cases from noncases, and we calculated sensitivity, specificity and predictive values. The sensitivity of the defintion was 68.9%, and the specificity of the definition was 49.6%. The presence of epidemiologic risk factors (i.e., recent contact with a known or suspected EVD case-patient) had the highest sensitivity (74.7%), and unexplained deaths had the highest specificity (92.8%). Results for case definition analyses were statistically significant (p<0.05 by χ2 test). Multiple components of the EVD case definition used in Guinea contributed to improved overall sensitivity and specificity. PMID:29260687

Evaluation of the SD Bioline Cholera Rapid Diagnostic Test During the 2016 Cholera Outbreak in Lusaka, Zambia.

PubMed

Mwaba, John; Ferreras, Eva; Chizema-Kawesa, Elizabeth; Mwimbe, Daniel; Tafirenyika, Francis; Rauzier, Jean; Blake, Alexandre; Rakesh, Ankur; Poncin, Marc; Stoitsova, Savina; Kwenda, Geoffrey; Azman, Andrew S; Chewe, Orbrie; Serafini, Micaela; Lukwesa-Musyani, Chileshe; Cohuet, Sandra; Quilici, Marie-Laure; Luquero, Francisco J; Page, Anne-Laure

2018-05-31

To assess the performance of the SD Bioline Cholera Ag O1/O139 rapid diagnostic test (RDT) compared to a reference standard combining culture and PCR for the diagnosis of cholera cases during an outbreak. RDT and bacterial culture were performed on site using fresh stools collected from cholera suspected cases, and from stools enriched in alkaline peptone water. Dried stool samples on filter paper were tested for V. cholerae by PCR in Lusaka (as part of a laboratory technology transfer project) and at a reference laboratory in Paris, France. A sample was considered positive for cholera by the reference standard if any of the culture or PCR tests was positive for V. cholerae O1 or O139. Among the 170 samples tested with SD Bioline and compared to the reference standard, the RDT showed a sensitivity of 90.9% (95% CI: 81.3-96.6) and specificity of 95.0% (95% CI: 89.1-98.4). After enrichment, the sensitivity was 95.5% (95% CI: 87.3-99.1) and specificity 100% (5% CI: 96.5-100). The observed sensitivity and specificity were within recommendations set by the Global Task Force for Cholera Control on the use of cholera RDT (sensitivity=90% : specificity=85%). Although the sample size was small, our findings suggest that the SD Bioline RDT could be used in the field to rapidly alert public health officials to the likely presence of cholera cases when an outbreak is suspected. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

CdSe/ZnS Quantum Dot-Labeled Lateral Flow Strips for Rapid and Quantitative Detection of Gastric Cancer Carbohydrate Antigen 72-4

NASA Astrophysics Data System (ADS)

Yan, Xinyu; Wang, Kan; Lu, Wenting; Qin, Weijian; Cui, Daxiang; He, Jinghua

2016-03-01

Carbohydrate antigen 72-4 (CA72-4) is an important biomarker associated closely with diagnosis and prognosis of early gastric cancer. How to realize quick, sensitive, specific, and quantitative detection of CA72-4 in clinical specimens has become a great requirement. Herein, we reported a CdSe/ZnS quantum dot-labeled lateral flow test strip combined with a charge-coupled device (CCD)-based reader was developed for rapid, sensitive, and quantitative detection of CA72-4. Two mouse monoclonal antibodies (mAbs) against CA72-4 were employed. One of them was coated as a test line, while another mAb was labeled with quantum dots and coated onto conjugate pad. The goat anti-mouse IgG was immobilized as a control line. After sample was added, a sandwich structure was formed with CA72-4 and these two mAbs. The fluorescent signal from quantum dots (QD)-labeled mAb in sandwich structure was related to the amount of detected CA72-4. A CCD-based reader was used to realize quantitative detection of CA72-4. Results showed that developed QD-labeled lateral flow strips to detect CA72-4 biomarker with the sensitivity of 2 IU/mL and 10 min detection time. One hundred sera samples from clinical patients with gastric cancer and healthy people were used to confirm specificity of this strip method; results showed that established strip method own 100 % reproducibility and 100 % specificity compared with Roche electrochemiluminescence assay results. In conclusion, CdSe/ZnS quantum dot-labeled lateral flow strips for detection of CA72-4 could realize rapid, sensitive, and specific detection of clinical samples and could own great potential in clinical translation in near future.

Impact of the rapid antigen detection test in diagnosis and treatment of acute pharyngotonsillitis in a pediatric emergency room.

PubMed

Cardoso, Débora Morais; Gilio, Alfredo Elias; Hsin, Shieh Huei; Machado, Beatriz Marcondes; de Paulis, Milena; Lotufo, João Paulo B; Martinez, Marina Baquerizo; Grisi, Sandra Josefina E

2013-01-01

To evaluate the impact of the routine use of rapid antigen detection test in the diagnosis and treatment of acute pharyngotonsillitis in children. This is a prospective and observational study, with a protocol compliance design established at the Emergency Unit of the University Hospital of Universidade de São Paulo for the care of children and adolescents diagnosed with acute pharyngitis. 650 children and adolescents were enrolled. Based on clinical findings, antibiotics would be prescribed for 389 patients (59.8%); using the rapid antigen detection test, they were prescribed for 286 patients (44.0%). Among the 261 children who would not have received antibiotics based on the clinical evaluation, 111 (42.5%) had positive rapid antigen detection test. The diagnosis based only on clinical evaluation showed 61.1% sensitivity, 47.7% specificity, 44.9% positive predictive value, and 57.5% negative predictive value. The clinical diagnosis of streptococcal pharyngotonsillitis had low sensitivity and specificity. The routine use of rapid antigen detection test led to the reduction of antibiotic use and the identification of a risk group for complications of streptococcal infection, since 42.5% positive rapid antigen detection test patients would not have received antibiotics based only on clinical diagnosis.

Colorimetric analysis of saliva–alcohol test strips by smartphone-based instruments using machine-learning algorithms

USDA-ARS?s Scientific Manuscript database

Strip lateral flow assays, similar to a home pregnancy test, are used widely in food safety applications to provide rapid and accurate tests for the presence of specific foodborne pathogens or other contaminants. Though these tests are very rapid, they are not very sensitive, are not quantitative, a...

Large-scale human immunodeficiency virus rapid test evaluation in a low-prevalence ugandan blood bank population.

PubMed

Eller, Leigh A; Eller, Michael A; Ouma, Benson J; Kataaha, Peter; Bagaya, Bernard S; Olemukan, Robert L; Erima, Simon; Kawala, Lilian; de Souza, Mark S; Kibuuka, Hannah; Wabwire-Mangen, Fred; Peel, Sheila A; O'Connell, Robert J; Robb, Merlin L; Michael, Nelson L

2007-10-01

The use of rapid tests for human immunodeficiency virus (HIV) has become standard in HIV testing algorithms employed in resource-limited settings. We report an extensive HIV rapid test validation study conducted among Ugandan blood bank donors at low risk for HIV infection. The operational characteristics of four readily available commercial HIV rapid test kits were first determined with 940 donor samples and were used to select a serial testing algorithm. Uni-Gold Recombigen HIV was used as the screening test, followed by HIV-1/2 STAT-PAK for reactive samples. OraQuick HIV-1 testing was performed if the first two test results were discordant. This algorithm was then tested with 5,252 blood donor samples, and the results were compared to those of enzyme immunoassays (EIAs) and Western blotting. The unadjusted algorithm sensitivity and specificity were 98.6 and 99.9%, respectively. The adjusted sensitivity and specificity were 100 and 99.96%, respectively. This HIV testing algorithm is a suitable alternative to EIAs and Western blotting for Ugandan blood donors.

Field performance of malaria rapid diagnostic test for the detection of Plasmodium falciparum infection in Odisha State, India.

PubMed

Sahu, S S; Gunasekaran, K; Jambulingam, P

2015-12-01

Rapid diagnostic tests (RDTs) have become an essential surveillance tool in the malaria control programme in India. The current study aimed to assess the performance of ParaHIT-f, a rapid test in diagnosis of Plasmodium falciparum infection through detecting its specific antigen, histidine rich protein 2 (PfHRP-2), in Odisha State, India. The study was undertaken in eight falciparum malaria endemic southern districts of Odisha State. Febrile patients included through active case detection, were diagnosed by Accredited Social Health Activists (ASHAs) for P. falciparum infection using the RDT, ParaHIT-f. The performance of ParaHIT-f was evaluated using microscopy as the gold standard. A total of 1030 febrile patients were screened by both microscopy and the RDT for P. falciparum infection. The sensitivity of ParaHIT-f was 63.6% (95% CI: 56.0-70.6) and specificity was 98.9% (95% CI: 97.9-99.5), with positive and negative predictive values (PPV and NPV) of 92.6% (95% CI: 86.0-96.3) and 93.0% (95% CI: 91.0-94.5), respectively. When related to parasitaemia, the RDT sensitivity was 47.8% at the low parasitaemia of 4 to 40 parasites/µl of blood. The results showed that the performance of the RDT, ParaHIT-f, was not as sensitive as microscopy in detecting true falciparum infections; a high specificity presented a low frequency of false-positive RDT results. t0 he sensitivity of ParaHIT-f was around 60 per cent. It is, therefore, essential to improve the efficiency (sensitivity) of the kit so that the true falciparum infections will not be missed especially in areas where P. falciparum has been the predominant species causing cerebral malaria.

Rapid specific and visible detection of porcine circovirus type 3 using loop-mediated isothermal amplification (LAMP).

PubMed

Zheng, S; Wu, X; Shi, J; Peng, Z; Gao, M; Xin, C; Liu, Y; Wang, S; Xu, S; Han, H; Yu, J; Sun, W; Cong, X; Li, J; Wang, J

2018-06-01

In this study, a rapid and specific assay for the detection of porcine circovirus type 3 (PCV3) was established using loop-mediated isothermal amplification (LAMP). Four primers were specifically designed to amplify PCV3. The LAMP assay was effectively optimized to amplify PCV3 by water bath at 60°C for 60 min. The detection limit was approximately 1 × 10 1 copy in this LAMP assay. Compared to porcine circovirus type 2 (PCV2), both gE and gD genes of pseudorabies virus (PRV) and porcine parvovirus (PPV), the LAMP assay showed a high specific detection of PCV3. A visible detection method was developed using SYBR Green I to recognize the results rapidly. Based on the detection of 20 clinical tissue samples, the LAMP assay was more practical and convenient than classical PCR due to its simplicity, high sensitivity, rapidity, specificity, visibility and cost efficiency. © 2018 Blackwell Verlag GmbH.

A sensitive and specific LC-MS/MS method for rapid diagnosis of Niemann-Pick C1 disease from human plasma[S

PubMed Central

Jiang, Xuntian; Sidhu, Rohini; Porter, Forbes D.; Yanjanin, Nicole M.; Speak, Anneliese O.; te Vruchte, Danielle Taylor; Platt, Frances M.; Fujiwara, Hideji; Scherrer, David E.; Zhang, Jessie; Dietzen, Dennis J.; Schaffer, Jean E.; Ory, Daniel S.

2011-01-01

Niemann-Pick type C1 (NPC1) disease is a rare, progressively fatal neurodegenerative disease for which there are no FDA-approved therapies. A major barrier to developing new therapies for this disorder has been the lack of a sensitive and noninvasive diagnostic test. Recently, we demonstrated that two cholesterol oxidation products, specifically cholestane-3β,5α,6β-triol (3β,5α,6β-triol) and 7-ketocholesterol (7-KC), were markedly increased in the plasma of human NPC1 subjects, suggesting a role for these oxysterols in diagnosis of NPC1 disease and evaluation of therapeutics in clinical trials. In the present study, we describe the development of a sensitive and specific LC-MS/MS method for quantifying 3β,5α,6β-triol and 7-KC human plasma after derivatization with N,N-dimethylglycine. We show that dimethylglycine derivatization successfully enhanced the ionization and fragmentation of 3β,5α,6β-triol and 7-KC for mass spectrometric detection of the oxysterol species in human plasma. The oxysterol dimethylglycinates were resolved with high sensitivity and selectivity, and enabled accurate quantification of 3β,5α,6β-triol and 7-KC concentrations in human plasma. The LC-MS/MS assay was able to discriminate with high sensitivity and specificity between control and NPC1 subjects, and offers for the first time a noninvasive, rapid, and highly sensitive method for diagnosis of NPC1 disease. PMID:21518695

Field evaluation of a malaria rapid diagnostic test (ICT Pf).

PubMed

Moonasar, Devanand; Goga, Ameena E; Kruger, Philip S; La Cock, Christine; Maharaj, Rajendra; Frean, John; Chandramohan, Daniel

2009-11-01

Malaria rapid diagnostic tests (MRDTs) are quick and easy to perform and useful for diagnosing malaria in primary health care settings. In South Africa most malaria infections are due to Plasmodium falciparurrm, and HRPII-based MRDTs have been used since 2001. Previous studies in Africa showed variability in sensitivity and specificity of HRPII-based MRDTs; hence, we conducted a field evaluation in Limpopo province to determine the accuracy of the MRDT currently used in public sector clinics and hospitals. A cross-sectional observational study was conducted to determine the sensitivity and specificity of an ICT Pf MRDT. We tested 405 patients with fever with ICT Pf MRDT and compared the results with blood film microscopy (the gold standard). RESULTS. The overall sensitivity of the ICT Pf MRDT was 99.48% (95% confidence interval (CI) 96.17-100%), while specificity was 96.26% (95% CI 94.7-100%). The positive predictive value of the test was 98.48 (99% CI 98.41-100%), and the negative predictive value was 99.52% (95% CI 96.47-100%). The ICT Pf MRDT is an appropriate test to use in the field in South Africa where laboratory facilities are not available. It has a high degree of sensitivity and acceptable level of specificity in accordance with the World Health Organization criteria. However, sensitivity of MRDT at low levels of parasitaemia (<100 parasites/microl of blood) in field conditions must still be established.

Evaluation of a Cryptococcal antigen Lateral Flow Assay in serum and cerebrospinal fluid for rapid diagnosis of cryptococcosis in Colombia.

PubMed

Cáceres, Diego H; Zuluaga, Alejandra; Tabares, Ángela M; Chiller, Tom; González, Ángel; Gómez, Beatriz L

2017-12-21

A Lateral Flow Assay to detect cryptococcal antigen (CrAg® LFA) in serum and cerebrospinal fluid for the rapid diagnosis of cryptococcosis was evaluated. A retrospective validation was performed. Sensitivity and specificity of the CrAg® LFA was 100%. High concordance (kappa index=1.0) between Cryptococcal Antigen Latex Agglutination System (CALAS®) and CrAg® LFA was observed. CrAg® LFA showed higher analytical sensitivity for detecting low concentrations of cryptococcal antigen.

Evaluation of a Cryptococcal antigen Lateral Flow Assay in serum and cerebrospinal fluid for rapid diagnosis of cryptococcosis in Colombia

PubMed Central

Cáceres, Diego H.; Zuluaga, Alejandra; Tabares, Ángela M.; Chiller, Tom; González, Ángel; Gómez, Beatriz L.

2017-01-01

ABSTRACT A Lateral Flow Assay to detect cryptococcal antigen (CrAg® LFA) in serum and cerebrospinal fluid for the rapid diagnosis of cryptococcosis was evaluated. A retrospective validation was performed. Sensitivity and specificity of the CrAg® LFA was 100%. High concordance (kappa index=1.0) between Cryptococcal Antigen Latex Agglutination System (CALAS®) and CrAg® LFA was observed. CrAg® LFA showed higher analytical sensitivity for detecting low concentrations of cryptococcal antigen. PMID:29267584

Accuracy of Two Malaria Rapid Diagnostic Tests (RDTS) for Initial Diagnosis and Treatment Monitoring in a High Transmission Setting in Uganda

PubMed Central

Mbabazi, Phoebe; Hopkins, Heidi; Osilo, Emmanuel; Kalungu, Michael; Byakika-Kibwika, Pauline; Kamya, Moses R.

2015-01-01

Malaria rapid diagnostic tests (RDTs) may improve fever management in areas without microscopy. We compared the accuracy of histidine-rich protein 2 (HRP2) and Plasmodium lactate dehydrogenase (pLDH)-based RDTs, using expert microscopy as a gold standard, for initial diagnosis, treatment monitoring, and diagnosis of recurrent malaria in a cohort of children followed longitudinally in a high-transmission area in Uganda. For 305 initial fever episodes, sensitivity was 98% for HRP2 and 87% for pLDH, whereas specificity was 55% and 96%, respectively. The HRP2 gave 51% false-positive results on Day 28, whereas pLDH gave no false positives after Day 7. For 59 recurrent fever episodes during follow-up, sensitivity was 100% for HRP2 and 91% for pLDH, whereas specificity was 33% and 100%, respectively. The HRP2-based RDTs are useful for initial diagnosis of malaria caused by superior sensitivity; however, as a result of superior specificity, pLDH-based RDTs are more appropriate to monitor treatment and diagnose recurrent malaria. PMID:25624399

Diagnosis of Normal and Abnormal Color Vision with Cone-Specific VEPs.

PubMed

Rabin, Jeff C; Kryder, Andrew C; Lam, Dan

2016-05-01

Normal color vision depends on normal long wavelength (L), middle wavelength (M), and short wavelength sensitive (S) cones. Hereditary "red-green" color vision deficiency (CVD) is due to a shift in peak sensitivity or lack of L or M cones. Hereditary S cone CVD is rare but can be acquired as an early sign of disease. Current tests detect CVD but few diagnose type or severity, critical for linking performance to real-world demands. The anomaloscope and newer subjective tests quantify CVD but are not applicable to infants or cognitively impaired patients. Our purpose was to develop an objective test of CVD with sensitivity and specificity comparable to current tests. A calibrated visual-evoked potential (VEP) display and Food and Drug Administration-approved system was used to record L, M, and S cone-specific pattern-onset VEPs from 18 color vision normals (CVNs) and 13 hereditary CVDs. VEP amplitudes and latencies were compared between groups to establish VEP sensitivity and specificity. Cone VEPs show 100% sensitivity for diagnosis of CVD and 94% specificity for confirming CVN. L cone (protan) CVDs showed a significant increase in L cone latency (53.1 msec, P < 0.003) and decreased amplitude (10.8 uV, P < 0.0000005) but normal M and S cone VEPs ( P > 0.31). M cone (deutan) CVDs showed a significant increase in M cone latency (31.0 msec, P < 0.000004) and decreased amplitude (8.4 uV, P < 0.006) but normal L and S cone VEPs ( P > 0.29). Cone-specific VEPs offer a rapid, objective test to diagnose hereditary CVD and show potential for detecting acquired CVD in various diseases. This paper describes the efficacy of cone-specific color VEPs for quantification of normal and abnormal color vision. The rapid, objective nature of this approach makes it suitable for detecting color sensitivity loss in infants and the cognitively impaired.

Multiplex PCR for simultaneous identification of E. coli O157:H7, Salmonella spp. and L. monocytogenes in food.

PubMed

Nguyen, Thuy Trang; Van Giau, Vo; Vo, Tuong Kha

2016-12-01

The rapid detection of pathogens in food is becoming increasingly critical for ensuring the safety of consumers, since the majority of food-borne illnesses and deaths are caused by pathogenic bacteria. Hence, rapid, sensitive, inexpensive and convenient approaches to detect food-borne pathogenic bacteria is essential in controlling food safety. In this study, a multiplex PCR assay for the rapid and simultaneous detection of Escherichia coli O157:H7, Salmonella spp. and Listeria monocytogenes was established. The invA, stx and hlyA genes specifically amplified DNA fragments of 284, 404 and 510 bp from Salmonella spp., L. monocytogenes and E. coli O157:H7, respectively. The 16S rRNA gene was targeted as an internal control gene in the presence of bacterial DNA. The specificity and sensitivity of the multiplex PCR were performed by testing different strains. The multiplex PCR assay was able to specifically simultaneously detect ten colony-forming unit/mL of each pathogen in artificially inoculated samples after enrichment for 12 h. The whole process took less than 24 h to complete, indicating that the assay is suitable for reliable and rapid identification of these three food-borne pathogens, which could be suitable in microbial epidemiology investigation.

Evaluation of SD BIOLINE H. pylori Ag rapid test against double ELISA with SD H. pylori Ag ELISA and EZ-STEP H. pylori Ag ELISA tests.

PubMed

Negash, Markos; Kassu, Afework; Amare, Bemnet; Yismaw, Gizachew; Moges, Beyene

2018-01-01

Helicobacter pylori antibody titters fall very slowly even after successful treatment. Therefore, tests detecting H. pylori antibody lack specificity and sensitivity. On the other hand, H. pylori stool antigen tests are reported as an alternative assay because of their reliability and simplicity. However, the comparative performance of H. pylori stool antigen tests for detecting the presence of the bacterium in clinical specimens in the study area is not assessed. Therefore, in this study we evaluated the performance of SD BIOLINE H. pylori Ag rapid test with reference to the commercially available EZ- STEP ELISA and SD BIOLINE H. pylori Ag ELISA tests. Stool samples were collected to analyse the diagnostic performance of SD BIOLINE H. pylori Ag rapid test kit using SD H. pylori Ag ELISA kit and EZ- STEP ELISA tests as a gold standard. Serum samples were also collected from each patient to test for the presence of H. pylori antibodies using dBest H. pylori Test Disk. Sensitivity, specificity, predictive values and kappa value are assessed. P values < 0.05 were taken statistically significant. Stool and serum samples were collected from 201 dyspeptic patients and analysed. The sensitivity, specificity, positive and negative predictive values of the SD BIOLINE H. pylori Ag rapid test were: 95.6% (95% CI, 88.8-98.8), 92.5% (95%CI, 89-94.1%), 86.7% (95% CI, 80.5-89.6), and 97.6% (95% CI, 993.9-99.3) respectively. The performance of SD BIOLINE H. pylori Ag rapid test was better than the currently available antibody test in study area. Therefore, the SD BIOLINE Ag rapid stool test could replace and be used to diagnose active H. pylori infection before the commencement of therapy among dyspeptic patients.

Nucleophile sensitivity of Drosophila TRPA1 underlies light-induced feeding deterrence

PubMed Central

Du, Eun Jo; Ahn, Tae Jung; Wen, Xianlan; Seo, Dae-Won; Na, Duk L; Kwon, Jae Young; Choi, Myunghwan; Kim, Hyung-Wook; Cho, Hana; Kang, KyeongJin

2016-01-01

Solar irradiation including ultraviolet (UV) light causes tissue damage by generating reactive free radicals that can be electrophilic or nucleophilic due to unpaired electrons. Little is known about how free radicals induced by natural sunlight are rapidly detected and avoided by animals. We discover that Drosophila Transient Receptor Potential Ankyrin 1 (TRPA1), previously known only as an electrophile receptor, sensitively detects photochemically active sunlight through nucleophile sensitivity. Rapid light-dependent feeding deterrence in Drosophila was mediated only by the TRPA1(A) isoform, despite the TRPA1(A) and TRPA1(B) isoforms having similar electrophile sensitivities. Such isoform dependence re-emerges in the detection of structurally varied nucleophilic compounds and nucleophilicity-accompanying hydrogen peroxide (H2O2). Furthermore, these isoform-dependent mechanisms require a common set of TRPA1(A)-specific residues dispensable for electrophile detection. Collectively, TRPA1(A) rapidly responds to natural sunlight intensities through its nucleophile sensitivity as a receptor of photochemically generated radicals, leading to an acute light-induced behavioral shift in Drosophila. DOI: http://dx.doi.org/10.7554/eLife.18425.001 PMID:27656903

Rapid assessment method for prevalence and intensity of Loa loa infection.

PubMed Central

Takougang, Innocent; Meremikwu, Martin; Wandji, Samuel; Yenshu, Emmanuel V.; Aripko, Ben; Lamlenn, Samson B.; Eka, Braide L.; Enyong, Peter; Meli, Jean; Kale, Oladele; Remme, Jan H.

2002-01-01

OBJECTIVE: To assess the validity of observations on eye worm and Calabar swellings for the rapid assessment of the prevalence and intensity of loiasis at the community level. METHOD: A total of 12895 individuals over the age of 15 years living in 102 communities in Cameroon and Nigeria took part in the study. A standardized questionnaire was administered to participants from whom finger-prick blood samples were collected and examined for Loa loa microfilariae. Rapid assessments of the prevalence and intensity of loiasis were made on the basis of a history of eye worm or Calabar swellings. FINDINGS: There was a strong correlation between the indices of the rapid assessment procedures and the parasitological indices of L. loa endemicity. The rapid assessment indices were effective in diagnosing high-risk communities (sensitivity 94-100%; specificity 66-92%). The highest sensitivity (100%) and specificity (92%) were obtained with a rapid assessment procedure based on a history of eye worm lasting 1-7 days together with confirmation by the guided recognition of a photograph of adult L. loa in the eye. CONCLUSION: Rapid assessment of the prevalence and intensity of loiasis at the community level can be achieved using a procedure based on the history of eye worm lasting 1-7 days together with confirmation by the guided recognition of a photograph of an adult L. loa in the eye. PMID:12481206

Rapid, Sensitive, Enzyme-Immunodotting Assay for Detecting Cow Milk Adulteration in Sheep Milk: A Modern Laboratory Project

NASA Astrophysics Data System (ADS)

Inda, Luis A.; Razquín, Pedro; Lampreave, Fermín; Alava, María A.; Calvo, Miguel

1998-12-01

Specificity, sensitivity, and experimental simplicity make the immunoenzymatic assay suitable for a variety of laboratories dedicated to diverse activities such as research, quality control in food analysis, or clinical biochemistry. In these assays, the antibody that specifically recognizes the antigen is covalently attached to an enzyme. Once the antigen-antibody immunocomplex is formed, the enzymatic reaction gives a colored product that allows the detection of the initial antigen. The aim of this work was the design of a new laboratory project appropriate for use in courses of biochemistry, immunochemistry, or analytical chemistry. The assay described here detects the presence of cow milk in milk of other species. The main application is the detection of cow milk in sheep milk and cheese. Specific proteins, immunoglobulins (IgG) of the fraudulent bovine milk, are specifically recognized and retained by antibodies immobilized on a membrane. The binding of a second antibody covalently attached to horseradish peroxidase (HRP) allows the development of a visible signal. Thus, students can rapidly detect milk adulterations using a specific, sensitive, and safe experimental approach. The experiment allows students to apply their theoretical knowledge, resulting in a stimulating experience of solving a real problem during a 4-hour laboratory period.

Multiplex polymerase chain reaction assay developed to diagnose adult bacterial meningitis in Taiwan.

PubMed

Lee, Chi-Tsung; Hsiao, Kuang-Ming; Chen, Jin-Cherng; Su, Cheng-Chuan

2015-11-01

Acute bacterial meningitis causes high morbidity and mortality; the associated clinical symptoms often are insensitive or non-specific; and the pathogenic bacteria are geographically diverse. Clinical diagnosis requires a rapid and accurate methodology. This study aimed to develop a new multiplex polymerase chain reaction (mPCR) assay to detect simultaneously six major bacteria that cause adult bacterial meningitis in Taiwan: Klebsiella pneumoniae, Pseudomonas aeruginosa, Streptococcus pneumoniae, Staphylococcus aureus, Escherichia coli, and Acinetobacter baumannii. Species-specific primers for the six bacteria were developed using reference strains. The specificities of the mPCRs for these bacteria were validated, and the sensitivities were evaluated via serial dilutions. The mPCR assay specifically detected all of the six pathogens, particularly with sensitivities of 12 colony forming units (CFU)/mL, 90 CFU/mL, and 390 CFU/mL for E. coli, S. pneumoniae, and K. pneumoniae, respectively. This mPCR assay is a rapid and specific tool to detect the six major bacterial pathogens that cause acute adult meningitis in Taiwan, particularly sensitive for detecting E. coli, S. pneumoniae, and K. pneumoniae. The assay may facilitate early diagnosis and guidance for antimicrobial therapy for adult patients with this deadly disease in Taiwan. © 2015 APMIS. Published by John Wiley & Sons Ltd.

Recombinase polymerase amplification combined with a lateral flow dipstick for rapid and visual detection of Schistosoma japonicum.

PubMed

Sun, Kui; Xing, Weiwei; Yu, Xinling; Fu, Wenliang; Wang, Yuanyuan; Zou, Minji; Luo, Zhihong; Xu, Donggang

2016-08-31

With the continuous decline in prevalence and intensity of Schistosoma japonicum infection in China, more accurate and sensitive methods suitable for field detection become much needed for schistosomiasis control. Here, a novel rapid and visual detection method based on the combination of recombinase polymerase amplification (RPA) and lateral flow dipstick (LFD) was developed to detect S. japonicum DNA in fecal samples. The LFD-RPA assay targeting SjR2 could detect 5 fg S. japonicum DNA, which was identical to qPCR and real-time RPA assay, and showed no cross-reaction with other parasites. The detection could be finished within 15-20 min at a wide temperature range (25-45 °C), and the results could be visualized by naked eye. The diagnostic validity of LFD-RPA assay was further assessed with 14 fecal samples of infected patients diagnosed by Kato-Katz method and 31 fecal samples of healthy persons, and compared with that of Enzyme-linked immunosorbent assay (ELSIA) and Indirect Hemagglutination Assay (IHA). The LFD-RPA assay showed 92.68 % sensitivity, 100 % specificity and excellent diagnostic agreement with the gold standard Kato-Katz test (k = 0.947, Z = 6.36, P < 0.001), whereas ELISA showed 85.71 % sensitivity, 93.55 % specificity, and substantial diagnostic agreement (k = 0.793, Z = 5.31, P < 0.001), and IHA showed 78.57 % sensitivity, 83.87 % specificity, and moderate diagnostic agreement (k = 0.600, Z = 4.05, P < 0.001), indicating that the LFD-RPA was much better than the traditional methods. The LFD-RPA assay established by us is a sensitive, specific, rapid and convenient method for the diagnosis of schistosomiasis, and shows a great potency in field application.

Multistix 10 SG Leukocyte Esterage Dipstick Testing in Rapid Bedside Diagnosis of Spontaneous Bacterial Peritonitis: A Prospective Study.

PubMed

Jha, Ashish K; Kumawat, Dal C; Bolya, Yasvant K; Goenka, Mahesh K

2012-09-01

Spontaneous bacterial peritonitis (SBP) requires rapid diagnosis and the initiation of antibiotics. Diagnosis of SBP is usually based on cytobacteriological examination of ascitic fluid. These tests require good laboratory facilities and reporting time of few hours to 1-2 day. However, the 24 h laboratory facilities not widely available in country like India. We evaluated the diagnostic utility of reagent strip (Multistix 10 SG(®)) for rapid diagnosis of SBP. The study was prospectively carried out on patients of cirrhosis with ascites. Bedside leukocyte esterase reagent strip testing was performed on ascitic fluid. Cell count as determined by colorimetric scale of reagent strip was compared with counting chamber method. Sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were calculated. Out of 100 patients with cirrhotic ascites, [72 males: 28 female; mean age 44.34 (SD 13.03) years] 18 patients were diagnosed to have SBP by counting chamber method as compared to 14 patients detected to have SBP by reagent strip test ≥++ positive. The sensitivity, specificity, positive predictive value, negative predictive value and accuracy of reagent strip ≥++ positive were 77.77%, 95.12%, 77.77%, 95.12% and 92% respectively compared to counting chamber method. Reagent strip to diagnose SBP is very specific but less sensitive as compared to counting chamber method. This can be performed rapidly, easily and efficiently even in remote area of developing countries. This bedside test could be a useful tool for the diagnosis of SBP in country like India.

Rapid diagnosis of sepsis with TaqMan-Based multiplex real-time PCR.

PubMed

Liu, Chang-Feng; Shi, Xin-Ping; Chen, Yun; Jin, Ye; Zhang, Bing

2018-02-01

The survival rate of septic patients mainly depends on a rapid and reliable diagnosis. A rapid, broad range, specific and sensitive quantitative diagnostic test is the urgent need. Thus, we developed a TaqMan-Based Multiplex real-time PCR assays to identify bloodstream pathogens within a few hours. Primers and TaqMan probes were designed to be complementary to conserved regions in the 16S rDNA gene of different kinds of bacteria. To evaluate accurately, sensitively, and specifically, the known bacteria samples (Standard strains, whole blood samples) are determined by TaqMan-Based Multiplex real-time PCR. In addition, 30 blood samples taken from patients with clinical symptoms of sepsis were tested by TaqMan-Based Multiplex real-time PCR and blood culture. The mean frequency of positive for Multiplex real-time PCR was 96% at a concentration of 100 CFU/mL, and it was 100% at a concentration greater than 1000 CFU/mL. All the known blood samples and Standard strains were detected positively by TaqMan-Based Multiplex PCR, no PCR products were detected when DNAs from other bacterium were used in the multiplex assay. Among the 30 patients with clinical symptoms of sepsis, 18 patients were confirmed positive by Multiplex real-time PCR and seven patients were confirmed positive by blood culture. TaqMan-Based Multiplex real-time PCR assay with highly sensitivity, specificity and broad detection range, is a rapid and accurate method in the detection of bacterial pathogens of sepsis and should have a promising usage in the diagnosis of sepsis. © 2017 Wiley Periodicals, Inc.

Novel high-speed droplet-allele specific-polymerase chain reaction: application in the rapid genotyping of single nucleotide polymorphisms.

PubMed

Taira, Chiaki; Matsuda, Kazuyuki; Yamaguchi, Akemi; Sueki, Akane; Koeda, Hiroshi; Takagi, Fumio; Kobayashi, Yukihiro; Sugano, Mitsutoshi; Honda, Takayuki

2013-09-23

Single nucleotide alterations such as single nucleotide polymorphisms (SNP) and single nucleotide mutations are associated with responses to drugs and predisposition to several diseases, and they contribute to the pathogenesis of malignancies. We developed a rapid genotyping assay based on the allele-specific polymerase chain reaction (AS-PCR) with our droplet-PCR machine (droplet-AS-PCR). Using 8 SNP loci, we evaluated the specificity and sensitivity of droplet-AS-PCR. Buccal cells were pretreated with proteinase K and subjected directly to the droplet-AS-PCR without DNA extraction. The genotypes determined using the droplet-AS-PCR were then compared with those obtained by direct sequencing. Specific PCR amplifications for the 8 SNP loci were detected, and the detection limit of the droplet-AS-PCR was found to be 0.1-5.0% by dilution experiments. Droplet-AS-PCR provided specific amplification when using buccal cells, and all the genotypes determined within 9 min were consistent with those obtained by direct sequencing. Our novel droplet-AS-PCR assay enabled high-speed amplification retaining specificity and sensitivity and provided ultra-rapid genotyping. Crude samples such as buccal cells were available for the droplet-AS-PCR assay, resulting in the reduction of the total analysis time. Droplet-AS-PCR may therefore be useful for genotyping or the detection of single nucleotide alterations. Copyright © 2013 Elsevier B.V. All rights reserved.

[Early diagnosis of streptococcal pharyngitis in paediatric practice: Validity of a rapid antigen detection test].

PubMed

Flores Mateo, Gemma; Conejero, Jaume; Grenzner Martinel, Elisabet; Baba, Zeki; Dicono, Susana; Echasabal, Mildrey; Gonzalo Santos, Concepción; Aliaga, Arantxa; Barredo, María; Ruiz, Luis; Carrau, Montserrat

2010-07-01

To determine the validity of the rapid antigen test for the diagnoses of acute pharyngitis caused by group A beta-haemolytic Streptococcus (GABHS) compared with culture. Observational study of a consecutive sample of paediatric patients. Two primary care centres (PCC) from the metropolitan area of Barcelona. Children aged 1-14 years with sore throat of no more than 5 days duration were chosen at PCC. Oropharyngeal samples were collected from tonsillar bed and posterior pharynx. A rapid diagnostic test was performed, as well as a throat culture. A total of 211 patients were studied. The overall prevalence of pharyngitis due to Streptococcus was 34.1%. Compared with the throat culture, the sensitivity of the rapid test was 90.3% (95% CI: 81.0-96.0), the specificity was 78.4% (95% CI: 70.6-84.9). The percentage of false negatives was 9.7% and the false positives was 21.6%. Spectrum bias was present, inasmuch as the rapid test sensitivity increased with Centor scores. The diagnostic value of a rapid antigen test for the diagnosis of streptococcal pharyngitis in paediatric patients at PCC is high. However, the percentage of false positives and negatives is too high, and also the sensitivity is too low in patients with fewer symptoms to support the use of rapid antigenic test without culture confirmation and bacterial sensitivity test. 2009 Elsevier España, S.L. All rights reserved.

The coeruleus/subcoeruleus complex in idiopathic rapid eye movement sleep behaviour disorder.

PubMed

Ehrminger, Mickael; Latimier, Alice; Pyatigorskaya, Nadya; Garcia-Lorenzo, Daniel; Leu-Semenescu, Smaranda; Vidailhet, Marie; Lehericy, Stéphane; Arnulf, Isabelle

2016-04-01

Idiopathic rapid eye movement sleep behaviour disorder is characterized by nocturnal violence, increased muscle tone during rapid eye movement sleep and the lack of any other neurological disease. However, idiopathic rapid eye movement sleep behaviour disorder can precede parkinsonism and dementia by several years. Using 3 T magnetic resonance imaging and neuromelanin-sensitive sequences, we previously found that the signal intensity was reduced in the locus coeruleus/subcoeruleus area of patients with Parkinson's disease and rapid eye movement sleep behaviour disorder. Here, we studied the integrity of the locus coeruleus/subcoeruleus complex with neuromelanin-sensitive imaging in 21 patients with idiopathic rapid eye movement sleep behaviour disorder and compared the results with those from 21 age- and gender-matched healthy volunteers. All subjects underwent a clinical examination, motor, cognitive, autonomous, psychological, olfactory and colour vision tests, and rapid eye movement sleep characterization using video-polysomnography and 3 T magnetic resonance imaging. The patients more frequently had preclinical markers of alpha-synucleinopathies, including constipation, olfactory deficits, orthostatic hypotension, and subtle motor impairment. Using neuromelanin-sensitive imaging, reduced signal intensity was identified in the locus coeruleus/subcoeruleus complex of the patients with idiopathic rapid eye movement sleep behaviour. The mean sensitivity of the visual analyses of the signal performed by neuroradiologists who were blind to the clinical diagnoses was 82.5%, and the specificity was 81% for the identification of idiopathic rapid eye movement sleep behaviour. The results confirm that this complex is affected in idiopathic rapid eye movement sleep behaviour (to the same degree as it is affected in Parkinson's disease). Neuromelanin-sensitive imaging provides an early marker of non-dopaminergic alpha-synucleinopathy that can be detected on an individual basis. © The Author (2016). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Development and validation of a quantitative PCR for rapid and specific detection of California sea lion adenovirus 1 and prevalence in wild and managed populations.

PubMed

Cortés-Hinojosa, Galaxia; Gulland, Frances M D; Goldstein, Tracey; Venn-Watson, Stephanie; Rivera, Rebecca; Archer, Linda L; Waltzek, Thomas B; Gray, Gregory C; Wellehan, James F X

2017-03-01

California sea lion adenovirus 1 (CSLAdV-1) has been associated with hepatitis and enteritis in several wild and captive populations of diverse pinniped species. Currently available tests have been limited to pan-adenoviral polymerase chain reaction (PCR) followed by sequencing. We present the development of a quantitative probe-hybridization PCR (qPCR) assay for rapid, sensitive, and specific detection of this virus in California sea lions ( Zalophus californianus) and other pinnipeds. This assay did not amplify other mammalian adenoviruses and is able to detect consistently down to 10 viral copies per well. Compared with the gold standard conventional pan-adenovirus PCR/sequencing assay, diagnostic sensitivity and specificity of 100% and 88.2% were found, respectively. The lower diagnostic specificity of this qPCR assay may be the result of the lower limit of detection of this assay compared with the gold standard rather than the result of detection of true false-positives.

Rapid and sensitive identification of the herbal tea ingredient Taraxacum formosanum using loop-mediated isothermal amplification.

PubMed

Lai, Guan-Hua; Chao, Jung; Lin, Ming-Kuem; Chang, Wen-Te; Peng, Wen-Huang; Sun, Fang-Chun; Lee, Meng-Shiunn; Lee, Meng-Shiou

2015-01-09

Taraxacum formosanum (TF) is a medicinal plant used as an important component of health drinks in Taiwan. In this study, a rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) assay for authenticating TF was established. A set of four specific LAMP primers was designed based on the nucleotide sequence of the internal transcribed spacer 2 (ITS2) nuclear ribosomal DNA (nrDNA) of TF. LAMP amplicons were successfully amplified and detected when purified genomic DNA of TF was added in the LAMP reaction under isothermal condition (65 °C) within 45 min. These specific LAMP primers have high specificity and can accurately discriminate Taraxacum formosanum from other adulterant plants; 1 pg of genomic DNA was determined to be the detection limit of the LAMP assay. In conclusion, using this novel approach, TF and its misused plant samples obtained from herbal tea markets were easily identified and discriminated by LAMP assay for quality control.

Rapid and Sensitive Identification of the Herbal Tea Ingredient Taraxacum formosanum Using Loop-Mediated Isothermal Amplification

PubMed Central

Lai, Guan-Hua; Chao, Jung; Lin, Ming-Kuem; Chang, Wen-Te; Peng, Wen-Huang; Sun, Fang-Chun; Lee, Meng-Shiunn; Lee, Meng-Shiou

2015-01-01

Taraxacum formosanum (TF) is a medicinal plant used as an important component of health drinks in Taiwan. In this study, a rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) assay for authenticating TF was established. A set of four specific LAMP primers was designed based on the nucleotide sequence of the internal transcribed spacer 2 (ITS2) nuclear ribosomal DNA (nrDNA) of TF. LAMP amplicons were successfully amplified and detected when purified genomic DNA of TF was added in the LAMP reaction under isothermal condition (65 °C) within 45 min. These specific LAMP primers have high specificity and can accurately discriminate Taraxacum formosanum from other adulterant plants; 1 pg of genomic DNA was determined to be the detection limit of the LAMP assay. In conclusion, using this novel approach, TF and its misused plant samples obtained from herbal tea markets were easily identified and discriminated by LAMP assay for quality control. PMID:25584616

Sonophoresis for Rapid Assessment of Interstitial Fluid and Drug Delivery

DTIC Science & Technology

2007-10-01

sensitive indicator of local diseases, for example skin cancer, psoriasis and eczema , but also of certain systemic diseases such as cardiovascular disease...cytokine " functionality -map" that precisely represents skin’s specific diseased milieu (Fig. 2b-CI and Fig. 3a-AD and PS, 3 additional data in. Supp...yielded a unique cytokine " functionality map", which correlates with the specific diseased state (figure 2d and 3a). A striking, rapid upregulation was

Horseradish peroxidase-labeled oligonucleotides and fluorescent tyramides for rapid detection of chromosome-specific repeat sequences.

PubMed

van Gijlswijk, R P; Wiegant, J; Vervenne, R; Lasan, R; Tanke, H J; Raap, A K

1996-01-01

We present a sensitive and rapid fluorescence in situ hybridization (FISH) strategy for detecting chromosome-specific repeat sequences. It uses horseradish peroxidase (HRP)-labeled oligonucleotide sequences in combination with fluorescent tyramide-based detection. After in situ hybridization, the HRP conjugated to the oligonucleotide probe is used to deposit fluorescently labeled tyramide molecules at the site of hybridization. The method features full chemical synthesis of probes, strong FISH signals, and short processing periods, as well as multicolor capabilities.

Development and evaluation of a polydiacetylene based biosensor for the detection of H5 influenza virus.

PubMed

Jiang, Lixiang; Luo, Jing; Dong, Wenjie; Wang, Chengmin; Jin, Wen; Xia, Yuetong; Wang, Haijing; Ding, Hua; Jiang, Long; He, Hongxuan

2015-07-01

H5N1 avian influenza has caused serious economic losses as well as posed significant threats to public health, agriculture and wildlife. It is important to develop a rapid, sensitive and specific detection platform suitable for disease surveillance and control. In this study, a highly sensitive, specific and rapid biosensor based on polydiacetylene was developed for detecting H5 influenza virus. The polydiacetylene based biosensor was produced from an optimized ratio of 10,12-pentacosadiynoic acid and 1,2-dimyristoyl-sn-glycero-3-phosphocholine, with the anti-H5 influenza antibody embedded onto the vesicle surface. The optimized polydiacetylene vesicle could detect H5 influenza virus sensitively with a detection limit of 0.53 copies/μL, showing a dramatic blue-to-red color change that can be observed directly by the naked eye and recorded by a UV-vis spectrometer. The sensitivity, specificity and accuracy of the biosensor were also evaluated. The sensor could specifically differentiate H5 influenza virus from H3 influenza virus, Newcastle disease virus and porcine reproductive and respiratory syndrome virus. Detection using tracheal swabs was in accord with virus isolation results, and comparable to the RT-PCR method. These results offer the possibility and potential of simple polydiacetylene based bio-analytical method for influenza surveillance. Copyright © 2015 Elsevier B.V. All rights reserved.

Development of a rapid and visual nucleotide detection method for a Chinese epidemic strain of Orientia tsutsugamushi based on recombinase polymerase amplification assay and lateral flow test.

PubMed

Qi, Yong; Yin, Qiong; Shao, Yinxiu; Cao, Min; Li, Suqin; Chen, Hongxia; Shen, Wanpeng; Rao, Jixian; Li, Jiameng; Li, Xiaoling; Sun, Yu; Lin, Yu; Deng, Yi; Zeng, Wenwen; Zheng, Shulong; Liu, Suyun; Li, Yuexi

2018-05-01

Orientia tsutsugamushi is an obligate intracellular pathogen that causes scrub typhus. Diagnosing scrub typhus remains a challenge, and a sensitive, specific, simple, and rapid diagnostic test is still needed. A recombinase polymerase amplification (RPA) assay combined with a lateral flow (LF) test targeting the 56-kDa gene of a Karp-like strain of O. tsutsugamushi was developed and optimized. The detection limits, sensitivity, specificity, and simulative clinical performance were evaluated. Primers and probe were screened to establish the RPA assay, and the reaction conditions were optimized. The detection limit was 10 copies/reaction in detecting plasmid DNA and 12 copies/reaction in detecting genomic DNA. The RPA-LF method could differentiate O. tsutsugamushi from other phylogenetically related bacteria. The sensitivity was 100% and specificity was over 90%, when evaluated using infected animal samples or simulative clinical samples. Furthermore, the method was completed in 20min at 37°C followed by a 3-5min incubation at room temperature for the development of an immunochromatographic strip, and the results could be determined visually. This method is promising for wide-ranging use in basic medical units considering that it requires minimal instruments and infrastructure and is highly time-efficient, sensitive, and specific for diagnosing scrub typhus. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Establishment and application of cross-priming isothermal amplification coupled with lateral flow dipstick (CPA-LFD) for rapid and specific detection of red-spotted grouper nervous necrosis virus.

PubMed

Su, Zi Dan; Shi, Cheng Yin; Huang, Jie; Shen, Gui Ming; Li, Jin; Wang, Sheng Qiang; Fan, Chao

2015-09-26

Red-spotted grouper nervous necrosis virus (RGNNV) is an important pathogen that causes diseases in many species of fish in marine aquaculture. The larvae and juveniles are more easily infected by RGNNV and the cumulative mortality is as high as 100 % after being infected with RGNNV. This virus imposes a serious threat to aquaculture of grouper fry. This study aimed to establish a simple, accurate and highly sensitive method for rapid detection of RGNNV on the spot. In this study, the primers specifically targeting RGNNV were designed and cross-priming isothermal amplification (CPA) system was established. The product amplified by CPA was detected through visualization with lateral flow dipstick (LFD). Three important parameters, including the amplification temperature, the concentration of dNTPs and the concentration of Mg(2+) for the CPA system, were optimized. The sensitivity and specificity of this method for RGNNV were tested and compared with those of the conventional RT-PCR and real-time quantitative RT-PCR (qRT-PCR). The optimized conditions for the CPA amplification system were determined as follows: the optimal amplification temperature, the optimized concentration of dNTPs and the concentration for Mg(2+) were 69 °C, 1.2 mmol/L and 5 mmol/L, respectively. The lowest limit of detection (LLOD) of this method for RGNNV was 10(1) copies/μL of RNA sample, which was 10 times lower than that of conventional RT-PCR and comparable to that of RT-qPCR. This method was specific for RGNNV in combination with SJNNV and had no cross-reactions with 8 types of virus and bacterial strains tested. This method was successfully applied to detect RGNNV in fish samples. This study established a CPA-LFD method for detection of RGNNV. This method is simple and rapid with high sensitivity and good specificity and can be widely applied for rapid detection of this virus on the spot.

Utility of dengue NS1 antigen rapid diagnostic test for use in difficult to reach areas and its comparison with dengue NS1 ELISA and qRT-PCR.

PubMed

Shukla, Mohan K; Singh, Neeru; Sharma, Ravendra K; Barde, Pradip V

2017-07-01

The objective of this study was to demonstrate the utility of dengue virus (DENV) non structural protein 1 (NS1) based rapid diagnostic test (RDT) for use in tribal and difficult to reach areas for early dengue (DEN) diagnosis in acute phase patients and evaluate its sensitivity and specificity against DENV NS1 enzyme linked immune sorbent assay (ELISA) and real time reverse transcriptase polymerase chain reaction (qRT-PCR). The DENV NS1 RDT was used for preliminary diagnosis during outbreaks in difficult to reach rural and tribal areas. The diagnosis was confirmed by DENV NS1 ELISA in the laboratory. The samples were also tested and serotyped by qRT-PCR. The results were evaluated using statistical tests. The DENV NS1 RDT showed 99.2% sensitivity and 96.0% specificity when analyzed using DENV NS1 ELISA as standard. The specificity and sensitivity of the RDT when compared with qRT-PCR was 93.6% and 91.1%, respectively. The serotype specific evaluation showed more than 90% sensitivity and specificity for DENV-1, 2, and 3. The RDT proved a good diagnostic tool in difficult to reach rural and tribal areas. Further evaluation studies with different commercially available RDTs in different field conditions are essential, that will help clinicians and patients for treatment and programme managers for timely intervention. © 2017 Wiley Periodicals, Inc.

Using Reference Quantitative Polymerase Chain Reaction to Assess the Clinical Performance of the Paracheck-Pf® Rapid Diagnostic Test in a Field Setting in Uganda.

PubMed

Mitran, Catherine J; Mbonye, Anthony K; Hawkes, Michael; Yanow, Stephanie K

2018-06-04

Malaria rapid diagnostic tests (RDTs) are widely used in clinical and surveillance settings. However, the performance of most RDTs has not been characterized at parasite densities below detection by microscopy. We present findings from Uganda, where RDT results from 491 participants with suspected malaria were correlated with quantitative polymerase chain reaction (qPCR)-defined parasitemia. Compared with qPCR, the sensitivity and specificity of the RDT for Plasmodium falciparum mono-infections were 76% (95% confidence interval [CI]: 68-83%) and 95% (95% CI: 92-97%), respectively. The sensitivity of the RDT at parasite densities between 0.2 and 200 parasites/μL was surprisingly high (87%, 95% CI: 74-94%). The high sensitivity of the RDT is likely because of histidine-rich protein 2 from submicroscopic infections, gametocytes, or sequestered parasites. These findings underscore the importance of evaluating different RDTs in field studies against qPCR reference testing to better define the sensitivity and specificity, particularly at low parasite densities.

Use of a Rapid Test for Diagnosis of Dengue during Suspected Dengue Outbreaks in Resource-Limited Regions.

PubMed

Hunsperger, Elizabeth A; Sharp, Tyler M; Lalita, Paul; Tikomaidraubuta, Kini; Cardoso, Yolanda Rebello; Naivalu, Taina; Khan, Aalisha Sahu; Marfel, Maria; Hancock, W Thane; Tomashek, Kay M; Margolis, Harold S

2016-08-01

Dengue is major public health problem, globally. Timely verification of suspected dengue outbreaks allows for public health response, leading to the initiation of appropriate clinical care. Because the clinical presentation of dengue is nonspecific, dengue diagnosis would benefit from a sensitive rapid diagnostic test (RDT). We evaluated the diagnostic performance of an RDT that detects dengue virus (DENV) nonstructural protein 1 (NS1) and anti-DENV IgM during suspected acute febrile illness (AFI) outbreaks in four countries. Real-time reverse transcription-PCR and anti-DENV IgM enzyme-linked immunosorbent assay were used to verify RDT results. Anti-DENV IgM RDT sensitivity and specificity ranged from 55.3 to 91.7% and 85.3 to 98.5%, respectively, and NS1 sensitivity and specificity ranged from 49.7 to 92.9% and 22.2 to 89.0%, respectively. Sensitivity varied by timing of specimen collection and DENV serotype. Combined test results moderately improved the sensitivity. The use of RDTs identified dengue as the cause of AFI outbreaks where reference diagnostic testing was limited or unavailable. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Evaluation of a Rapid Diagnostic Test for Yaws Infection in a Community Surveillance Setting

PubMed Central

Marks, Michael; Goncalves, Adriana; Vahi, Ventis; Sokana, Oliver; Puiahi, Elliot; Zhang, Zaixing; Dalipanda, Tenneth; Bottomley, Christian; Mabey, David; Solomon, Anthony W.

2014-01-01

Yaws is a non-venereal treponemal infection caused by Treponema pallidum ssp. pertenue. The WHO has launched a worldwide control programme, which aims to eradicate yaws by 2020. The development of a rapid diagnostic test (RDT) for serological diagnosis in the isolated communities affected by yaws is a key requirement for the successful implementation of the WHO strategy. We conducted a study to evaluate the utility of the DPP test in screening for yaws, utilizing samples collected as part of a community prevalence survey conducted in the Solomon Islands. 415 serum samples were tested using both traditional syphilis serology (TPPA and quantitative RPR) and the Chembio DPP Syphilis Screen and Confirm RDT. We calculated the sensitivity and specificity of the RDT as compared to gold standard serology. The sensitivity of the RDT against TPPA was 58.5% and the specificity was 97.6%. The sensitivity of the RDT against RPR was 41.7% and the specificity was 95.2%. The sensitivity of the DPP was strongly related to the RPR titre with a sensitivity of 92.0% for an RPR titre of >1/16. Wider access to DPP testing would improve our understanding of worldwide yaws case reporting and the test may play a key role in assessing patients presenting with yaws like lesions in a post-mass drug administration (MDA) setting. PMID:25211018

Evaluation of a rapid diagnostic test for yaws infection in a community surveillance setting.

PubMed

Marks, Michael; Goncalves, Adriana; Vahi, Ventis; Sokana, Oliver; Puiahi, Elliot; Zhang, Zaixing; Dalipanda, Tenneth; Bottomley, Christian; Mabey, David; Solomon, Anthony W

2014-09-01

Yaws is a non-venereal treponemal infection caused by Treponema pallidum ssp. pertenue. The WHO has launched a worldwide control programme, which aims to eradicate yaws by 2020. The development of a rapid diagnostic test (RDT) for serological diagnosis in the isolated communities affected by yaws is a key requirement for the successful implementation of the WHO strategy. We conducted a study to evaluate the utility of the DPP test in screening for yaws, utilizing samples collected as part of a community prevalence survey conducted in the Solomon Islands. 415 serum samples were tested using both traditional syphilis serology (TPPA and quantitative RPR) and the Chembio DPP Syphilis Screen and Confirm RDT. We calculated the sensitivity and specificity of the RDT as compared to gold standard serology. The sensitivity of the RDT against TPPA was 58.5% and the specificity was 97.6%. The sensitivity of the RDT against RPR was 41.7% and the specificity was 95.2%. The sensitivity of the DPP was strongly related to the RPR titre with a sensitivity of 92.0% for an RPR titre of >1/16. Wider access to DPP testing would improve our understanding of worldwide yaws case reporting and the test may play a key role in assessing patients presenting with yaws like lesions in a post-mass drug administration (MDA) setting.

Clinical Evaluation of Anti-Tuberculous Glycolipid Immunoglobulin G Antibody Assay for Rapid Serodiagnosis of Pulmonary Tuberculosis

PubMed Central

Maekura, Ryoji; Okuda, Yoshinari; Nakagawa, Masaru; Hiraga, Touru; Yokota, Souichirou; Ito, Masami; Yano, Ikuya; Kohno, Hiroaki; Wada, Masako; Abe, Chiyoji; Toyoda, Takeo; Kishimoto, Toshio; Ogura, Takeshi

2001-01-01

Previously we reported the development of a highly sensitive enzyme-linked immunosorbent assay specific for anti-tuberculous glycolipid (anti-TBGL) for the rapid serodiagnosis of tuberculosis. In this study, the usefulness of an anti-TBGL antibody assay kit for rapid serodiagnosis was evaluated in a controlled multicenter study. Antibody titers in sera from 318 patients with active pulmonary tuberculosis (216 positive for Mycobacterium tuberculosis in smear and/or culture tests and 102 smear and culture negative and clinically diagnosed), 58 patients with old tuberculosis, 177 patients with other respiratory diseases, 156 patients with nonrespiratory diseases, and 454 healthy subjects were examined. Sera from 256 younger healthy subjects from among the 454 healthy subjects were examined as a control. When the cutoff point of anti-TBGL antibody titer was determined as 2.0 U/ml, the sensitivity for active tuberculosis patients was 81.1% and the specificity was 95.7%. Sensitivity in patients with smear-negative and culture-negative active pulmonary tuberculosis was 73.5%. Even in patients with noncavitary minimally advanced lesions, the positivity rate (60.0%) and the antibody titer (4.6 ± 9.4 U/ml) were significantly higher than those in the healthy group. These results indicate that this assay using anti-TBGL antibody is useful for the rapid serodiagnosis of active pulmonary tuberculosis. PMID:11574580

Validity of rapid antigen detection testing in group A beta-hemolytic streptococcal tonsillopharyngitis.

PubMed

Küçük, Oznur; Biçer, Suat; Giray, Tuba; Cöl, Defne; Erdağ, Gülay Ciler; Gürol, Yeşim; Kaspar, Ciğdem E; Vitrinel, Ayça

2014-02-01

To evaluate the utility of rapid antigen detection testing (RADT) for the diagnosis of group A beta-hemolytic streptococcal tonsillopharyngitis in children, and to detect the sensitivity and specificity of rapid antigen detection of group A beta-hemolytic streptococci from throat specimen compared with throat culture. Rapid antigen detection and throat culture results for group A beta-hemolytic streptococci from outpatients attending university hospital between 1st January 2011 and 31st of December 2011 were evaluated retrospectively. The antigen test negative-throat culture positive patients were investigated for streptococcal carriage. For this purpose, the throat culture results taken from these patients were reviewed after treatment. Eight hundred and ninetytwo children were included in the studywith a mean age of 5.34 y. There were 639 and 253 children in two groups with age of 0-6 and 7-17 y, RADT sensitivity and specificity were found to be 59.5 % and 97.2 %, respectively. The positive predictive value was 87.1 %, whereas negative predictive value was 88.4 %. After treatment of 74 patients with throat culture positive and antigen test negative. Group A beta-hemolytic streptococci were isolated in 12 of them (16.2 %) and accepted as a carrier. The low sensitivity of the RADT may be related to streptococcal carriage in some patients. The throat culture should be repeated after treatment to detect streptococcal carriage.

Rapid detection of the Klebsiella pneumoniae carbapenemase (KPC) gene by loop-mediated isothermal amplification (LAMP).

PubMed

Nakano, Ryuichi; Nakano, Akiyo; Ishii, Yoshikazu; Ubagai, Tsuneyuki; Kikuchi-Ueda, Takane; Kikuchi, Hirotoshi; Tansho-Nagakawa, Shigeru; Kamoshida, Go; Mu, Xiaoqin; Ono, Yasuo

2015-03-01

Klebsiella pneumoniae carbapenemases (KPC), which are associated with resistance to carbapenem, have recently spread worldwide and have become a global concern. It is necessary to detect KPC-producing organisms in clinical settings to be able to control the spread of this resistance. We have developed a loop-mediated isothermal amplification (LAMP) method for rapid detection of KPC producers. LAMP primer sets were designed to recognize the homologous regions of blaKPC-2 to blaKPC-17 and could amplify blaKPC rapidly. The specificity and sensitivity of the primers in the LAMP reactions for blaKPC detection were determined. This LAMP assay was able to specifically detect KPC producers at 68 °C, and no cross-reactivity was observed for other types of β-lactamase (class A, B, C, or D) producers. The detection limit for this assay was found to be 10(0) CFU per tube, in 25 min, which was 10-fold more sensitive than a PCR assay for blaKPC detection. Then, the sensitivity of the LAMP reactions for blaKPC detection in human specimens (sputum samples, urine samples, fecal samples and blood samples) was analyzed; it was observed that the LAMP assay had almost the same sensitivity in these samples as when using purified DNA. The LAMP assay is easy to perform and rapid. It may therefore be routinely applied for detection of KPC producers in the clinical laboratory. Copyright © 2014 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Rapid and accurate detection of KRAS mutations in colorectal cancers using the isothermal-based optical sensor for companion diagnostics

PubMed Central

Koo, Bonhan; Lee, Tae Yoon; Lee, Jeong Hoon; Shin, Yong; Lim, Seok-Byung

2017-01-01

Although KRAS mutational status testing is becoming a companion diagnostic tool for managing patients with colorectal cancer (CRC), there are still several difficulties when analyzing KRAS mutations using the existing assays, particularly with regard to low sensitivity, its time-consuming, and the need for large instruments. We developed a rapid, sensitive, and specific mutation detection assay based on the bio-photonic sensor termed ISAD (isothermal solid-phase amplification/detection), and used it to analyze KRAS gene mutations in human clinical samples. To validate the ISAD-KRAS assay for use in clinical diagnostics, we examined for hotspot KRAS mutations (codon 12 and codon 13) in 70 CRC specimens using PCR and direct sequencing methods. In a serial dilution study, ISAD-KRAS could detect mutations in a sample containing only 1% of the mutant allele in a mixture of wild-type DNA, whereas both PCR and direct sequencing methods could detect mutations in a sample containing approximately 30% of mutant cells. The results of the ISAD-KRAS assay from 70 clinical samples matched those from PCR and direct sequencing, except in 5 cases, wherein ISAD-KRAS could detect mutations that were not detected by PCR and direct sequencing. We also found that the sensitivity and specificity of ISAD-KRAS were 100% within 30 min. The ISAD-KRAS assay provides a rapid, highly sensitive, and label-free method for KRAS mutation testing, and can serve as a robust and near patient testing approach for the rapid detection of patients most likely to respond to anti-EGFR drugs. PMID:29137388

Rapid-scan Fourier-transform coherent anti-Stokes Raman scattering spectroscopy with heterodyne detection.

PubMed

Hiramatsu, Kotaro; Luo, Yizhi; Ideguchi, Takuro; Goda, Keisuke

2017-11-01

High-speed Raman spectroscopy has become increasingly important for analyzing chemical dynamics in real time. To address the need, rapid-scan Fourier-transform coherent anti-Stokes Raman scattering (FT-CARS) spectroscopy has been developed to realize broadband CARS measurements at a scan rate of more than 20,000 scans/s. However, the detection sensitivity of FT-CARS spectroscopy is inherently low due to the limited number of photons detected during each scan. In this Letter, we show our experimental demonstration of enhanced sensitivity in rapid-scan FT-CARS spectroscopy by heterodyne detection. Specifically, we implemented heterodyne detection by superposing the CARS electric field with an external local oscillator (LO) for their interference. The CARS signal was amplified by simply increasing the power of the LO without the need for increasing the incident power onto the sample. Consequently, we achieved enhancement in signal intensity and the signal-to-noise ratio by factors of 39 and 5, respectively, compared to FT-CARS spectroscopy with homodyne detection. The sensitivity-improved rapid-scan FT-CARS spectroscopy is expected to enable the sensitive real-time observation of chemical dynamics in a broad range of settings, such as combustion engines and live biological cells.

Rapid, ultrasensitive detection of microorganisms based on interferometry and lab-on-a-chip nanotechnology

NASA Astrophysics Data System (ADS)

Ymeti, Aurel; Nederkoorn, Paul H. J.; Dudia, Alma; Subramaniam, Vinod; Kanger, Johannes S.

2009-05-01

Future viral outbreaks are a major threat to societal and economic development throughout the world. A rapid, sensitive, and easy-to-use test for viral infections is essential to prevent and to control such viral pandemics. Furthermore, a compact, portable device is potentially very useful in remote or developing regions without easy access to sophisticated laboratory facilities. We have developed a rapid, ultrasensitive sensor that could be used in a handheld device to detect various viruses and measure their concentration. The essential innovation in this technique is the combination of an integrated optical interferometric sensor with antibody-antigen recognition approaches to yield a very sensitive, very rapid test for virus detection. The sensor is able to spot the herpes virus at concentrations of just 850 particles per milliliter under physiological conditions. The sensitivity of the sensor approaches detection of a single virus particle, yielding a sensor of unprecedented sensitivity with wide applications for viral diagnostics. The sensor's detection principle can be extended to any biological target such as bacteria, cells and proteins and for which there are specific antibodies. The nature of the sensor enables multiplexed detection of several analytes at the same time.

Direct detection of falciparum and non-falciparum malaria DNA from a drop of blood with high sensitivity by the dried-LAMP system.

PubMed

Hayashida, Kyoko; Kajino, Kiichi; Simukoko, Humphrey; Simuunza, Martin; Ndebe, Joseph; Chota, Amos; Namangala, Boniface; Sugimoto, Chihiro

2017-01-13

Because of the low sensitivity of conventional rapid diagnostic tests (RDTs) for malaria infections, the actual prevalence of the diseases, especially those caused by non-Plasmodium falciparum (non-Pf) species, in asymptomatic populations remain less defined in countries lacking in well-equipped facilities for accurate diagnoses. Our direct blood dry LAMP system (CZC-LAMP) was applied to the diagnosis of malaria as simple, rapid and highly sensitive method as an alternative for conventional RDTs in malaria endemic areas where laboratory resources are limited. LAMP primer sets for mitochondria DNAs of Plasmodium falciparum (Pf) and human-infective species other than Pf (non-Pf; P. vivax, P. ovale, P. malariae) were designed and tested by using human blood DNA samples from 74 residents from a malaria endemic area in eastern Zambia. These malaria dry-LAMPs were optimized for field or point-of-care operations, and evaluated in the field at a malaria endemic area in Zambia with 96 human blood samples. To determine the sensitivities and specificities, results obtained by the on-site LAMP diagnosis were compared with those by the nested PCR and nucleotide sequencing of its product. The dry LAMPs showed the sensitivities of 89.7% for Pf and 85.7% for non-Pf, and the specificities of 97.2% for Pf and 100% for non-Pf, with purified blood DNA samples. The direct blood LAMP diagnostic methods, in which 1 μl of anticoagulated blood were used as the template, showed the sensitivities of 98.1% for Pf, 92.1% for non-Pf, and the specificities of 98.1% for Pf, 100% for non-Pf. The prevalences of P. falciparum, P. malariae and P. ovale in the surveyed area were 52.4, 25.3 and 10.6%, respectively, indicating high prevalence of asymptomatic carriers in endemic areas in Zambia. We have developed new field-applicable malaria diagnostic tests. The malaria CZC-LAMPs showed high sensitivity and specificity to both P. falciparum and non-P. falciparum. These malaria CZC-LAMPs provide new means for rapid, sensitive and reliable point-of-care diagnosis for low-density malaria infections, and are expected to help update current knowledge of malaria epidemiology, and can contribute to the elimination of malaria from endemic areas.

Rapid micromotor-based naked-eye immunoassay.

PubMed

de Ávila, Berta Esteban-Fernández; Zhao, Mingjiao; Campuzano, Susana; Ricci, Francesco; Pingarrón, José M; Mascini, Marcello; Wang, Joseph

2017-05-15

A dynamic micromotor-based immunoassay, exemplified by cortisol detection, based on the use of tubular micromotors functionalized with a specific antibody is described. The use of antibody-functionalized micromotors offers huge acceleration of both direct and competitive cortisol immunoassays, along with greatly enhanced sensitivity of direct and competitive immunoassays. The dramatically improved speed and sensitivity reflect the greatly increased likelihood of antibody-cortisol contacts and fluid mixing associated with the dynamic movement of these microtube motors and corresponding bubble generation that lead to a highly efficient and rapid recognition process. Rapid naked-eye detection of cortisol in the sample is achieved in connection to use of horseradish peroxidase (HRP) tag and TMB/H 2 O 2 system. Key parameters of the competitive immunoassay (e.g., incubation time and reaction volume) were optimized. This fast visual micromotor-based sensing approach enables "on the move" specific detection of the target cortisol down to 0.1μgmL -1 in just 2min, using ultrasmall (50µL) sample volumes. Copyright © 2017 Elsevier B.V. All rights reserved.

Mind the Gap: Two Dissociable Mechanisms of Temporal Processing in the Auditory System

PubMed Central

Anderson, Lucy A.

2016-01-01

High temporal acuity of auditory processing underlies perception of speech and other rapidly varying sounds. A common measure of auditory temporal acuity in humans is the threshold for detection of brief gaps in noise. Gap-detection deficits, observed in developmental disorders, are considered evidence for “sluggish” auditory processing. Here we show, in a mouse model of gap-detection deficits, that auditory brain sensitivity to brief gaps in noise can be impaired even without a general loss of central auditory temporal acuity. Extracellular recordings in three different subdivisions of the auditory thalamus in anesthetized mice revealed a stimulus-specific, subdivision-specific deficit in thalamic sensitivity to brief gaps in noise in experimental animals relative to controls. Neural responses to brief gaps in noise were reduced, but responses to other rapidly changing stimuli unaffected, in lemniscal and nonlemniscal (but not polysensory) subdivisions of the medial geniculate body. Through experiments and modeling, we demonstrate that the observed deficits in thalamic sensitivity to brief gaps in noise arise from reduced neural population activity following noise offsets, but not onsets. These results reveal dissociable sound-onset-sensitive and sound-offset-sensitive channels underlying auditory temporal processing, and suggest that gap-detection deficits can arise from specific impairment of the sound-offset-sensitive channel. SIGNIFICANCE STATEMENT The experimental and modeling results reported here suggest a new hypothesis regarding the mechanisms of temporal processing in the auditory system. Using a mouse model of auditory temporal processing deficits, we demonstrate the existence of specific abnormalities in auditory thalamic activity following sound offsets, but not sound onsets. These results reveal dissociable sound-onset-sensitive and sound-offset-sensitive mechanisms underlying auditory processing of temporally varying sounds. Furthermore, the findings suggest that auditory temporal processing deficits, such as impairments in gap-in-noise detection, could arise from reduced brain sensitivity to sound offsets alone. PMID:26865621

Use of bacteriophage cell wall-binding proteins for rapid diagnostics of Listeria.

PubMed

Schmelcher, Mathias; Loessner, Martin J

2014-01-01

Diagnostic protocols for food-borne bacterial pathogens such as Listeria need to be sensitive, specific, rapid, and inexpensive. Conventional culture methods are hampered by lengthy enrichment and incubation steps. Bacteriophage-derived high-affinity binding molecules (cell wall-binding domains, CBDs) specific for Listeria cells have recently been introduced as tools for detection and differentiation of this pathogen in foods. When coupled with magnetic separation, these proteins offer advantages in sensitivity and speed compared to the standard diagnostic methods. Furthermore, fusion of CBDs to differently colored fluorescent reporter proteins enables differentiation of Listeria strains in mixed cultures. This chapter provides protocols for detection of Listeria in food by CBD-based magnetic separation and subsequent multiplexed identification of strains of different serotypes with reporter-CBD fusion proteins.

Direct Application of the INNO-LiPA Rif.TB Line-Probe Assay for Rapid Identification of Mycobacterium tuberculosis Complex Strains and Detection of Rifampin Resistance in 360 Smear-Positive Respiratory Specimens from an Area of High Incidence of Multidrug-Resistant Tuberculosis

PubMed Central

Viveiros, Miguel; Leandro, Clara; Rodrigues, Liliana; Almeida, Josefina; Bettencourt, Rosário; Couto, Isabel; Carrilho, Lurdes; Diogo, José; Fonseca, Ana; Lito, Luís; Lopes, João; Pacheco, Teresa; Pessanha, Mariana; Quirim, Judite; Sancho, Luísa; Salfinger, Max; Amaral, Leonard

2005-01-01

The INNO-LiPA Rif.TB assay for the identification of Mycobacterium tuberculosis complex strains and the detection of rifampin (RIF) resistance has been evaluated with 360 smear-positive respiratory specimens from an area of high incidence of multidrug-resistant tuberculosis (MDR-TB). The sensitivity when compared to conventional identification/culture methods was 82.2%, and the specificity was 66.7%; the sensitivity and specificity were 100.0% and 96.9%, respectively, for the detection of RIF resistance. This assay has the potential to provide rapid information that is essential for the effective management of MDR-TB. PMID:16145166

Improved Detection of Botulinum Neurotoxin Serotype A by Endopep-MS through Peptide Substrate Modification

PubMed Central

Wang, Dongxia; Baudys, Jakub; Ye, Yiming; Rees, Jon C.; Barr, John R.; Pirkle, James L.; Kalb, Suzanne R.

2015-01-01

Botulinum neurotoxins (BoNTs) are a family of seven toxin serotypes that are the most toxic substances known to man. Intoxication with BoNT causes flaccid paralysis and can lead to death if untreated with serotype specific antibodies. Supportive care, including ventilation, may be necessary. Rapid and sensitive detection of BoNT is necessary for timely clinical confirmation of clinical botulism. Previously, our laboratory developed a fast and sensitive mass spectrometry (MS) method termed the Endopep-MS assay. The BoNT serotypes are rapidly detected and differentiated by extracting the toxin with serotype specific antibodies and detecting the unique and serotype specific cleavage products of peptide substrates that mimic the sequence of the BoNT native targets. To further improve the sensitivity of the Endopep-MS assay, we report here the optimization of the substrate peptide for the detection of BoNT/A. Modifications on the terminal groups of the original peptide substrate with acetylation and amidation significantly improved the detection of BoNT/A cleavage products. The replacement of some internal amino acid residues with single or multiple substitutions led to further improvement. An optimized peptide increased assay sensitivity five fold with toxin spiked into buffer solution or different biological matrices. PMID:23017875

Sensitive and rapid detection of Chlamydia trachomatis by recombinase polymerase amplification directly from urine samples.

PubMed

Krõlov, Katrin; Frolova, Jekaterina; Tudoran, Oana; Suhorutsenko, Julia; Lehto, Taavi; Sibul, Hiljar; Mäger, Imre; Laanpere, Made; Tulp, Indrek; Langel, Ülo

2014-01-01

Chlamydia trachomatis is the most common sexually transmitted human pathogen. Infection results in minimal to no symptoms in approximately two-thirds of women and therefore often goes undiagnosed. C. trachomatis infections are a major public health concern because of the potential severe long-term consequences, including an increased risk of ectopic pregnancy, chronic pelvic pain, and infertility. To date, several point-of-care tests have been developed for C. trachomatis diagnostics. Although many of them are fast and specific, they lack the required sensitivity for large-scale application. We describe a rapid and sensitive form of detection directly from urine samples. The assay uses recombinase polymerase amplification and has a minimum detection limit of 5 to 12 pathogens per test. Furthermore, it enables detection within 20 minutes directly from urine samples without DNA purification before the amplification reaction. Initial analysis of the assay from clinical patient samples had a specificity of 100% (95% CI, 92%-100%) and a sensitivity of 83% (95% CI, 51%-97%). The whole procedure is fairly simple and does not require specific machinery, making it potentially applicable in point-of-care settings. Copyright © 2014 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

Rapid and sensitive multiplex single-tube nested PCR for the identification of five human Plasmodium species.

PubMed

Saito, Takahiro; Kikuchi, Aoi; Kaneko, Akira; Isozumi, Rie; Teramoto, Isao; Kimura, Masatsugu; Hirasawa, Noriyasu; Hiratsuka, Masahiro

2018-06-01

Malaria is caused by five species of Plasmodium in humans. Microscopy is currently used for pathogen detection, requiring considerable training and technical expertise as the parasites are often difficult to differentiate morphologically. Rapid diagnostic tests are as reliable as microscopy and offer faster diagnoses but possess lower detection limits and are incapable of distinguishing among the parasitic species. To improve global health efforts towards malaria control, a rapid, sensitive, species-specific, and economically viable diagnostic method is needed. In this study, we designed a malaria diagnostic method involving a multiplex single-tube nested PCR targeting Plasmodium mitochondrial cytochrome c oxidase III and single-stranded tag hybridization chromatographic printed-array strip. The detection sensitivity was found to be at least 40 times higher than that of agarose gel electrophoresis with ethidium bromide. This system also enables the identification of both single- and mixed-species malaria infections. The assay was validated with 152 Kenyan samples; using nested PCR as the standard, the assay's sensitivity and specificity were 88.7% and 100.0%, respectively. The turnaround time required, from PCR preparation to signal detection, is 90min. Our method should improve the diagnostic speed, treatment efficacy, and control of malaria, in addition to facilitating surveillance within global malaria eradication programs. Copyright © 2018 Elsevier B.V. All rights reserved.

Nanoswitch-linked immunosorbent assay (NLISA) for fast, sensitive, and specific protein detection.

PubMed

Hansen, Clinton H; Yang, Darren; Koussa, Mounir A; Wong, Wesley P

2017-09-26

Protein detection and quantification play critical roles in both basic research and clinical practice. Current detection platforms range from the widely used ELISA to more sophisticated, and more expensive, approaches such as digital ELISA. Despite advances, there remains a need for a method that combines the simplicity and cost-effectiveness of ELISA with the sensitivity and speed of modern approaches in a format suitable for both laboratory and rapid, point-of-care applications. Building on recent developments in DNA structural nanotechnology, we introduce the nanoswitch-linked immunosorbent assay (NLISA), a detection platform based on easily constructed DNA nanodevices that change conformation upon binding to a target protein with the results read out by gel electrophoresis. NLISA is surface-free and includes a kinetic-proofreading step for purification, enabling both enhanced sensitivity and reduced cross-reactivity. We demonstrate femtomolar-level detection of prostate-specific antigen in biological fluids, as well as reduced cross-reactivity between different serotypes of dengue and also between a single-mutation and wild-type protein. NLISA is less expensive, uses less sample volume, is more rapid, and, with no washes, includes fewer hands-on steps than ELISA, while also achieving superior sensitivity. Our approach also has the potential to enable rapid point-of-care assays, as we demonstrate by performing NLISA with an iPad/iPhone camera for imaging.

A genetic variant of the sperm-specific SLO3 K+ channel has altered pH and Ca2+ sensitivities.

PubMed

Geng, Yanyan; Ferreira, Juan J; Dzikunu, Victor; Butler, Alice; Lybaert, Pascale; Yuan, Peng; Magleby, Karl L; Salkoff, Lawrence; Santi, Celia M

2017-05-26

To fertilize an oocyte, sperm must first undergo capacitation in which the sperm plasma membrane becomes hyperpolarized via activation of potassium (K + ) channels and resultant K + efflux. Sperm-specific SLO3 K + channels are responsible for these membrane potential changes critical for fertilization in mouse sperm, and they are only sensitive to pH i However, in human sperm, the major K + conductance is both Ca 2+ - and pH i -sensitive. It has been debated whether Ca 2+ -sensitive SLO1 channels substitute for human SLO3 (hSLO3) in human sperm or whether human SLO3 channels have acquired Ca 2+ sensitivity. Here we show that hSLO3 is rapidly evolving and reveal a natural structural variant with enhanced apparent Ca 2+ and pH sensitivities. This variant allele (C382R) alters an amino acid side chain at a principal interface between the intramembrane-gated pore and the cytoplasmic gating ring of the channel. Because the gating ring contains sensors to intracellular factors such as pH and Ca 2+ , the effectiveness of transduction between the gating ring and the pore domain appears to be enhanced. Our results suggest that sperm-specific genes can evolve rapidly and that natural genetic variation may have led to a SLO3 variant that differs from wild type in both pH and intracellular Ca 2+ sensitivities. Whether this physiological variation confers differences in fertility among males remains to be established. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

A novel restriction endonuclease GlaI for rapid and highly sensitive detection of DNA methylation coupled with isothermal exponential amplification reaction.

PubMed

Sun, Yueying; Sun, Yuanyuan; Tian, Weimin; Liu, Chenghui; Gao, Kejian; Li, Zhengping

2018-02-07

Sensitive and accurate detection of site-specific DNA methylation is of critical significance for early diagnosis of human diseases, especially cancers. Herein, for the first time we employ a novel methylation-dependent restriction endonuclease GlaI to detect site-specific DNA methylation in a highly specific and sensitive way by coupling with isothermal exponential amplification reaction (EXPAR). GlaI can only cut the methylated target site with excellent selectivity but leave the unmethylated DNA intact. Then the newly exposed end fragments of methylated DNA can trigger EXPAR for highly efficient signal amplification while the intact unmethylated DNA will not initiate EXPAR at all. As such, only the methylated DNA is quantitatively and faithfully reflected by the real-time fluorescence signal of the GlaI-EXPAR system, and the potential false positive interference from unmethylated DNA can be effectively eliminated. Therefore, by integrating the unique features of GlaI for highly specific methylation discrimination and EXPAR for rapid and powerful signal amplification, the elegant GlaI-EXPAR assay allows the direct quantification of methylated DNA with ultrahigh sensitivity and accuracy. The detection limit of methylated DNA target has been pushed down to the aM level and the whole detection process of GlaI-EXPAR can be accomplished within a short time of 2 h. More importantly, ultrahigh specificity is achieved and as low as 0.01% methylated DNA can be clearly identified in the presence of a large excess of unmethylated DNA. This GlaI-EXPAR is also demonstrated to be capable of determining site-specific DNA methylations in real genomic DNA samples. Sharing the distinct advantages of ultrahigh sensitivity, outstanding specificity and facile operation, this new GlaI-EXPAR strategy may provide a robust and reliable platform for the detection of site-specific DNA methylations with low abundances.

Serological Diagnosis of Acute Scrub Typhus in Southern India: Evaluation of InBios Scrub Typhus Detect IgM Rapid Test and Comparison with other Serological Tests.

PubMed

Anitharaj, Velmurugan; Stephen, Selvaraj; Pradeep, Jothimani; Park, Sungman; Kim, Seung-Han; Kim, Young Jin; Kim, Eun-Ye; Kim, Yoon-Won

2016-11-01

Scrub Typhus (ST) is being reported from different parts of India in the recent past. However, the diagnosis and confirmation of ST cases require specific serological and molecular diagnostic tests. Both rapid and conventional ELISA tests need to be properly evaluated. Evaluation of a new ST IgM Immunochromatography (ICT) test kit (InBios Scrub Typhus Detect IgM Rapid Test) and compare it with another rapid kit, conventional ELISA kit and Weil-Felix (WF) test. This prospective study was carried out in Mahatma Gandhi Medical College and Research Institute, Puducherry, during November 2015 to June 2016. Clinically suspected 220 ST patients were examined by a new kit, InBios Scrub Typhus Detect IgM Rapid Test, taking the conventional InBios Scrub Typhus Detect IgM ELISA as reference. Additional comparison was made with ImmuneMed Scrub Typhus Rapid, and WF test (single OXK titers ≥1:320). Statistical analysis was performed (Chi-square, Spearman's correlation and Kappa) using IBM SPSS Statistics 17 for Windows (SPSS Inc; Chicago, USA). Percentage Sensitivity, Specificity, Positive Predictive and Negative Predictive Values for InBios, ImmuneMed and WF were 99.25, 93.02, 95.68, 98.77; 94.87, 94.19, 96.21, 92.05 and 50.38, 95.51, 94.29, 56.67 respectively. A total of 134 patients were positive in reference standard InBios IgM ELISA. This new rapid ST IgM kit validated for the first time in India, showed good sensitivity and specificity. As a Point-of-Care (PoC) test, the kit would be helpful in both urban and remote rural parts of India.

Serological Diagnosis of Acute Scrub Typhus in Southern India: Evaluation of InBios Scrub Typhus Detect IgM Rapid Test and Comparison with other Serological Tests

PubMed Central

Anitharaj, Velmurugan; Pradeep, Jothimani; Park, Sungman; Kim, Seung-Han; Kim, Young Jin; Kim, Eun-Ye; Kim, Yoon-Won

2016-01-01

Introduction Scrub Typhus (ST) is being reported from different parts of India in the recent past. However, the diagnosis and confirmation of ST cases require specific serological and molecular diagnostic tests. Both rapid and conventional ELISA tests need to be properly evaluated. Aim Evaluation of a new ST IgM Immunochromatography (ICT) test kit (InBios Scrub Typhus Detect IgM Rapid Test) and compare it with another rapid kit, conventional ELISA kit and Weil-Felix (WF) test. Materials and Methods This prospective study was carried out in Mahatma Gandhi Medical College and Research Institute, Puducherry, during November 2015 to June 2016. Clinically suspected 220 ST patients were examined by a new kit, InBios Scrub Typhus Detect IgM Rapid Test, taking the conventional InBios Scrub Typhus Detect IgM ELISA as reference. Additional comparison was made with ImmuneMed Scrub Typhus Rapid, and WF test (single OXK titers ≥1:320). Statistical analysis was performed (Chi-square, Spearman’s correlation and Kappa) using IBM SPSS Statistics 17 for Windows (SPSS Inc; Chicago, USA). Results Percentage Sensitivity, Specificity, Positive Predictive and Negative Predictive Values for InBios, ImmuneMed and WF were 99.25, 93.02, 95.68, 98.77; 94.87, 94.19, 96.21, 92.05 and 50.38, 95.51, 94.29, 56.67 respectively. A total of 134 patients were positive in reference standard InBios IgM ELISA. Conclusion This new rapid ST IgM kit validated for the first time in India, showed good sensitivity and specificity. As a Point-of-Care (PoC) test, the kit would be helpful in both urban and remote rural parts of India. PMID:28050364

Detection of Entamoeba histolytica by Recombinase Polymerase Amplification

PubMed Central

Nair, Gayatri; Rebolledo, Mauricio; White, A. Clinton; Crannell, Zachary; Richards-Kortum, R. Rebecca; Pinilla, A. Elizabeth; Ramírez, Juan David; López, M. Consuelo; Castellanos-Gonzalez, Alejandro

2015-01-01

Amebiasis is an important cause of diarrheal disease worldwide and has been associated with childhood malnutrition. Traditional microscopy approaches are neither sensitive nor specific for Entamoeba histolytica. Antigen assays are more specific, but many cases are missed unless tested by molecular methods. Although polymerase chain reaction (PCR) is effective, the need for sophisticated, expensive equipment, infrastructure, and trained personnel limits its usefulness, especially in the resource-limited, endemic areas. Here, we report development of a recombinase polymerase amplification (RPA) method to detect E. histolytica specifically. Using visual detection by lateral flow (LF), the test was highly sensitive and specific and could be performed without additional equipment. The availability of this inexpensive, sensitive, and field-applicable diagnostic test could facilitate rapid diagnosis and treatment of amebiasis in endemic regions. PMID:26123960

Luciferase-Zinc-Finger System for the Rapid Detection of Pathogenic Bacteria.

PubMed

Shi, Chu; Xu, Qing; Ge, Yue; Jiang, Ling; Huang, He

2017-08-09

Rapid and reliable detection of pathogenic bacteria is crucial for food safety control. Here, we present a novel luciferase-zinc finger system for the detection of pathogens that offers rapid and specific profiling. The system, which uses a zinc-finger protein domain to probe zinc finger recognition sites, was designed to bind the amplified conserved regions of 16S rDNA, and the obtained products were detected using a modified luciferase. The luciferase-zinc finger system not only maintained luciferase activity but also allowed the specific detection of different bacterial species, with a sensitivity as low as 10 copies and a linear range from 10 to 10 4 copies per microliter of the specific PCR product. Moreover, the system is robust and rapid, enabling the simultaneous detection of 6 species of bacteria in artificially contaminated samples with excellent accuracy. Thus, we envision that our luciferase-zinc finger system will have far-reaching applications.

Rapid and Sensitive Enumeration of Viable Diluted Cells of Members of the Family Enterobacteriaceae in Freshwater and Drinking Water

PubMed Central

Baudart, Julia; Coallier, Josée; Laurent, Patrick; Prévost, Michèle

2002-01-01

Water quality assessment involves the specific, sensitive, and rapid detection of bacterial indicators and pathogens in water samples, including viable but nonculturable (VBNC) cells. This work evaluates the specificity and sensitivity of a new method which combines a fluorescent in situ hybridization (FISH) approach with a physiological assay (direct viable count [DVC]) for the direct enumeration, at the single-cell level, of highly diluted viable cells of members of the family Enterobacteriaceae in freshwater and drinking water after membrane filtration. The approach (DVC-FISH) uses a new direct detection device, the laser scanning cytometer (Scan RDI). Combining the DVC-FISH method on a membrane with Scan RDI detection makes it possible to detect as few as one targeted cell in approximately 108 nontargeted cells spread over the membrane. The ability of this new approach to detect and enumerate VBNC enterobacterial cells in freshwater and drinking water distribution systems was investigated and is discussed. PMID:12324357

Comparative performance of aldolase and lactate dehydrogenase rapid diagnostic tests in Plasmodium vivax detection

PubMed Central

2014-01-01

Background Misdiagnosis of malaria by commercial rapid diagnostic tests (RDTs) is a major cause of concern in the diagnosis of malaria. This retrospective study was aimed at assessing the relative performance of four RDTs with emphasis on the detection of two Plasmodium vivax antigens: aldolase and lactate dehydrogenase (LDH). Methods Three commercially available Plasmodium LDH or aldolase antigen detection kits (One Step Malaria P.f/P.v, ParaHit Total ver. 1.0, SD Bioline Malaria) and an anti-P. vivax aldolase-specific monoclonal antibody (mAb) pair 1C3-12 F10 were evaluated with P. vivax positive as well as non-P. vivax samples and healthy samples using blood smear examination as standard. Each test was read according to the manufacturer’s instructions. Results MAb 1C3-12 F10 pair targeting P. vivax-specific aldolase exhibited very good specificity and sensitivity of 100 and 97.4%, respectively. Positive predictive value (PPV) and negative predictive value (NPV) of 100 and 99.5%, respectively, were also observed. The anti-P. vivax LDH in the One-Step Malaria P.f/P.v test showed sensitivity, specificity, PPV and NPV of 93.5, 98.0, 88.9 and 98.8%, respectively. ParaHit Total ver. 1.0 targeting the pan-aldolase antigen showed sensitivity, specificity of 97.4 and 99.6%, respectively. PPV and NPV were both 99.5%. SD Bioline had sensitivity, specificity, PPV and NPV of 93.5, 100, 100 and 98.8%, respectively. The overall sensitivity and specificity of all four RDTs were acceptable, especially for the aldolase detection tests. Five (6.5%) of the P. vivax-positive samples (n = 77) that were confirmed by microscopic examination as well as the two aldolase detection RDTs (mAb 1C3-12 F10 and ParaHit Total ver.1.0) were undetected by the two LDH detection RDTs (One Step Malaria P.f/P.v and SD Bioline). Similarly, two positive samples (2.6%) that were positively confirmed by the LDH detection RDTs were also undetected by the aldolase detection test kits. Conclusion Aldolase and LDH antigens perform differently in different P. vivax samples; hence there is a high risk of misdiagnosis when monoclonal antibodies are used against only one particular antigen in the test. A combination of both aldolase and LDH in RDTs for the rapid diagnosis of P. vivax will enhance the sensitivity of the assay and reduce misdiagnosis. PMID:25015737

An improved, rapid competitive ELISA using a novel conserved 3B epitope for the detection of serum antibodies to foot-and-mouth disease virus.

PubMed

Chung, Chungwon J; Clavijo, Alfonso; Bounpheng, Mangkey A; Uddowla, Sabena; Sayed, Abu; Dancho, Brooke; Olesen, Ian C; Pacheco, Juan; Kamicker, Barbara J; Brake, David A; Bandaranayaka-Mudiyanselage, Carey L; Lee, Stephen S; Rai, Devendra K; Rieder, Elizabeth

2018-06-01

The highly contagious foot-and-mouth disease virus (FMDV) afflicts cloven-hoofed animals, resulting in significant costs because of loss of trade and recovery from disease. We developed a sensitive, specific, and rapid competitive ELISA (cELISA) to detect serum antibodies to FMDV. The cELISA utilized a monoclonal blocking antibody specific for a highly conserved FMDV nonstructural 3B epitope, a recombinant mutant FMDV 3ABC coating protein, and optimized format variables including serum incubation for 90 min at 20-25°C. Samples from 16 animals experimentally infected with one FMDV serotype (A, O, Asia, or SAT-1) demonstrated early detection capacity beginning 7 d post-inoculation. All samples from 55 vesicular stomatitis virus antibody-positive cattle and 44 samples from cloven-hoofed animals affected by non-FMD vesicular diseases were negative in the cELISA, demonstrating 100% analytical specificity. The diagnostic sensitivity was 100% against sera from 128 cattle infected with isolates of all FMDV serotypes, emphasizing serotype-agnostic results. Diagnostic specificities of U.S. cattle ( n = 1135) and swine ( n = 207) sera were 99.4% and 100%, respectively. High repeatability and reproducibility were demonstrated with 3.1% coefficient of variation in percent inhibition data and 100% agreement using 2 kit lots and 400 negative control serum samples, with no difference between bench and biosafety cabinet operation. Negative results from vaccinated, uninfected cattle, pig, and sheep sera confirmed the DIVA (differentiate infected from vaccinated animals) capability. This rapid (<3 h), select agent-free assay with high sensitivity and specificity, DIVA capability, and room temperature processing capability will serve as a useful tool in FMDV surveillance, emergency preparedness, response, and outbreak recovery programs.

Preparation of immunochromatographic strips for rapid detection of early secreted protein ESAT-6 and culture filtrate protein CFP-10 from Mycobacterium tuberculosis

PubMed Central

Wu, Xiaoxin; Wang, Yeping; Weng, Tianhao; Hu, Chenyu; Wang, Frederick X.C.; Wu, Zhigang; Yu, Dongshan; Lu, Huoquan; Yao, Hangping

2017-01-01

Abstract The early secreted protein early secretory antigenic target 6(ESAT-6) and the culture filtrate protein 10 (CFP-10) are 2 antigens that are specific to Mycobacterium tuberculosis. These 2 antigens are good targets for tuberculosis (TB) detection. To rapidly diagnose TB across a variety of samples, we developed colloidal gold immunochromatographic strips (ICSs) based on ESAT-6 and CFP-10. The strips were evaluated using 233 samples, including sputum, plasma, and pleural effusion samples. The positive detection rates for ICSs for ESAT-6 and CFP-10 in sputum (culture-positive for M tuberculosis) were 100% and 91.2%, respectively. The positive detection rates for ICSs for ESAT-6 and CFP-10 in plasma were 34.1% and 29.4%, respectively. The positive detection rates for ICSs for ESAT-6 and CFP-10 in pleural effusion were 64.7% and 55.9%, respectively. Experimental analysis of culture supernatant showing that the ICS developed for ESAT-6 had a sensitivity of 100% and a specificity of 91.2%. While the ICS developed for CFP-10 had a sensitivity of 91.2% and a specificity of 88.2%. The validity of the test is limited by source of sample. The technique is sensitive and specific for samples in sputum and culture media but not for plasma or pleural effusion samples. Detection of M tuberculosis using ICSs is rapid, simple, and relatively effective; thus, ICSs are a potential screening tool for TB. PMID:29390519

Preparation of immunochromatographic strips for rapid detection of early secreted protein ESAT-6 and culture filtrate protein CFP-10 from Mycobacterium tuberculosis.

PubMed

Wu, Xiaoxin; Wang, Yeping; Weng, Tianhao; Hu, Chenyu; Wang, Frederick X C; Wu, Zhigang; Yu, Dongshan; Lu, Huoquan; Yao, Hangping

2017-12-01

The early secreted protein early secretory antigenic target 6(ESAT-6) and the culture filtrate protein 10 (CFP-10) are 2 antigens that are specific to Mycobacterium tuberculosis. These 2 antigens are good targets for tuberculosis (TB) detection.To rapidly diagnose TB across a variety of samples, we developed colloidal gold immunochromatographic strips (ICSs) based on ESAT-6 and CFP-10.The strips were evaluated using 233 samples, including sputum, plasma, and pleural effusion samples.The positive detection rates for ICSs for ESAT-6 and CFP-10 in sputum (culture-positive for M tuberculosis) were 100% and 91.2%, respectively. The positive detection rates for ICSs for ESAT-6 and CFP-10 in plasma were 34.1% and 29.4%, respectively. The positive detection rates for ICSs for ESAT-6 and CFP-10 in pleural effusion were 64.7% and 55.9%, respectively. Experimental analysis of culture supernatant showing that the ICS developed for ESAT-6 had a sensitivity of 100% and a specificity of 91.2%. While the ICS developed for CFP-10 had a sensitivity of 91.2% and a specificity of 88.2%.The validity of the test is limited by source of sample. The technique is sensitive and specific for samples in sputum and culture media but not for plasma or pleural effusion samples. Detection of M tuberculosis using ICSs is rapid, simple, and relatively effective; thus, ICSs are a potential screening tool for TB. Copyright © 2017 The Authors. Published by Wolters Kluwer Health, Inc. All rights reserved.

SPECIES-SPECIFIC DETECTION OF THREE HUMAN-PATHOGENIC MICROSPORIDIAL SPECIES FROM THE GENUS ENCEPHALITOZOON VIA FLUOROGENIC 5' NUCLEASE PCR ASSAYS

EPA Science Inventory

This describes fluorogenic 5' nuclease PCR assays suitable for rapid, sensitive, quantitative, high-throughput detection of the human-pathogenic microsporidial species Encephalitozoon hellem, E. cunicli and E. intestinalis. The assays utilize species-specific primer sets and a g...

Rapid detection of urinary soluble intercellular adhesion molecule-1 for determination of lupus nephritis activity.

PubMed

Wang, Yanyun; Tao, Ye; Liu, Yi; Zhao, Yi; Song, Chao; Zhou, Bin; Wang, Tao; Gao, Linbo; Zhang, Lin; Hu, Huaizhong

2018-06-01

The current methods of monitoring the activity of lupus nephritis (LN) may cause unnecessary hospital visits or delayed immunosuppressive therapy. We aimed to find a urinary biomarker that could be developed as a home-based test for monitoring the activity of LN.Urine samples were collected immediately before a renal biopsy from patients of suspected active LN, and also from patients with inactive LN, systemic lupus erythematous without LN or healthy controls. Biomarker search was conducted on a cytokine antibody array and confirmation was done by quantitative evaluation with enzyme-linked immunosorbent assay. The Mann-Whiney test or Student t test was used to compare the levels of 9 cytokines between different groups. The sensitivity and specificity of each cytokine for diagnosis of LN was evaluated by receiver operating characteristic curve. A rapid test based on colloidal gold immunochromatography was then developed for bedside or home use. Furthermore, an experimental e-healthcare system was constructed for recording and sharing the results of the rapid test a cloud-assisted internet of things (IoT) consisting of a sensing device, an IoT device and a cloud server.Adiponectin (Acrp30), soluble intercellular cell adhesion molecule-1 (sICAM-1), neural cell adhesion molecule 1 (NCAM-1), and CD26 were significantly higher in urine samples of active LN patients. sICAM-1 appeared more sensitive and specific among these candidates. When the cut-off value of sICAM-1 was set at 1.44 ng/mL, the sensitivity reached 98.33% with a specificity at 85.71%. The sICAM-1 strip test showed comparable sensitivity of 95% and a specificity of 83.3% for assessing the LN activity. Meanwhile, the e-healthcare system was able to conveniently digitize and share the sICAM-1 rapid test results.sICAM-1 appeared to be an excellent biomarker for monitoring LN activity. The e-healthcare system with cloud-assisted IoT could assist the digitalization and sharing of the bedside or home-based sICAM-1 test results.

Variola virus-specific diagnostic assays: characterization, sensitivity, and specificity.

PubMed

Kondas, Ashley V; Olson, Victoria A; Li, Yu; Abel, Jason; Laker, Miriam; Rose, Laura; Wilkins, Kimberly; Turner, Jonathan; Kline, Richard; Damon, Inger K

2015-04-01

A public health response relies upon rapid and reliable confirmation of disease by diagnostic assays. Here, we detail the design and validation of two variola virus-specific real-time PCR assays, since previous assays cross-reacted with newly identified cowpox viruses. The assay specificity must continually be reassessed as other closely related viruses are identified. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Multiplex qRT-PCR for the Detection of Western Equine Encephalomyelitis, St. Louis Encephalitis, and West Nile Viral RNA in Mosquito Pools (Diptera: Culicidae).

PubMed

Brault, Aaron C; Fang, Ying; Reisen, William K

2015-05-01

Following the introduction of West Nile virus into California during the summer of 2003, public health and vector control programs expanded surveillance efforts and were in need of diagnostics capable of rapid, sensitive, and specific detection of arbovirus infections of mosquitoes to inform decision support for intervention. Development of a multiplex TaqMan or real-time semiquantitative reverse transcription polymerase chain reaction (RT-PCR) assay in which three virus specific primer-probe sets were used in the same reaction is described herein for the detection of western equine encephalomyelitis, St. Louis encephalitis and West Nile viral RNA. Laboratory validation and field data from 10 transmission seasons are reported. The comparative sensitivity and specificity of this multiplex assay to singleplex RT-PCR as well as an antigen detection (rapid analyte measurement platform) and standard plaque assays indicate this assay to be rapid and useful in providing mosquito infection data to estimate outbreak risk. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Bioconjugated fluorescent silica nanoparticles for the rapid detection of Entamoeba histolytica.

PubMed

Hemadi, Ahmad; Ekrami, Alireza; Oormazdi, Hormozd; Meamar, Ahmad Reza; Akhlaghi, Lame; Samarbaf-Zadeh, Ali Reza; Razmjou, Elham

2015-05-01

Rapid detection of Entamoeba histolytica based on fluorescent silica nanoparticle (FSNP) indirect immunofluorescence microscopy was evaluated. Silica nanoparticles were synthesized using Stöber's method, with their surface activated to covalently bind to, and immobilize, protein A. For biolabeling, FSNP was added to conjugated E. histolytica trophozoites with monoclonal anti-E. histolytica IgG1 for microscopic observation of fluorescence. Fluorescent silica nanoparticle sensitivity was determined with axenically cultured E. histolytica serially diluted to seven concentrations. Specificity was evaluated using other intestinal protozoa. Fluorescent silica nanoparticles detected E. histolytica at the lowest tested concentration with no cross-reaction with Entamoeba dispar, Entamoeba moshkovskii, Blastocystis sp., or Giardia lamblia. Visualization of E. histolytica trophozoites with anti-E. histolytica antibody labeled with fluorescein isothiocyanate (FITC) was compared with that using anti-E. histolytica antibody bioconjugated FSNP. Although FITC and FSNP produced similar results, the amount of specific antibody required for FITC to induce fluorescence of similar intensity was fivefold that for FSNP. Fluorescent silica nanoparticles delivered a rapid, simple, cost-effective, and highly sensitive and specific method of detecting E. histolytica. Further study is needed before introducing FSNP for laboratory diagnosis of amoebiasis. Copyright © 2015 Elsevier B.V. All rights reserved.

Reliable detection of Bacillus anthracis, Francisella tularensis and Yersinia pestis by using multiplex qPCR including internal controls for nucleic acid extraction and amplification.

PubMed

Janse, Ingmar; Hamidjaja, Raditijo A; Bok, Jasper M; van Rotterdam, Bart J

2010-12-08

Several pathogens could seriously affect public health if not recognized timely. To reduce the impact of such highly pathogenic micro-organisms, rapid and accurate diagnostic tools are needed for their detection in various samples, including environmental samples. Multiplex real-time PCRs were designed for rapid and reliable detection of three major pathogens that have the potential to cause high morbidity and mortality in humans: B. anthracis, F. tularensis and Y. pestis. The developed assays detect three pathogen-specific targets, including at least one chromosomal target, and one target from B. thuringiensis which is used as an internal control for nucleic acid extraction from refractory spores as well as successful DNA amplification. Validation of the PCRs showed a high analytical sensitivity, specificity and coverage of diverse pathogen strains. The multiplex qPCR assays that were developed allow the rapid detection of 3 pathogen-specific targets simultaneously, without compromising sensitivity. The application of B. thuringiensis spores as internal controls further reduces false negative results. This ensures highly reliable detection, while template consumption and laboratory effort are kept at a minimum.

Reliable detection of Bacillus anthracis, Francisella tularensis and Yersinia pestis by using multiplex qPCR including internal controls for nucleic acid extraction and amplification

PubMed Central

2010-01-01

Background Several pathogens could seriously affect public health if not recognized timely. To reduce the impact of such highly pathogenic micro-organisms, rapid and accurate diagnostic tools are needed for their detection in various samples, including environmental samples. Results Multiplex real-time PCRs were designed for rapid and reliable detection of three major pathogens that have the potential to cause high morbidity and mortality in humans: B. anthracis, F. tularensis and Y. pestis. The developed assays detect three pathogen-specific targets, including at least one chromosomal target, and one target from B. thuringiensis which is used as an internal control for nucleic acid extraction from refractory spores as well as successful DNA amplification. Validation of the PCRs showed a high analytical sensitivity, specificity and coverage of diverse pathogen strains. Conclusions The multiplex qPCR assays that were developed allow the rapid detection of 3 pathogen-specific targets simultaneously, without compromising sensitivity. The application of B. thuringiensis spores as internal controls further reduces false negative results. This ensures highly reliable detection, while template consumption and laboratory effort are kept at a minimum PMID:21143837

Detection of Citrus tristeza virus by using fluorescence resonance energy transfer-based biosensor

NASA Astrophysics Data System (ADS)

Shojaei, Taha Roodbar; Salleh, Mohamad Amran Mohd; Sijam, Kamaruzaman; Rahim, Raha Abdul; Mohsenifar, Afshin; Safarnejad, Reza; Tabatabaei, Meisam

2016-12-01

Due to the low titer or uneven distribution of Citrus tristeza virus (CTV) in field samples, detection of CTV by using conventional detection techniques may be difficult. Therefore, in the present work, the cadmium-telluride quantum dots (QDs) was conjugated with a specific antibody against coat protein (CP) of CTV, and the CP were immobilized on the surface of gold nanoparticles (AuNPs) to develop a specific and sensitive fluorescence resonance energy transfer (FRET)-based nanobiosensor for detecting CTV. The maximum FRET efficiency for the developed nano-biosensor was observed at 60% in AuNPs-CP/QDs-Ab ratio of 1:8.5. The designed system showed higher sensitivity and specificity over enzyme linked immunosorbent assay (ELISA) with a limit of detection of 0.13 μg mL- 1 and 93% and 94% sensitivity and specificity, respectively. As designed sensor is rapid, sensitive, specific and efficient in detecting CTV, this could be envisioned for diagnostic applications, surveillance and plant certification program.

Evaluation of CHROMagar KPC for Rapid Detection of Carbapenem-Resistant Enterobacteriaceae▿

PubMed Central

Samra, Zmira; Bahar, Judi; Madar-Shapiro, Liora; Aziz, Nazi; Israel, Sara; Bishara, Jihad

2008-01-01

A new CHROMagar KPC medium was compared to MacConkey agar with carbapenem discs and PCR for the blaKPC gene for rapid detection of carbapenem-resistant Klebsiella pneumoniae. The sensitivity and specificity relative to PCR were 100% and 98.4%, respectively, for CHROMagar KPC and 92.7% and 95.9%, respectively, for MacConkey agar. PMID:18632915

A novel strategy for rapid detection of NT-proBNP

NASA Astrophysics Data System (ADS)

Cui, Qiyao; Sun, Honghao; Zhu, Hui

2017-09-01

In order to establish a simple, rapid, sensitive, and specific quantitative assay to detect the biomarkers of heart failure, in this study, biotin-streptavidin technology was employed with fluorescence immunochromatographic assay to detect the concentration of the biomarkers in serum, and this method was applied to detect NT-proBNP, which is valuable for diagnostic evaluation of heart failure.

An improved, rapid cELISA using a novel conserved 3B epitope for the serological diagnosis of foot-and-mouth disease

USDA-ARS?s Scientific Manuscript database

The highly contagious foot-and-mouth disease virus (FMDV) afflicts cloven-hoofed livestock and wildlife resulting in heavy economic losses due to loss of trade and recovery from disease. We developed a rapid, sensitive, and specific competitive ELISA to detect serum antibodies to FMDV. This new cELI...

Rapid, Efficient Detection and Drug Susceptibility Testing of Mycobacterium tuberculosis in Sputum by Microscopic Observation of Broth Cultures

PubMed Central

Caviedes, Luz; Lee, Tien-Shun; Gilman, Robert H.; Sheen, Patricia; Spellman, Emily; Lee, Ellen H.; Berg, Douglas E.; Montenegro-James, Sonia

2000-01-01

Inexpensive, rapid, and reliable methods of detecting infection by and drug susceptibility of Mycobacterium tuberculosis (MTB) are crucial to the control of tuberculosis. The novel microscopic observation broth-drug susceptibility assay (MODS) detects early growth of MTB in liquid medium, allowing more timely diagnosis and drug susceptibility testing. Sputum samples from hospitalized patients in Peru were analyzed by using stains, culture, and PCR. Sensitivity of MODS (92%) compared favorably with the most sensitive of the other culture methods (93%). Sputum samples positive for tuberculosis were tested for susceptibility to isoniazid and rifampin with the microwell alamar blue assay (MABA) and MODS. In 89% of cases, there was concordance between MODS and MABA. Of the diagnostic and susceptibility testing methods used, MODS yielded results most rapidly (median, 9.0 and 9.5 days, respectively). MODS is a rapid, inexpensive, sensitive, and specific method for MTB detection and susceptibility testing; it is particularly appropriate for use in developing countries burdened by significant infection rates and increasing numbers of multiple-drug-resistant cases. PMID:10699023

Rapid, efficient detection and drug susceptibility testing of Mycobacterium tuberculosis in sputum by microscopic observation of broth cultures. The Tuberculosis Working Group in Peru.

PubMed

Caviedes, L; Lee, T S; Gilman, R H; Sheen, P; Spellman, E; Lee, E H; Berg, D E; Montenegro-James, S

2000-03-01

Inexpensive, rapid, and reliable methods of detecting infection by and drug susceptibility of Mycobacterium tuberculosis (MTB) are crucial to the control of tuberculosis. The novel microscopic observation broth-drug susceptibility assay (MODS) detects early growth of MTB in liquid medium, allowing more timely diagnosis and drug susceptibility testing. Sputum samples from hospitalized patients in Peru were analyzed by using stains, culture, and PCR. Sensitivity of MODS (92%) compared favorably with the most sensitive of the other culture methods (93%). Sputum samples positive for tuberculosis were tested for susceptibility to isoniazid and rifampin with the microwell alamar blue assay (MABA) and MODS. In 89% of cases, there was concordance between MODS and MABA. Of the diagnostic and susceptibility testing methods used, MODS yielded results most rapidly (median, 9.0 and 9.5 days, respectively). MODS is a rapid, inexpensive, sensitive, and specific method for MTB detection and susceptibility testing; it is particularly appropriate for use in developing countries burdened by significant infection rates and increasing numbers of multiple-drug-resistant cases.

Specific and sensitive detection of the conifer pathogen Gremmeniella abietina by nested PCR

PubMed Central

Zeng, Qing-Yin; Hansson, Per; Wang, Xiao-Ru

2005-01-01

Background Gremmeniella abietina (Lagerb.) Morelet is an ascomycete fungus that causes stem canker and shoot dieback in many conifer species. The fungus is widespread and causes severe damage to forest plantations in Europe, North America and Asia. To facilitate early diagnosis and improve measures to control the spread of the disease, rapid, specific and sensitive detection methods for G. abietina in conifer hosts are needed. Results We designed two pairs of specific primers for G. abietina based on the 18S rDNA sequence variation pattern. These primers were validated against a wide range of fungi and 14 potential conifer hosts. Based on these specific primers, two nested PCR systems were developed. The first system employed universal fungal primers to enrich the fungal DNA targets in the first round, followed by a second round selective amplification of the pathogen. The other system employed G. abietina-specific primers in both PCR steps. Both approaches can detect the presence of G. abietina in composite samples with high sensitivity, as little as 7.5 fg G. abietina DNA in the host genomic background. Conclusion The methods described here are rapid and can be applied directly to a wide range of conifer species, without the need for fungal isolation and cultivation. Therefore, it represents a promising alternative to disease inspection in forest nurseries, plantations and quarantine control facilities. PMID:16280082

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

PubMed Central

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

Sensitivity ranking for freshwater invertebrates towards hydrocarbon contaminants.

PubMed

Gerner, Nadine V; Cailleaud, Kevin; Bassères, Anne; Liess, Matthias; Beketov, Mikhail A

2017-11-01

Hydrocarbons have an utmost economical importance but may also cause substantial ecological impacts due to accidents or inadequate transportation and use. Currently, freshwater biomonitoring methods lack an indicator that can unequivocally reflect the impacts caused by hydrocarbons while being independent from effects of other stressors. The aim of the present study was to develop a sensitivity ranking for freshwater invertebrates towards hydrocarbon contaminants, which can be used in hydrocarbon-specific bioindicators. We employed the Relative Sensitivity method and developed the sensitivity ranking S hydrocarbons based on literature ecotoxicological data supplemented with rapid and mesocosm test results. A first validation of the sensitivity ranking based on an earlier field study has been conducted and revealed the S hydrocarbons ranking to be promising for application in sensitivity based indicators. Thus, the first results indicate that the ranking can serve as the core component of future hydrocarbon-specific and sensitivity trait based bioindicators.

Rapid detection of multidrug resistant Gram-negative bacilli by Cica-Beta-Test strips.

PubMed

Lavigne, J-P; Pfeiffer, C; Vidal, L; Sotto, A

2011-02-01

This study aimed to evaluate the sensitivity and specificity of a new system (Cica-Beta-Test, Kanto Chemical, Japan) for rapid detection of AmpC-derepressed, extended-spectrum β-lactamases (ESβL) and metallo-β-lactamases (MβL). Two hundred Multi-Drug Resistant (MDR)-Gram-negative bacilli were studied: 170 Enterobacteriaceae and 30 Gram-negative non-fermentative bacteria. One hundred and eighteen strains produced an ESβL, seven MβL and 75 derepressed cephalosporinases. One drop of substrate was dispensed onto the filter pad of the Cica-Beta-Test strip. The bacterial colonies were spread on the filter pad of strip. The reading was performed after 2 to 15 min: turning chromogenic indicated the positive test. Three tests were used: Cica-Beta I for detection of MDR bacteria; Cica-Beta MβL for detection of MβL-producing bacteria and Cica-Beta CVA, which distinguish ESβL and AmpC-derepressed producers. Results were compared with molecular assays. Cica-Beta-Test I has detected 194 MDR (sensitivity 97%), Cica-Beta-Test MβL has shown the presence of six MβL tested (sensitivity 85.7%). Five strains were non-MβL false positive (specificity 97.3%). Cica-Beta-Test CVA allowed the differentiation of ESβL-producing strains (109/115) and AmpC-derepressed strains (56/67) (sensitivity 94.8%, specificity 83.8%). Because of their epidemic nature, the MDR strains are screened and require strict hygienic measures patients. The simultaneous use of three strips can quickly determine the presence of MDR including ESβL and MβL. Rapid screening of MDR avoids transmission and limits the use of broad-spectrum antibiotics. Copyright © 2010 Elsevier Masson SAS. All rights reserved.

Evaluation of a rapid diagnostic test for the detection of Burkholderia pseudomallei in the Lao People's Democratic Republic.

PubMed

Woods, Kate L; Boutthasavong, Latsaniphone; NicFhogartaigh, Caoimhe; Lee, Sue J; Davong, Viengmon; AuCoin, David P; Dance, David A B

2018-05-02

Burkholderia pseudomallei causes significant global morbidity and mortality, with the highest disease burden in parts of Asia where culture-based diagnosis is often not available. We prospectively evaluated the Active Melioidosis Detect (AMD, InBios International, USA) lateral flow immunoassay (LFI) for rapid detection of B. pseudomallei in turbid blood cultures, pus, sputum, sterile fluid, urine, and sera. Performance was compared to B. pseudomallei detection using monoclonal antibody latex agglutination (LA) and immunofluorescence assays (IFA), with culture as the gold standard. AMD was 99% (99/100; 94.6 - 100%) sensitive and 100% (308/308; 98.8-100%) specific on turbid blood culture bottles, with no difference to LA or IFA. AMD specificity was 100% on pus (122/122; 97.0-100%), sputum (20/20; 83.2-100%), and sterile fluid (44/44; 92 - 100%). Sensitivity on these samples was: pus 47.1% (8/17; 23.0 - 72.2%), sputum 33.3% (1/3; 0.84 - 90.6%), and sterile fluid 0% (0/2; 0 - 84.2%). Urine AMD had a positive predictive value of 94% (32/34; 79.7 - 98.5%) for diagnosing melioidosis in our cohort. AMD sensitivity on stored sera, collected prospectively from melioidosis cases during this study, was 13.9% (5/36; 4.7% - 29.5%) when compared to blood culture samples taken on the same day. In conclusion, the AMD is an excellent tool for rapid diagnosis of melioidosis from turbid blood cultures, and maintains specificity across all sample types. It is a promising tool for urinary antigen detection, which could revolutionise diagnosis of melioidosis in resource-limited settings. Further work is required to improve sensitivity on non-blood culture samples. Copyright © 2018 Woods et al.

Performance of two strategies for urgent ANCA and anti-GBM analysis in vasculitis.

PubMed

de Joode, Anoek A E; Roozendaal, Caroline; van der Leij, Marcel J; Bungener, Laura B; Sanders, Jan Stephan F; Stegeman, Coen A

2014-02-01

In anti-neutrophil cytoplasmic antibodies (ANCA) associated small vessel vasculitis (AAV), rapid testing for ANCA and anti-glomerular basement membrane (GBM) antibodies may be beneficial for therapeutic purpose. We analysed the diagnostic performance of two rapid ANCA and anti-GBM test methods in 260 patients with suspected AAV. Between January 2004 and November 2010, we analysed 260 samples by qualitative Dotblot (Biomedical Diagnostics); retrospective analysis followed with directly coated highly sensitive automated Phadia ELiA and ELiA anti-GBM. Results were related to the final clinical diagnosis and compared with routine capture ELISA. Seventy-four patients had a final diagnosis of AAV (n=62) or anti-GBM disease (n=12). Both Dotblot and ELiA detected all 12 cases of anti-GBM disease; 2 false positive results were found. Dotblot detected ANCA in 56 of 62 AAV patients (sensitivity 90%, NPV 97%), and showed 5 false positives (specificity 97%, PPV 90%). The Phadia ELiA anti-PR3(s) or anti-MPO(s) was positive in 57 of 62 AAV patients (sensitivity 92%, NPV 97%), and had 5 false positives (specificity 97%, PPV 88%). Routine capture ELISA was equally accurate (sensitivity 94%, specificity 97%, PPV 88%, NPV 98%). The Dotblot and Phadia ELiA on anti-GBM, anti-PR3(s) and anti-MPO(s) performed excellently; results were almost identical to routine ELISA. When suspicion of AAV or anti-GBM disease is high and diagnosis is urgently needed, both tests are very powerful for rapid serological diagnosis. Further studies have to confirm the test performances in samples routinely presented for ANCA testing and in follow-up of positive patients. Copyright © 2013 European Federation of Internal Medicine. Published by Elsevier B.V. All rights reserved.

Performance of chromogenic media for Candida in rapid presumptive identification of Candida species from clinical materials

PubMed Central

Pravin Charles, M. V.; Kali, Arunava; Joseph, Noyal Mariya

2015-01-01

Background: In perspective of the worldwide increase in a number of immunocompromised patients, the need for identification of Candida species has become a major concern. The development of chromogenic differential media, introduced recently, facilitate rapid speciation. However, it can be employed for routine mycology workup only after an exhaustive evaluation of its benefit and cost effectiveness. This study was undertaken to evaluate the benefit and cost effectiveness of chromogenic media for speciation of Candida clinical isolates. Materials and Methods: Sputum samples of 382 patients were screened for the presence of Candida spp. by Gram stain and culture on sabouraud dextrose agar. Candida species were identified using Gram stain morphology, germ tube formation, cornmeal agar with Tween-80, sugar fermentation tests and morphology on HiCrome Candida differential agar. All the Candida isolates were inoculated on HiCrome Candida agar (HiMedia, Mumbai, India). Results: The sensitivity and specificity of HiCrome agar for identification of Candida albicans were 90% and 96.42%, respectively whereas sensitivity and specificity of carbohydrate fermentation test were 86.67% and 74.07%, respectively. Sensitivity and specificity values of HiCrome agar for detection of C. albicans, Candida parapsilosis and Candida glabrata were above 90%. Conclusions: We found HiCrome agar has high sensitivity and specificity comparable to that of the conventional method. In addition, use of this differential media could significantly cut down the turnaround time as well as cost of sample processing. PMID:26109791

Performance of chromogenic media for Candida in rapid presumptive identification of Candida species from clinical materials.

PubMed

Pravin Charles, M V; Kali, Arunava; Joseph, Noyal Mariya

2015-06-01

In perspective of the worldwide increase in a number of immunocompromised patients, the need for identification of Candida species has become a major concern. The development of chromogenic differential media, introduced recently, facilitate rapid speciation. However, it can be employed for routine mycology workup only after an exhaustive evaluation of its benefit and cost effectiveness. This study was undertaken to evaluate the benefit and cost effectiveness of chromogenic media for speciation of Candida clinical isolates. Sputum samples of 382 patients were screened for the presence of Candida spp. by Gram stain and culture on sabouraud dextrose agar. Candida species were identified using Gram stain morphology, germ tube formation, cornmeal agar with Tween-80, sugar fermentation tests and morphology on HiCrome Candida differential agar. All the Candida isolates were inoculated on HiCrome Candida agar (HiMedia, Mumbai, India). The sensitivity and specificity of HiCrome agar for identification of Candida albicans were 90% and 96.42%, respectively whereas sensitivity and specificity of carbohydrate fermentation test were 86.67% and 74.07%, respectively. Sensitivity and specificity values of HiCrome agar for detection of C. albicans, Candida parapsilosis and Candida glabrata were above 90%. We found HiCrome agar has high sensitivity and specificity comparable to that of the conventional method. In addition, use of this differential media could significantly cut down the turnaround time as well as cost of sample processing.

Diagnostic accuracy of a loop-mediated isothermal PCR assay for detection of Orientia tsutsugamushi during acute Scrub Typhus infection.

PubMed

Paris, Daniel H; Blacksell, Stuart D; Nawtaisong, Pruksa; Jenjaroen, Kemajittra; Teeraratkul, Achara; Chierakul, Wirongrong; Wuthiekanun, Vanaporn; Kantipong, Pacharee; Day, Nicholas P J

2011-09-01

There is an urgent need to develop rapid and accurate point-of-care (POC) technologies for acute scrub typhus diagnosis in low-resource, primary health care settings to guide clinical therapy. In this study we present the clinical evaluation of loop-mediated isothermal PCR assay (LAMP) in the context of a prospective fever study, including 161 patients from scrub typhus-endemic Chiang Rai, northern Thailand. A robust reference comparator set comprising following 'scrub typhus infection criteria' (STIC) was used: a) positive cell culture isolate and/or b) an admission IgM titer ≥1∶12,800 using the 'gold standard' indirect immunofluorescence assay (IFA) and/or c) a 4-fold rising IFA IgM titer and/or d) a positive result in at least two out of three PCR assays. Compared to the STIC criteria, all PCR assays (including LAMP) demonstrated high specificity ranging from 96-99%, with sensitivities varying from 40% to 56%, similar to the antibody based rapid test, which had a sensitivity of 47% and a specificity of 95%. The diagnostic accuracy of the LAMP assay was similar to realtime and nested conventional PCR assays, but superior to the antibody-based rapid test in the early disease course. The combination of DNA- and antibody-based detection methods increased sensitivity with minimal reduction of specificity, and expanded the timeframe of adequate diagnostic coverage throughout the acute phase of scrub typhus.

Evaluation of an immunochromatographic assay for the detection of anti-hepatitis A virus IgM

PubMed Central

2010-01-01

Background Hepatitis A virus (HAV) is a causative agent of acute hepatitis, which is transmitted by person-to-person contact and via the faecal-oral route. Acute HAV infection is usually confirmed by anti-HAV IgM detection. In order to detect anti-HAV IgM in the serum of patients infected with HAV, we developed a rapid assay based on immunochromatography (ICA) and evaluated the sensitivity of this assay by comparing it with a commercial microparticle enzyme immunoassay (MEIA) that is widely used for serological diagnosis. Results The newly developed ICA showed 100% sensitivity and specificity when used to test 150 anti-HAV IgM-positive sera collected from infected patients and 75 negative sera from healthy subjects. Also, the sensitivity of ICA is about 10 times higher than MEIA used in this study by determining end point to detect independent on infected genotype of HAV. In addition, the ICA was able to detect 1 positive sample from among 50 sera from acute hepatitis patients that had tested negative for anti-HAV IgM using the MEIA. Conclusion Conclusively, ICA for the detection of anti-HAV IgM will be very effective for rapid assay to apply clinical diagnosis and epidemiological investigation on epidemics due to the simplicity, rapidity and specificity. PMID:20637129

Sensitive Detection of Xanthomonas oryzae Pathovars oryzae and oryzicola by Loop-Mediated Isothermal Amplification

PubMed Central

Lang, Jillian M.; Langlois, Paul; Nguyen, Marian Hanna R.; Triplett, Lindsay R.; Purdie, Laura; Holton, Timothy A.; Djikeng, Appolinaire; Vera Cruz, Casiana M.; Verdier, Valérie

2014-01-01

Molecular diagnostics for crop diseases can enhance food security by enabling the rapid identification of threatening pathogens and providing critical information for the deployment of disease management strategies. Loop-mediated isothermal amplification (LAMP) is a PCR-based tool that allows the rapid, highly specific amplification of target DNA sequences at a single temperature and is thus ideal for field-level diagnosis of plant diseases. We developed primers highly specific for two globally important rice pathogens, Xanthomonas oryzae pv. oryzae, the causal agent of bacterial blight (BB) disease, and X. oryzae pv. oryzicola, the causal agent of bacterial leaf streak disease (BLS), for use in reliable, sensitive LAMP assays. In addition to pathovar distinction, two assays that differentiate X. oryzae pv. oryzae by African or Asian lineage were developed. Using these LAMP primer sets, the presence of each pathogen was detected from DNA and bacterial cells, as well as leaf and seed samples. Thresholds of detection for all assays were consistently 104 to 105 CFU ml−1, while genomic DNA thresholds were between 1 pg and 10 fg. Use of the unique sequences combined with the LAMP assay provides a sensitive, accurate, rapid, simple, and inexpensive protocol to detect both BB and BLS pathogens. PMID:24837384

Detection of Entamoeba histolytica by Recombinase Polymerase Amplification.

PubMed

Nair, Gayatri; Rebolledo, Mauricio; White, A Clinton; Crannell, Zachary; Richards-Kortum, R Rebecca; Pinilla, A Elizabeth; Ramírez, Juan David; López, M Consuelo; Castellanos-Gonzalez, Alejandro

2015-09-01

Amebiasis is an important cause of diarrheal disease worldwide and has been associated with childhood malnutrition. Traditional microscopy approaches are neither sensitive nor specific for Entamoeba histolytica. Antigen assays are more specific, but many cases are missed unless tested by molecular methods. Although polymerase chain reaction (PCR) is effective, the need for sophisticated, expensive equipment, infrastructure, and trained personnel limits its usefulness, especially in the resource-limited, endemic areas. Here, we report development of a recombinase polymerase amplification (RPA) method to detect E. histolytica specifically. Using visual detection by lateral flow (LF), the test was highly sensitive and specific and could be performed without additional equipment. The availability of this inexpensive, sensitive, and field-applicable diagnostic test could facilitate rapid diagnosis and treatment of amebiasis in endemic regions. © The American Society of Tropical Medicine and Hygiene.

Self-Reported HIV-Positive Status But Subsequent HIV-Negative Test Result Using Rapid Diagnostic Testing Algorithms Among Seven Sub-Saharan African Military Populations

DTIC Science & Technology

2017-07-07

RESEARCH ARTICLE Self-reported HIV-positive status but subsequent HIV-negative test result using rapid diagnostic testing algorithms among seven sub...America * judith.harbertson.ctr@mail.mil Abstract HIV rapid diagnostic tests (RDTs) combined in an algorithm are the current standard for HIV diagnosis...in many sub-Saharan African countries, and extensive laboratory testing has con- firmed HIV RDTs have excellent sensitivity and specificity. However

Clinical Evaluation of Rapid Diagnostic Test Kit for Scrub Typhus with Improved Performance.

PubMed

Kim, Young-Jin; Park, Sungman; Premaratna, Ranjan; Selvaraj, Stephen; Park, Sang-Jin; Kim, Sora; Kim, Donghwan; Kim, Min Soo; Shin, Dong Hoon; Choi, Kyung-Chan; Kwon, Soon-Hwan; Seo, Wonjun; Lee, Nam Taek; Kim, Seung-Han; Kang, Heui Keun; Kim, Yoon-Won

2016-08-01

Diagnosis of scrub typhus is challenging due to its more than twenty serotypes and the similar clinical symptoms with other acute febrile illnesses including leptospirosis, murine typhus and hemorrhagic fever with renal syndrome. Accuracy and rapidity of a diagnostic test to Orientia tsutsugamushi is an important step to diagnose this disease. To discriminate scrub typhus from other diseases, the improved ImmuneMed Scrub Typhus Rapid Diagnostic Test (RDT) was evaluated in Korea and Sri Lanka. The sensitivity at the base of each IgM and IgG indirect immunofluorescent assay (IFA) in Korean patients was 98.6% and 97.1%, and the specificity was 98.2% and 97.7% respectively. The sensitivity and specificity for retrospective diagnosis at the base of IFA in Sri Lanka was 92.1% and 96.1%. ImmuneMed RDT was not reactive to any serum from seventeen diseases including hemorrhagic fever with renal syndrome (n = 48), leptospirosis (n = 23), and murine typhus (n = 48). ImmuneMed RDT shows superior sensitivity (98.6% and 97.1%) compared with SD Bioline RDT (84.4% at IgM and 83.3% at IgG) in Korea. The retrospective diagnosis of ImmuneMed RDT exhibits 94.0% identity with enzyme-linked Immunosorbent assay (ELISA) using South India patient serum samples. These results suggest that this RDT can replace other diagnostic tests and is applicable for global diagnosis of scrub typhus. This rapid and accurate diagnosis will be beneficial for diagnosing and managing scrub typhus.

Ultrasound Fracture Diagnosis in Space

NASA Technical Reports Server (NTRS)

Dulchavsky, Scott A.; Amponsah, David; Sargsyan, Ashot E.; Garcia, Kathleen M.; Hamilton, Douglas R.; vanHolsbeeck, Marnix

2010-01-01

Introduction: This ground-based investigation accumulated high-level clinical evidence on the sensitivity and specificity of point of care ultrasound performed by expert and novice users for the rapid diagnosis of musculoskeletal (MSK) injuries. We developed preliminary educational methodologies to provide just-in-time training of novice users by creating multi-media training tools and imaging procedures for non expert operators and evaluated the sensitivity and specificity of non-expert performed musculoskeletal ultrasound to diagnose acute injuries in a Level 1 Trauma Center. Methods: Patients with potential MSK injuries were identified in the emergency room. A focused MSK ultrasound was performed by expert operators and compared to standard radiographs. A repeat examination was performed by non-expert operators who received a short, just-in-time multimedia education aid. The sensitivity and specificity of the expert and novice ultrasound examinations were compared to gold standard radiography. Results: Over 800 patients were enrolled in this study. The sensitivity and specificity of expert performed ultrasound exceeded 98% for MSK injuries. Novice operators achieved 97% sensitivity and 99% specificity for targeted examinations with the greatest error in fractures involving the hand and foot. Conclusion: Point of care ultrasound is a sensitive and specific diagnostic test for MSK injury when performed by experts and just-in-time trained novice operators.

Specific and straightforward molecular investigation of β-thalassemia mutations in the Malaysian Malays and Chinese using direct TaqMan genotyping assays.

PubMed

Kho, S L; Chua, K H; George, E; Tan, J A M A

2013-07-15

Beta-thalassemia is a life-threatening inherited blood disorder. Rapid characterization of β-globin gene mutations is necessary because of the high frequency of Malaysian β-thalassemia carriers. A combination real-time polymerase chain reaction genotyping assay using TaqMan probes was developed to confirm β-globin gene mutations. In this study, primers and probes were designed to specifically identify 8 common β-thalassemia mutations in the Malaysian Malay and Chinese ethnic groups using the Primer Express software. "Blind tests" using DNA samples from healthy individuals and β-thalassemia patients with different genotypes were performed to determine the specificity and sensitivity of this newly designed assay. Our results showed 100% sensitivity and specificity for this novel assay. In conclusion, the TaqMan genotyping assay is a straightforward assay that allows detection of β-globin gene mutations in less than 40 min. The simplicity and reproducibility of the TaqMan genotyping assay permit its use in laboratories as a rapid and cost-effective diagnostic tool for confirmation of common β-thalassemia mutations in Malaysia.

A rapid and simple method of detection of Blepharisma japonicum using PCR and immobilisation on FTA paper

PubMed Central

Hide, Geoff; Hughes, Jacqueline M; McNuff, Robert

2003-01-01

Background The rapid expansion in the availability of genome and DNA sequence information has opened up new possibilities for the development of methods for detecting free-living protozoa in environmental samples. The protozoan Blepharisma japonicum was used to investigate a rapid and simple detection system based on polymerase chain reaction amplification (PCR) from organisms immobilised on FTA paper. Results Using primers designed from the α-tubulin genes of Blepharisma, specific and sensitive detection to the equivalent of a single Blepharisma cell could be achieved. Similar detection levels were found using water samples, containing Blepharisma, which were dried onto Whatman FTA paper. Conclusion This system has potential as a sensitive convenient detection system for Blepharisma and could be applied to other protozoan organisms. PMID:14516472

Performance characteristics of magnetic resonance imaging without contrast agents or sedation in pediatric appendicitis.

PubMed

Didier, Ryne A; Hopkins, Katharine L; Coakley, Fergus V; Krishnaswami, Sanjay; Spiro, David M; Foster, Bryan R

2017-09-01

Magnetic resonance imaging (MRI) has emerged as a promising modality for evaluating pediatric appendicitis. However optimal imaging protocols, including roles of contrast agents and sedation, have not been established and diagnostic criteria have not been fully evaluated. To investigate performance characteristics of rapid MRI without contrast agents or sedation in the diagnosis of pediatric appendicitis. We included patients ages 4-18 years with suspicion of appendicitis who underwent rapid MRI between October 2013 and March 2015 without contrast agent or sedation. After two-radiologist review, we determined performance characteristics of individual diagnostic criteria and aggregate diagnostic criteria by comparing MRI results to clinical outcomes. We used receiver operating characteristic (ROC) curves to determine cut-points for appendiceal diameter and wall thickness for optimization of predictive power, and we calculated area under the curve (AUC) as a measure of test accuracy. Ninety-eight MRI examinations were performed in 97 subjects. Overall, MRI had a 94% sensitivity, 95% specificity, 91% positive predictive value and 97% negative predictive value. Optimal cut-points for appendiceal diameter and wall thickness were ≥7 mm and ≥2 mm, respectively. Independently, those cut-points produced sensitivities of 91% and 84% and specificities of 84% and 43%. Presence of intraluminal fluid (30/33) or localized periappendiceal fluid (32/33) showed a significant association with acute appendicitis (P<0.01), with sensitivities of 91% and 97% and specificities of 60% and 50%. For examinations in which the appendix was not identified by one or both reviewers (23/98), the clinical outcome was negative. Rapid MRI without contrast agents or sedation is accurate for diagnosis of pediatric appendicitis when multiple diagnostic criteria are considered in aggregate. Individual diagnostic criteria including optimized cut-points of ≥7 mm for diameter and ≥2 mm for wall thickness demonstrate high sensitivities but relatively low specificities. Nonvisualization of the appendix favors a negative diagnosis.

The sensitivity and specificity of a urine based Rapid Diagnostic Test for the diagnosis of plasmodium falciparum in a malaria endemic area in Odisha, India.

PubMed

Samal, Ajit Gopal; Behera, Prativa Kumari; Mohanty, Akshay Kumar; Satpathi, Sanghamitra; Kumar, Abhishek; Panda, Rabi Ratna; Minz, Aruna Mukti; Mohanty, Sanjib; Samal, Abhijit; Van Der Pluijm, Rob W

2017-10-01

Rapid and accurate diagnosis is crucial in the treatment of malaria. Rapid Diagnostic Tests (RDTs) using blood have been recommended by the WHO as an acceptable method for the diagnosis of malaria. RDTs provide results quickly, is simple to use and easy to interpret. However, its use requires collection of blood by skin puncture. Hence the aim of the pilot study is to explore the sensitivity and specificity of RDTs using urine (collected non-invasively) for diagnosis of Plasmodium falciparum malaria and to assess the relation between parasite density in blood with HRP-2 Ag detection in urine. All fever cases admitted to Ispat General Hospital (IGH) Rourkela, India, during June 2012-March 2013 with a clinical diagnosis of malaria were examined for the presence of asexual forms of P. falciparum in peripheral blood smears. All smear positive febrile patients who met the eligibility criteria were enrolled. Smear negative fever cases were enrolled as control cases. RDTs were performed using both urine and blood samples by using commercially available blood specific kits. Sixty blood smear positive cases and 51 febrile blood smear negative cases were enrolled. Sensitivity and specificity of RDT urine were 86.67% (95%CI:75.83-93.09) and 94.12% (95%CI:84.08-97.98) respectively whereas those of RDT blood were 91.67% (95% CI: 81.93-96.39) and 98.04% (95% CI 89.7-99.65). The sensitivity of both RDT urine as well as RDT blood were found to be dependent on the level of parasitemia. Results of this study are promising. Larger studies are needed to assess whether RDTs using urine could serve as a practical, reliable method for the detection of P. falciparum in a non-invasive manner where invasive blood taking is less feasible.

Detection of shigella in lettuce by the use of a rapid molecular assay with increased sensitivity

PubMed Central

Jiménez, Kenia Barrantes; McCoy², Clyde B.; Achí, Rosario

2010-01-01

A Multiplex Polymerase Chain Reaction (PCR) assay to be used as an alternative to the conventional culture method in detecting Shigella and enteroinvasive Escherichia coli (EIEC) virulence genes ipaH and ial in lettuce was developed. Efficacy and rapidity of the molecular method were determined as compared to the conventional culture. Lettuce samples were inoculated with different Shigella flexneri concentrations (from 10 CFU/ml to 107 CFU/ml). DNA was extracted directly from lettuce after inoculation (direct-PCR) and after an enrichment step (enrichment PCR). Multiplex PCR detection limit was 104CFU/ml, diagnostic sensitivity and specificity were 100% accurate. An internal amplification control (IAC) of 100 bp was used in order to avoid false negative results. This method produced results in 1 to 2 days while the conventional culture method required 5 to 6 days. Also, the culture method detection limit was 106 CFU/ml, diagnostic sensitivity was 53% and diagnostic specificity was 100%. In this study a Multiplex PCR method for detection of virulence genes in Shigella and EIEC was shown to be effective in terms of diagnostic sensitivity, detection limit and amount of time as compared to Shigella conventional culture. PMID:24031579

Plasmonic Nanobubbles Rapidly Detect and Destroy Drug-Resistant Tumors

PubMed Central

Lukianova-Hleb, Ekaterina Y.; Ren, Xiaoyang; Townley, Debra; Wu, Xiangwei; Kupferman, Michael E.; Lapotko, Dmitri O.

2012-01-01

The resistance of residual cancer cells after oncological resection to adjuvant chemoradiotherapies results in both high recurrence rates and high non-specific tissue toxicity, thus preventing the successful treatment of such cancers as head and neck squamous cell carcinoma (HNSCC). The patients' survival rate and quality of life therefore depend upon the efficacy, selectivity and low non-specific toxicity of the adjuvant treatment. We report a novel, theranostic in vivo technology that unites both the acoustic diagnostics and guided intracellular delivery of anti-tumor drug (liposome-encapsulated doxorubicin, Doxil) in one rapid process, namely a pulsed laser-activated plasmonic nanobubble (PNB). HNSCC-bearing mice were treated with gold nanoparticle conjugates, Doxil, and single near-infrared laser pulses of low energy. Tumor-specific clusters of gold nanoparticles (solid gold spheres) converted the optical pulses into localized PNBs. The acoustic signals of the PNB detected the tumor with high specificity and sensitivity. The mechanical impact of the PNB, co-localized with Doxil liposomes, selectively ejected the drug into the cytoplasm of cancer cells. Cancer cell-specific generation of PNBs and their intracellular co-localization with Doxil improved the in vivo therapeutic efficacy from 5-7% for administration of only Doxil or PNBs alone to 90% thus demonstrating the synergistic therapeutic effect of the PNB-based intracellular drug release. This mechanism also reduced the non-specific toxicity of Doxil below a detectable level and the treatment time to less than one minute. Thus PNBs combine highly sensitive diagnosis, overcome drug resistance and minimize non-specific toxicity in a single rapid theranostic procedure for intra-operative treatment. PMID:23139725

An insulated isothermal PCR method on a field-deployable device for rapid and sensitive detection of canine parvovirus type 2 at points of need.

PubMed

Wilkes, Rebecca P; Lee, Pei-Yu A; Tsai, Yun-Long; Tsai, Chuan-Fu; Chang, Hsiu-Hui; Chang, Hsiao-Fen G; Wang, Hwa-Tang T

2015-08-01

Canine parvovirus type 2 (CPV-2), including subtypes 2a, 2b and 2c, causes an acute enteric disease in both domestic and wild animals. Rapid and sensitive diagnosis aids effective disease management at points of need (PON). A commercially available, field-deployable and user-friendly system, designed with insulated isothermal PCR (iiPCR) technology, displays excellent sensitivity and specificity for nucleic acid detection. An iiPCR method was developed for on-site detection of all circulating CPV-2 strains. Limit of detection was determined using plasmid DNA. CPV-2a, 2b and 2c strains, a feline panleukopenia virus (FPV) strain, and nine canine pathogens were tested to evaluate assay specificity. Reaction sensitivity and performance were compared with an in-house real-time PCR using serial dilutions of a CPV-2b strain and 100 canine fecal clinical samples collected from 2010 to 2014, respectively. The 95% limit of detection of the iiPCR method was 13 copies of standard DNA and detection limits for CPV-2b DNA were equivalent for iiPCR and real-time PCR. The iiPCR reaction detected CPV-2a, 2b and 2c and FPV. Non-targeted pathogens were not detected. Test results of real-time PCR and iiPCR from 99 fecal samples agreed with each other, while one real-time PCR-positive sample tested negative by iiPCR. Therefore, excellent agreement (k = 0.98) with sensitivity of 98.41% and specificity of 100% in detecting CPV-2 in feces was found between the two methods. In conclusion, the iiPCR system has potential to serve as a useful tool for rapid and accurate PON, molecular detection of CPV-2. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

[Multiplex real-time PCR method for rapid detection of Marburg virus and Ebola virus].

PubMed

Yang, Yu; Bai, Lin; Hu, Kong-Xin; Yang, Zhi-Hong; Hu, Jian-Ping; Wang, Jing

2012-08-01

Marburg virus and Ebola virus are acute infections with high case fatality rates. A rapid, sensitive detection method was established to detect Marburg virus and Ebola virus by multiplex real-time fluorescence quantitative PCR. Designing primers and Taqman probes from highly conserved sequences of Marburg virus and Ebola virus through whole genome sequences alignment, Taqman probes labeled by FAM and Texas Red, the sensitivity of the multiplex real-time quantitative PCR assay was optimized by evaluating the different concentrations of primers and Probes. We have developed a real-time PCR method with the sensitivity of 30.5 copies/microl for Marburg virus positive plasmid and 28.6 copies/microl for Ebola virus positive plasmids, Japanese encephalitis virus, Yellow fever virus, Dengue virus were using to examine the specificity. The Multiplex real-time PCR assays provide a sensitive, reliable and efficient method to detect Marburg virus and Ebola virus simultaneously.

Targeting Conserved Genes in Fusarium Species.

PubMed

Gil-Serna, Jéssica; Patiño, Belén; Jurado, Miguel; Mirete, Salvador; Vázquez, Covadonga; González-Jaén, M Teresa

2017-01-01

Fumonisins are important mycotoxins contaminating foods and feeds which are mainly produced by F. verticillioides and F. proliferatum. Additionally, both are pathogens of maize and other cereals. We describe two highly sensitive, rapid, and species-specific PCR protocols which enable detection and discrimination of these closely related species in cereal flour or grain samples. The specific primer pairs of these assays were based on the intergenic spacer region of the multicopy rDNA unit which highly improves the sensitivity of the PCR assay in comparison with single-copy target regions.

Use of synthetic oligonucleotide DNA probes for the identification of Bacteroides gingivalis.

PubMed Central

Moncla, B J; Braham, P; Dix, K; Watanabe, S; Schwartz, D

1990-01-01

Six different oligonucleotide probes complementary to the hypervariable regions of 16S rRNA of Bacteroides gingivalis were tested for specificity and sensitivity against 77 field strains of B. gingivalis and 105 strains of 12 other Bacteroides species. The data demonstrated that these probes were very specific (range, 0.85 to 1.00) and sensitive (1.00). Some limited cross-reactions with other Bacteroides species were observed. Four of these probes should be useful for rapid detection and identification of B. gingivalis. Images PMID:1690217

Neptune: a bioinformatics tool for rapid discovery of genomic variation in bacterial populations

PubMed Central

Marinier, Eric; Zaheer, Rahat; Berry, Chrystal; Weedmark, Kelly A.; Domaratzki, Michael; Mabon, Philip; Knox, Natalie C.; Reimer, Aleisha R.; Graham, Morag R.; Chui, Linda; Patterson-Fortin, Laura; Zhang, Jian; Pagotto, Franco; Farber, Jeff; Mahony, Jim; Seyer, Karine; Bekal, Sadjia; Tremblay, Cécile; Isaac-Renton, Judy; Prystajecky, Natalie; Chen, Jessica; Slade, Peter

2017-01-01

Abstract The ready availability of vast amounts of genomic sequence data has created the need to rethink comparative genomics algorithms using ‘big data’ approaches. Neptune is an efficient system for rapidly locating differentially abundant genomic content in bacterial populations using an exact k-mer matching strategy, while accommodating k-mer mismatches. Neptune’s loci discovery process identifies sequences that are sufficiently common to a group of target sequences and sufficiently absent from non-targets using probabilistic models. Neptune uses parallel computing to efficiently identify and extract these loci from draft genome assemblies without requiring multiple sequence alignments or other computationally expensive comparative sequence analyses. Tests on simulated and real datasets showed that Neptune rapidly identifies regions that are both sensitive and specific. We demonstrate that this system can identify trait-specific loci from different bacterial lineages. Neptune is broadly applicable for comparative bacterial analyses, yet will particularly benefit pathogenomic applications, owing to efficient and sensitive discovery of differentially abundant genomic loci. The software is available for download at: http://github.com/phac-nml/neptune. PMID:29048594

Rapid and label-free detection of protein a by aptamer-tethered porous silicon nanostructures.

PubMed

Urmann, Katharina; Reich, Peggy; Walter, Johanna-Gabriela; Beckmann, Dieter; Segal, Ester; Scheper, Thomas

2017-09-10

Protein A, which is secreted by and displayed on the cell membrane of Staphylococcus aureus is an important biomarker for S. aureus. Thus, its rapid and specific detection may facilitate the pathogen identification and initiation of proper treatment. Herein, we present a simple, label-free and rapid optical biosensor enabling specific detection of protein A. Protein A-binding aptamer serves as the capture probe and is immobilized onto a nanostructured porous silicon thin film, which serves as the optical transducer element. We demonstrate high sensitivity of the biosensor with a linear detection range between 8 and 23μM. The apparent dissociation constant was determined as 13.98μM and the LoD is 3.17μM. Harnessing the affinity between protein A and antibodies, a sandwich assay format was developed to amplify the optical signal associated with protein A capture by the aptamer. Using this approach, we increase the sensitivity of the biosensor, resulting in a three times lower LoD. Copyright © 2017 Elsevier B.V. All rights reserved.

Surface enhanced Raman spectroscopy (SERS) for in vitro diagnostic testing at the point of care

NASA Astrophysics Data System (ADS)

Marks, Haley; Schechinger, Monika; Garza, Javier; Locke, Andrea; Coté, Gerard

2017-06-01

Point-of-care (POC) device development is a growing field that aims to develop low-cost, rapid, sensitive in-vitro diagnostic testing platforms that are portable, self-contained, and can be used anywhere - from modern clinics to remote and low resource areas. In this review, surface enhanced Raman spectroscopy (SERS) is discussed as a solution to facilitating the translation of bioanalytical sensing to the POC. The potential for SERS to meet the widely accepted "ASSURED" (Affordable, Sensitive, Specific, User-friendly, Rapid, Equipment-free, and Deliverable) criterion provided by the World Health Organization is discussed based on recent advances in SERS in vitro assay development. As SERS provides attractive characteristics for multiplexed sensing at low concentration limits with a high degree of specificity, it holds great promise for enhancing current efforts in rapid diagnostic testing. In outlining the progression of SERS techniques over the past years combined with recent developments in smart nanomaterials, high-throughput microfluidics, and low-cost paper diagnostics, an extensive number of new possibilities show potential for translating SERS biosensors to the POC.

Validation and clinical application of a molecular method for the identification of Cryptococcus neoformans/Cryptococcus gattii complex DNA in human clinical specimens.

PubMed

Rivera, Vanessa; Gaviria, Marcela; Muñoz-Cadavid, Cesar; Cano, Luz; Naranjo, Tonny

2015-01-01

The diagnosis of cryptococcosis is usually performed based on cultures of tissue or body fluids and isolation of the fungus, but this method may require several days. Direct microscopic examination, although rapid, is relatively insensitive. Biochemical and immunodiagnostic rapid tests are also used. However, all of these methods have limitations that may hinder final diagnosis. The increasing incidence of fungal infections has focused attention on tools for rapid and accurate diagnosis using molecular biological techniques. Currently, PCR-based methods, particularly nested, multiplex and real-time PCR, provide both high sensitivity and specificity. In the present study, we evaluated a nested PCR targeting the gene encoding the ITS-1 and ITS-2 regions of rDNA in samples from a cohort of patients diagnosed with cryptococcosis. The results showed that in our hands, this Cryptococcus nested PCR assay has 100% specificity and 100% sensitivity and was able to detect until 2 femtograms of Cryptococcus DNA. Copyright © 2015 Elsevier Editora Ltda. All rights reserved.

[Diagnostic tools for canine parvovirus infection].

PubMed

Proksch, A L; Hartmann, K

2015-01-01

Canine parvovirus (CPV) infection is one of the most important and common infectious diseases in dogs, in particular affecting young puppies when maternal antibodies have waned and vaccine-induced antibodies have not yet developed. The mortality rate remains high. Therefore, a rapid and safe diagnostic tool is essential to diagnose the disease to 1) provide intensive care treatment and 2) to identify virus-shedding animals and thus prevent virus spread. Whilst the detection of antibodies against CPV is considered unsuitable to diagnose the disease, there are several different methods to directly detect complete virus, virus antigen or DNA. Additionally, to test in commercial laboratories, rapid in-house tests based on ELISA are available worldwide. The specificity of the ELISA rapid in-house tests is reported to be excellent. However, results on sensitivity vary and high numbers of false-negative results are commonly reported, which potentially leads to misdiagnosis. Polymerase chain reaction (PCR) is a very sensitive and specific diagnostic tool. It also provides the opportunity to differentiate vaccine strains from natural infection when sequencing is performed after PCR.

Development of 2, 7-Diamino-1, 8-Naphthyridine (DANP) Anchored Hairpin Primers for RT-PCR Detection of Chikungunya Virus Infection

PubMed Central

Chen, Huixin; Parimelalagan, Mariya; Takei, Fumie; Hapuarachchi, Hapuarachchige Chanditha; Koay, Evelyn Siew-Chuan; Ng, Lee Ching; Ho, Phui San; Nakatani, Kazuhiko; Chu, Justin Jang Hann

2016-01-01

A molecular diagnostic platform with DANP-anchored hairpin primer was developed and evaluated for the rapid and cost-effective detection of Chikungunya virus (CHIKV) with high sensitivity and specificity. The molecule 2, 7-diamino-1, 8-naphthyridine (DANP) binds to a cytosine-bulge and emits fluorescence at 450 nm when it is excited by 400 nm light. Thus, by measuring the decline in fluorescence emitted from DANP—primer complexes after PCR reaction, we could monitor the PCR progress. By adapting this property of DANP, we have previously developed the first generation DANP-coupled hairpin RT-PCR assay. In the current study, we improved the assay performance by conjugating the DANP molecule covalently onto the hairpin primer to fix the DANP/primer ratio at 1:1; and adjusting the excitation emission wavelength to 365/430 nm to minimize the background signal and a ‘turn-on’ system is achieved. After optimizing the PCR cycle number to 30, we not only shortened the total assay turnaround time to 60 minutes, but also further reduced the background fluorescence. The detection limit of our assay was 0.001 PFU per reaction. The DANP-anchored hairpin primer, targeting nsP2 gene of CHIKV genome, is highly specific to CHIKV, having no cross-reactivity to a panel of other RNA viruses tested. In conclusion, we report here a molecular diagnostic assay that is sensitive, specific, rapid and cost effective for CHIKV detection and can be performed where no real time PCR instrumentation is required. Our results from patient samples indicated 93.62% sensitivity and 100% specificity of this method, ensuring that it can be a useful tool for rapid detection of CHIKV for outbreaks in many parts of the world. PMID:27571201

Development of 2, 7-Diamino-1, 8-Naphthyridine (DANP) Anchored Hairpin Primers for RT-PCR Detection of Chikungunya Virus Infection.

PubMed

Chen, Huixin; Parimelalagan, Mariya; Takei, Fumie; Hapuarachchi, Hapuarachchige Chanditha; Koay, Evelyn Siew-Chuan; Ng, Lee Ching; Ho, Phui San; Nakatani, Kazuhiko; Chu, Justin Jang Hann

2016-08-01

A molecular diagnostic platform with DANP-anchored hairpin primer was developed and evaluated for the rapid and cost-effective detection of Chikungunya virus (CHIKV) with high sensitivity and specificity. The molecule 2, 7-diamino-1, 8-naphthyridine (DANP) binds to a cytosine-bulge and emits fluorescence at 450 nm when it is excited by 400 nm light. Thus, by measuring the decline in fluorescence emitted from DANP-primer complexes after PCR reaction, we could monitor the PCR progress. By adapting this property of DANP, we have previously developed the first generation DANP-coupled hairpin RT-PCR assay. In the current study, we improved the assay performance by conjugating the DANP molecule covalently onto the hairpin primer to fix the DANP/primer ratio at 1:1; and adjusting the excitation emission wavelength to 365/430 nm to minimize the background signal and a 'turn-on' system is achieved. After optimizing the PCR cycle number to 30, we not only shortened the total assay turnaround time to 60 minutes, but also further reduced the background fluorescence. The detection limit of our assay was 0.001 PFU per reaction. The DANP-anchored hairpin primer, targeting nsP2 gene of CHIKV genome, is highly specific to CHIKV, having no cross-reactivity to a panel of other RNA viruses tested. In conclusion, we report here a molecular diagnostic assay that is sensitive, specific, rapid and cost effective for CHIKV detection and can be performed where no real time PCR instrumentation is required. Our results from patient samples indicated 93.62% sensitivity and 100% specificity of this method, ensuring that it can be a useful tool for rapid detection of CHIKV for outbreaks in many parts of the world.

Sensitivity, specificity, and confounding factors of novel serological tests used for the rapid diagnosis of bovine tuberculosis in farmed red deer (Cervus elaphus).

PubMed

Buddle, Bryce M; Wilson, Tania; Denis, Michel; Greenwald, Rena; Esfandiari, Javan; Lyashchenko, Konstantin P; Liggett, Simon; Mackintosh, Colin G

2010-04-01

In this study, novel serological tests were used to detect tuberculosis (TB) in groups of farmed red deer (Cervus elaphus) varying in disease status or possible confounding factors. Groups of deer naturally or experimentally infected with Mycobacterium bovis and animals vaccinated against paratuberculosis were studied, as were uninfected animals and animals naturally or experimentally infected with Mycobacterium avium subsp. paratuberculosis. Sera were assayed using two rapid lateral-flow tests, Chembio's CervidTB STAT-PAK and DPP VetTB tests, and results were compared to those from tuberculin skin tests. Both serological tests had a high sensitivity, but specificity was adversely affected after animals had received a vaccine against paratuberculosis and were subsequently skin tested. The specificity of the DPP VetTB test was higher than that of the CervidTB STAT-PAK test, with natural infection with M. avium subsp. paratuberculosis adversely affecting the specificity of only the CervidTB STAT-PAK test. The sera from M. avium subsp. paratuberculosis-infected deer that produced false-positive reactions in the CervidTB STAT-PAK test were retested with a multiantigen print immunoassay (MAPIA), and some of these sera were shown to react with the MPB83 antigen. Combining the results from the serological tests and the skin tests showed only a slight increase in the sensitivity of detection of M. bovis-infected animals. It is concluded that both the CervidTB STAT-PAK and DPP VetTB tests offer rapid, convenient, and easy detection of bovine tuberculosis in deer, albeit with significant interference from paratuberculosis vaccination status and subsequent skin testing. The latter finding illustrates one of the limitations of currently available vaccines against paratuberculosis.

Quantification of Kryptofix 2.2.2 in [18F]fluorine-labelled radiopharmaceuticals by rapid-resolution liquid chromatography.

PubMed

Lao, Yexing; Yang, Cuiping; Zou, Wei; Gan, Manquan; Chen, Ping; Su, Weiwei

2012-05-01

The cryptand Kryptofix 2.2.2 is used extensively as a phase-transfer reagent in the preparation of [18F]fluoride-labelled radiopharmaceuticals. However, it has considerable acute toxicity. The aim of this study was to develop and validate a method for rapid (within 1 min), specific and sensitive quantification of Kryptofix 2.2.2 at trace levels. Chromatographic separations were carried out by rapid-resolution liquid chromatography (Agilent ZORBAX SB-C18 rapid-resolution column, 2.1 × 30 mm, 3.5 μm). Tandem mass spectra were acquired using a triple quadrupole mass spectrometer equipped with an electrospray ionization interface. Quantitative mass spectrometric analysis was conducted in positive ion mode and multiple reaction monitoring mode for the m/z 377.3 → 114.1 transition for Kryptofix 2.2.2. The external standard method was used for quantification. The method met the precision and efficiency requirements for PET radiopharmaceuticals, providing satisfactory results for specificity, matrix effect, stability, linearity (0.5-100 ng/ml, r(2)=0.9975), precision (coefficient of variation < 5%), accuracy (relative error < ± 3%), sensitivity (lower limit of quantification=0.5 ng) and detection time (<1 min). Fluorodeoxyglucose (n=6) was analysed, and the Kryptofix 2.2.2 content was found to be well below the maximum permissible levels approved by the US Food and Drug Administration. The developed method has a short analysis time (<1 min) and high sensitivity (lower limit of quantification=0.5 ng/ml) and can be successfully applied to rapid quantification of Kryptofix 2.2.2 at trace levels in fluorodeoxyglucose. This method could also be applied to other [18F]fluorine-labelled radiopharmaceuticals that use Kryptofix 2.2.2 as a phase-transfer reagent.

Development of a rapid, sensitive and specific diagnostic assay for fish Aquareovirus based on RT-PCR.

PubMed

Seng, E K; Fang, Q; Lam, T J; Sin, Y M

2004-06-15

A rapid, sensitive and highly specific detection method for Aquareovirus based on reverse-transcription polymerase chain reaction (RT-PCR) was developed. Based on multiple sequence alignment of the cloned sequences of a local isolates, the Threadfin reovirus (TFV) and Guppy reovirus (GPV) with Grass carp reovirus (GCRV), a pair of degenerate primers was selected carefully and synthesized. Using this primer combination, only one specific product, approximately 450 bp in length was obtained when RT-PCR was carried out using the genomic double-stranded RNA (dsRNA) of TFV, GPV and GCRV. Similar results were also obtained when Chum salmon reovirus (CSRV) and Striped bass reovirus (SBRV) dsRNA were used as templates. No products were observed when nucleic acids other than the dsRNA of the aquareoviruses described above were used as RT-PCR templates. This technique could detect not only TFV but also GPV and GCRV in low titer virus-infected cell cultured cells. Furthermore, this method has also been shown to be able to diagnose GPV-infected guppy (Poecilia reticulata) that exhibit clinical symptoms as well as GPV-carrier guppy. Collectively, these results showed that the RT-PCR amplification method using specific degenerate primers described below is very useful for rapid and accurate detection of a variety of aquareovirus strains isolated from different host species and origin.

Gold nanoprobe functionalized with specific fusion protein selection from phage display and its application in rapid, selective and sensitive colorimetric biosensing of Staphylococcus aureus.

PubMed

Liu, Pei; Han, Lei; Wang, Fei; Petrenko, Valery A; Liu, Aihua

2016-08-15

Staphylococcus aureus (S. aureus) is one of the most ubiquitous pathogens in public healthcare worldwide. It holds great insterest in establishing robust analytical method for S. aureus. Herein, we report a S. aureus-specific recognition element, isolated from phage monoclone GQTTLTTS, which was selected from f8/8 landscape phage library against S. aureus in a high-throughput way. By functionalizing cysteamine (CS)-stabilized gold nanoparticles (CS-AuNPs) with S. aureus-specific pVIII fusion protein (fusion-pVIII), a bifunctional nanoprobe (CS-AuNPs@fusion-pVIII) for S. aureus was developed. In this strategy, the CS-AuNPs@fusion-pVIII could be induced to aggregate quickly in the presence of target S. aureus, resulting in a rapid colorimetric response of gold nanoparticles. More importantly, the as-designed probe exhibited excellent selectivity over other bacteria. Thus, the CS-AuNPs@fusion-pVIII could be used as the indicator of target S. aureus. This assay can detect as low as 19CFUmL(-1)S. aureus within 30min. Further, this approach can be applicable to detect S. aureus in real water samples. Due to its sensitivity, specificity and rapidness, this proposed method is promising for on-site testing of S. aureus without using any costly instruments. Copyright © 2016 Elsevier B.V. All rights reserved.

A DNA Barcode-Based RPA Assay (BAR-RPA) for Rapid Identification of the Dry Root of Ficus hirta (Wuzhimaotao).

PubMed

Tian, Enwei; Liu, Qianqian; Ye, Haoting; Li, Fang; Chao, Zhi

2017-12-18

Background: Wuzhimaotao (the dry root of Ficus hirta ) is used as both medicine and food ingredient by the locals in areas around Nanling Mountains of China. Due to its very similar external morphologies with Duanchangcao (the root of Gelsemium elegans , which contains gelsemine that is extremely neurotoxic) and the associated growth of these two plants, incidents of food poisoning and even death frequently occur, resulting from the misuse of Duanchangcao as Wuzhimaotao. The aim of this study is to develop a fast, even, on-spot approach to identification of Wuzhimaotao. Methods: We used DNA barcode-based recombinase polymerase amplification (BAR-RPA) with species-specific primers targeting the internal transcribed spacer (ITS) region of the rDNA of F. hirta. BAR-RPA reaction time and temperature were optimized and the specificity and sensitivity of BAR-RPA species-specific primers were assessed. Results: This technique showed a high specificity and sensitivity to amplify the genomic DNA of F. hirta and allowed for rapid amplification (within 15 min) of the ITS region under a constant and mild temperature range of 37-42 °C without using thermocyclers. Conclusions: The BAR-RPA assay with a fast DNA extraction protocol provides a simple, energy-saving, and rapid method for identification of Wuzhimaotao in both laboratory and field settings.

Standardization of a two-step real-time polymerase chain reaction based method for species-specific detection of medically important Aspergillus species.

PubMed

Das, P; Pandey, P; Harishankar, A; Chandy, M; Bhattacharya, S; Chakrabarti, A

2017-01-01

Standardization of Aspergillus polymerase chain reaction (PCR) poses two technical challenges (a) standardization of DNA extraction, (b) optimization of PCR against various medically important Aspergillus species. Many cases of aspergillosis go undiagnosed because of relative insensitivity of conventional diagnostic methods such as microscopy, culture or antigen detection. The present study is an attempt to standardize real-time PCR assay for rapid sensitive and specific detection of Aspergillus DNA in EDTA whole blood. Three nucleic acid extraction protocols were compared and a two-step real-time PCR assay was developed and validated following the recommendations of the European Aspergillus PCR Initiative in our setup. In the first PCR step (pan-Aspergillus PCR), the target was 28S rDNA gene, whereas in the second step, species specific PCR the targets were beta-tubulin (for Aspergillus fumigatus, Aspergillus flavus, Aspergillus terreus), gene and calmodulin gene (for Aspergillus niger). Species specific identification of four medically important Aspergillus species, namely, A. fumigatus, A. flavus, A. niger and A. terreus were achieved by this PCR. Specificity of the PCR was tested against 34 different DNA source including bacteria, virus, yeast, other Aspergillus sp., other fungal species and for human DNA and had no false-positive reactions. The analytical sensitivity of the PCR was found to be 102 CFU/ml. The present protocol of two-step real-time PCR assays for genus- and species-specific identification for commonly isolated species in whole blood for diagnosis of invasive Aspergillus infections offers a rapid, sensitive and specific assay option and requires clinical validation at multiple centers.

Novel aptamer-linked nanoconjugate approach for detection of waterborne bacterial pathogens: an update

NASA Astrophysics Data System (ADS)

Singh, Gulshan; Manohar, Murli; Adegoke, Anthony Ayodeji; Stenström, Thor Axel; Shanker, Rishi

2017-01-01

The lack of microbiologically safe water in underdeveloped nations is the prime cause of infectious disease outbreaks. The need for the specific identification and detection of microorganisms encourages the development of advanced, rapid, sensitive and highly specific methods for the monitoring of pathogens and management of potential risk to human health. The rapid molecular assays based on detection of specific molecular signatures offer advantages over conventional methods in terms of specificity and sensitivity but require complex instrumentation and skilled personnel. Nanotechnology is an emerging area and provides a robust approach for the identification of pathogenic microorganism utilizing the peculiar properties of nanomaterials, i.e. small size (1-100 nm) and large surface area. This emerging technology promises to fulfill the urgent need of a novel strategy to enhance the bacterial identification and quantitation in the environment. In this context, the peculiar properties of gold nanoparticles, their plasmonic shifts, and changes in magnetic properties have been utilized for the simple and cost-effective detection of bacterial nucleic acids, antigens and toxins with quite improved sensitivity. One of the promising leads to develop an advance detection method might be the coupling of nucleic acid aptamers (capable of interacting specifically with bacteria, protozoa, and viruses) with nanomaterials. Such aptamer-nano conjugate can be used for the specific recognition of infectious agents in different environmental matrices. This review summarizes the application of nanotechnology in the area of pathogen detection and discusses the prospects of coupling nucleic acid aptamers with nanoparticles for the specific detection of targeted pathogens.

Hepatitis B virus surface antigen and anti-hepatitis C virus rapid tests underestimate hepatitis prevalence among HIV-infected patients.

PubMed

Hønge, Bl; Jespersen, S; Medina, C; Té, Ds; da Silva, Zj; Ostergaard, L; Laursen, Al; Wejse, C; Krarup, H; Erikstrup, C

2014-10-01

In the case of coinfection with HIV and hepatitis B virus (HBV) and/or hepatitis C virus (HCV), hepatic disease progression is often accelerated, with higher rates of liver cirrhosis and liver-related mortality. We aimed to evaluate the performance of the rapid tests used routinely to detect HBV surface antigen (HBsAg) and anti-HCV among HIV-infected patients in Guinea-Bissau. Blood samples from HIV-infected patients in Guinea-Bissau were stored after testing for HBsAg and anti-HCV with rapid tests. Samples were subsequently re-tested for HBsAg and anti-HCV in Denmark. Two rapid tests were used in Guinea-Bissau: HBsAg Strip Ref 2034 (VEDA.LAB, Alençon, France; sensitivity 62.3%; specificity 99.2%) and HEPA-SCAN (Bhat Bio-Tech, Bangalore, India; sensitivity 57.1%; specificity 99.7%). In the two tests the ability to obtain the correct outcome depended on the antigen and antibody concentrations, respectively. Sex, age, CD4 cell count and antiretroviral therapy status did not differ between false negative and true positive samples in either of the tests. The study is limited by a low number of anti-HCV positive samples. New diagnostic rapid tests should always be evaluated in the setting in which they will be used before implementation. © 2014 British HIV Association.

Comparative Evaluation of Serological and Molecular Methods for the Diagnosis of Scrub Typhus in Indian Settings.

PubMed

Gupta, Nitin; Chaudhry, Rama; Kabra, Sushil K; Lodha, Rakesh; Mirdha, Bijay Ranjan; Das, Bimal Kumar; Dey, Aparajit Ballay; Sreenivas, Vishnubhatla

2017-03-24

The major brunt of scrub typhus is borne by developing countries like India but the disease remains neglected. The rapid progression of this infection to serious complications and associated mortality calls for improved diagnostics. The immunoglobulin (Ig)M immunofluorescence assay (IFA), with all of its limitations remains the gold standard. Of 218 suspected cases from patients enrolled in a 2-year study, 30 cases of scrub typhus were detected by IgM IFA, using a 1:64 dilution. The sensitivities of the IgM enzyme-linked immunosorbent assay, IgM rapid flow assay (RFA), and IgG RFA were found to be 97%, 87%, and 77%, respectively. Their respective specificities were 100%, 100%, and 92%, respectively. The sensitivity and specificity of a nested PCR assay targeting a gene encoding a 56-kDa protein were found to be 50% and 100%, respectively.

Design, optimisation and preliminary validation of a human specific loop-mediated amplification assay for the rapid detection of human DNA at forensic crime scenes.

PubMed

Hird, H J; Brown, M K

2017-11-01

The identification of samples at a crime scene which require forensic DNA typing has been the focus of recent research interest. We propose a simple, but sensitive analysis system which can be deployed at a crime scene to identify crime scene stains as human or non-human. The proposed system uses the isothermal amplification of DNA in a rapid assay format, which returns results in as little as 30min from sampling. The assay system runs on the Genie II device, a proven in-field detection system which could be deployed at a crime scene. The results presented here demonstrate that the system was sufficiently specific and sensitive and was able to detect the presence of human blood, semen and saliva on mock forensic samples. Copyright © 2017. Published by Elsevier B.V.

Rapid and highly sensitive detection of Enterovirus 71 by using nanogold-enhanced electrochemical impedance spectroscopy

NASA Astrophysics Data System (ADS)

Li, Hsing-Yuan; Tseng, Shing-Hua; Cheng, Tsai-Mu; Chu, Hsueh-Liang; Lu, Yu-Ning; Wang, Fang-Yu; Tsai, Li-Yun; Shieh, Juo-Yu; Yang, Jyh-Yuan; Juan, Chien-Chang; Tu, Lung-Chen; Chang, Chia-Ching

2013-07-01

Enterovirus 71 (EV71) infection is an emerging infectious disease causing neurological complications and/or death within two to three days after the development of fever and rash. A low viral titre in clinical specimens makes the detection of EV71 difficult. Conventional approaches for detecting EV71 are time consuming, poorly sensitive, or complicated, and cannot be used effectively for clinical diagnosis. Furthermore, EV71 and Coxsackie virus A16 (CA16) may cross react in conventional assays. Therefore, a rapid, highly sensitive, specific, and user-friendly test is needed. We developed an EV71-specific nanogold-modified working electrode for electrochemical impedance spectroscopy in the detection of EV71. Our results show that EV71 can be distinguished from CA16, Herpes simplex virus, and lysozyme, with the modified nanogold electrode being able to detect EV71 in concentrations as low as 1 copy number/50 μl reaction volume, and the duration between sample preparation and detection being 11 min. This detection platform may have the potential for use in point-of-care diagnostics.

Rapid antibiotic-resistance predictions from genome sequence data for Staphylococcus aureus and Mycobacterium tuberculosis

PubMed Central

Bradley, Phelim; Gordon, N. Claire; Walker, Timothy M.; Dunn, Laura; Heys, Simon; Huang, Bill; Earle, Sarah; Pankhurst, Louise J.; Anson, Luke; de Cesare, Mariateresa; Piazza, Paolo; Votintseva, Antonina A.; Golubchik, Tanya; Wilson, Daniel J.; Wyllie, David H.; Diel, Roland; Niemann, Stefan; Feuerriegel, Silke; Kohl, Thomas A.; Ismail, Nazir; Omar, Shaheed V.; Smith, E. Grace; Buck, David; McVean, Gil; Walker, A. Sarah; Peto, Tim E. A.; Crook, Derrick W.; Iqbal, Zamin

2015-01-01

The rise of antibiotic-resistant bacteria has led to an urgent need for rapid detection of drug resistance in clinical samples, and improvements in global surveillance. Here we show how de Bruijn graph representation of bacterial diversity can be used to identify species and resistance profiles of clinical isolates. We implement this method for Staphylococcus aureus and Mycobacterium tuberculosis in a software package (‘Mykrobe predictor') that takes raw sequence data as input, and generates a clinician-friendly report within 3 minutes on a laptop. For S. aureus, the error rates of our method are comparable to gold-standard phenotypic methods, with sensitivity/specificity of 99.1%/99.6% across 12 antibiotics (using an independent validation set, n=470). For M. tuberculosis, our method predicts resistance with sensitivity/specificity of 82.6%/98.5% (independent validation set, n=1,609); sensitivity is lower here, probably because of limited understanding of the underlying genetic mechanisms. We give evidence that minor alleles improve detection of extremely drug-resistant strains, and demonstrate feasibility of the use of emerging single-molecule nanopore sequencing techniques for these purposes. PMID:26686880

Aptamer-Based Dual-Functional Probe for Rapid and Specific Counting and Imaging of MCF-7 Cells.

PubMed

Yang, Bin; Chen, Beibei; He, Man; Yin, Xiao; Xu, Chi; Hu, Bin

2018-02-06

Development of multimodal detection technologies for accurate diagnosis of cancer at early stages is in great demand. In this work, we report a novel approach using an aptamer-based dual-functional probe for rapid, sensitive, and specific counting and visualization of MCF-7 cells by inductively coupled plasma-mass spectrometry (ICP-MS) and fluorescence imaging. The probe consists of a recognition unit of aptamer to catch cancer cells specifically, a fluorescent dye (FAM) moiety for fluorescence resonance energy transfer (FRET)-based "off-on" fluorescence imaging as well as gold nanoparticles (Au NPs) tag for both ICP-MS quantification and fluorescence quenching. Due to the signal amplification effect and low spectral interference of Au NPs in ICP-MS, an excellent linearity and sensitivity were achieved. Accordingly, a limit of detection of 81 MCF-7 cells and a relative standard deviation of 5.6% (800 cells, n = 7) were obtained. The dynamic linear range was 2 × 10 2 to 1.2 × 10 4 cells, and the recoveries in human whole blood were in the range of 98-110%. Overall, the established method provides quantitative and visualized information on MCF-7 cells with a simple and rapid process and paves the way for a promising strategy for biomedical research and clinical diagnostics.

Development and Evaluation of Reverse Transcription-Loop-Mediated Isothermal Amplification (RT-LAMP) Assay Coupled with a Portable Device for Rapid Diagnosis of Ebola Virus Disease in Guinea

PubMed Central

Kurosaki, Yohei; Magassouba, N’Faly; Oloniniyi, Olamide K.; Cherif, Mahamoud S.; Sakabe, Saori; Takada, Ayato; Hirayama, Kenji; Yasuda, Jiro

2016-01-01

Given the current absence of specific drugs or vaccines for Ebola virus disease (EVD), rapid, sensitive, and reliable diagnostic methods are required to stem the transmission chain of the disease. We have developed a rapid detection assay for Zaire ebolavirus based on reverse transcription-loop-mediated isothermal amplification (RT-LAMP) and coupled with a novel portable isothermal amplification and detection platform. The RT-LAMP assay is based on primer sets that target the untranscribed trailer region or nucleoprotein coding region of the viral RNA. The test could specifically detect viral RNAs of Central and West African Ebola virus strains within 15 minutes with no cross-reactivity to other hemorrhagic fever viruses and arboviruses, which cause febrile disease. The assay was evaluated using a total of 100 clinical specimens (serum, n = 44; oral swab, n = 56) collected from suspected EVD cases in Guinea. The specificity of this diagnostic test was 100% for both primer sets, while the sensitivity was 100% and 97.9% for the trailer and nucleoprotein primer sets, respectively, compared with a reference standard RT-PCR test. These observations suggest that our diagnostic assay is useful for identifying EVD cases, especially in the field or in settings with insufficient infrastructure. PMID:26900929

Point-Counterpoint: A Nucleic Acid Amplification Test for Streptococcus pyogenes Should Replace Antigen Detection and Culture for Detection of Bacterial Pharyngitis.

PubMed

Pritt, Bobbi S; Patel, Robin; Kirn, Thomas J; Thomson, Richard B

2016-10-01

Nucleic acid amplification tests (NAATs) have frequently been the standard diagnostic approach when specific infectious agents are sought in a clinic specimen. They can be applied for specific agents such as S. pyogenes, or commercial multiplex NAATs for detection of a variety of pathogens in gastrointestinal, bloodstream, and respiratory infections may be used. NAATs are both rapid and sensitive. For many years, S. pyogenes testing algorithms used a rapid and specific group A streptococcal antigen test to screen throat specimens, followed, in some clinical settings, by a throat culture for S. pyogenes to increase the sensitivity of its detection. Now S. pyogenes NAATs are being used with increasing frequency. Given their accuracy, rapidity, and ease of use, should they replace antigen detection and culture for the detection of bacterial pharyngitis? Bobbi Pritt and Robin Patel of the Mayo Clinic, where S. pyogenes NAATs have been used for well over a decade with great success, will explain the advantages of this approach, while Richard (Tom) Thomson and Tom Kirn of the NorthShore University HealthSystem will discuss their concerns about this approach to diagnosing bacterial pharyngitis. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Development and Evaluation of Reverse Transcription-Loop-Mediated Isothermal Amplification (RT-LAMP) Assay Coupled with a Portable Device for Rapid Diagnosis of Ebola Virus Disease in Guinea.

PubMed

Kurosaki, Yohei; Magassouba, N'Faly; Oloniniyi, Olamide K; Cherif, Mahamoud S; Sakabe, Saori; Takada, Ayato; Hirayama, Kenji; Yasuda, Jiro

2016-02-01

Given the current absence of specific drugs or vaccines for Ebola virus disease (EVD), rapid, sensitive, and reliable diagnostic methods are required to stem the transmission chain of the disease. We have developed a rapid detection assay for Zaire ebolavirus based on reverse transcription-loop-mediated isothermal amplification (RT-LAMP) and coupled with a novel portable isothermal amplification and detection platform. The RT-LAMP assay is based on primer sets that target the untranscribed trailer region or nucleoprotein coding region of the viral RNA. The test could specifically detect viral RNAs of Central and West African Ebola virus strains within 15 minutes with no cross-reactivity to other hemorrhagic fever viruses and arboviruses, which cause febrile disease. The assay was evaluated using a total of 100 clinical specimens (serum, n = 44; oral swab, n = 56) collected from suspected EVD cases in Guinea. The specificity of this diagnostic test was 100% for both primer sets, while the sensitivity was 100% and 97.9% for the trailer and nucleoprotein primer sets, respectively, compared with a reference standard RT-PCR test. These observations suggest that our diagnostic assay is useful for identifying EVD cases, especially in the field or in settings with insufficient infrastructure.

Endpoint visual detection of three genetically modified rice events by loop-mediated isothermal amplification.

PubMed

Chen, Xiaoyun; Wang, Xiaofu; Jin, Nuo; Zhou, Yu; Huang, Sainan; Miao, Qingmei; Zhu, Qing; Xu, Junfeng

2012-11-07

Genetically modified (GM) rice KMD1, TT51-1, and KF6 are three of the most well known transgenic Bt rice lines in China. A rapid and sensitive molecular assay for risk assessment of GM rice is needed. Polymerase chain reaction (PCR), currently the most common method for detecting genetically modified organisms, requires temperature cycling and relatively complex procedures. Here we developed a visual and rapid loop-mediated isothermal amplification (LAMP) method to amplify three GM rice event-specific junction sequences. Target DNA was amplified and visualized by two indicators (SYBR green or hydroxy naphthol blue [HNB]) within 60 min at an isothermal temperature of 63 °C. Different kinds of plants were selected to ensure the specificity of detection and the results of the non-target samples were negative, indicating that the primer sets for the three GM rice varieties had good levels of specificity. The sensitivity of LAMP, with detection limits at low concentration levels (0.01%−0.005% GM), was 10- to 100-fold greater than that of conventional PCR. Additionally, the LAMP assay coupled with an indicator (SYBR green or HNB) facilitated analysis. These findings revealed that the rapid detection method was suitable as a simple field-based test to determine the status of GM crops.

Establishment of a multiplex real-time RT-PCR assay for rapid identification of H6 subtype avian influenza viruses.

PubMed

Yang, Fan; Wu, Haibo; Liu, Fumin; Lu, Xiangyun; Peng, Xiuming; Wu, Nanping

2018-06-01

The H6 subtype avian influenza viruses (AIVs) possess the capacity for zoonotic transmission from avian species to humans. Establishment of a specific, rapid and sensitive method to screen H6 AIVs is necessary. Based on the conserved domain of the matrix and H6 AIV hemagglutinin genes, two TaqMan minor-groove-binder probes and multiplex real-time RT-PCR primers were designed in this study. The multiplex real-time RT-PCR assay developed in this study had high specificity and repeatability and a detection limit of 30 copies per reaction. This rapid diagnostic method will be useful for clinical detection and surveillance of H6 AIVs in China.

LH750 hematology analyzers to identify malaria and dengue and distinguish them from other febrile illnesses.

PubMed

Sharma, P; Bhargava, M; Sukhachev, D; Datta, S; Wattal, C

2014-02-01

Tropical febrile illnesses such as malaria and dengue are challenging to differentiate clinically. Automated cellular indices from hematology analyzers may afford a preliminary rapid distinction. Blood count and VCS parameters from 114 malaria patients, 105 dengue patients, and 105 febrile controls without dengue or malaria were analyzed. Statistical discriminant functions were generated, and their diagnostic performances were assessed by ROC curve analysis. Three statistical functions were generated: (i) malaria-vs.-controls factor incorporating platelet count and standard deviations of lymphocyte volume and conductivity that identified malaria with 90.4% sensitivity, 88.6% specificity; (ii) dengue-vs.-controls factor incorporating platelet count, lymphocyte percentage and standard deviation of lymphocyte conductivity that identified dengue with 81.0% sensitivity and 77.1% specificity; and (iii) febrile-controls-vs.-malaria/dengue factor incorporating mean corpuscular hemoglobin concentration, neutrophil percentage, mean lymphocyte and monocyte volumes, and standard deviation of monocyte volume that distinguished malaria and dengue from other febrile illnesses with 85.1% sensitivity and 91.4% specificity. Leukocyte abnormalities quantitated by automated analyzers successfully identified malaria and dengue and distinguished them from other fevers. These economic discriminant functions can be rapidly calculated by analyzer software programs to generate electronic flags to trigger-specific testing. They could potentially transform diagnostic approaches to tropical febrile illnesses in cost-constrained settings. © 2013 John Wiley & Sons Ltd.

Clinical Evaluation of the Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test for Rapid Diagnosis of Tuberculosis Lymphadenitis

PubMed Central

Kerleguer, A.; Fabre, M.; Bernatas, J. J.; Gerome, P.; Nicand, E.; Herve, V.; Koeck, J. L.

2004-01-01

This prospective study evaluated the performance of the Amplified Mycobacterium Tuberculosis Direct Test (MTD) for the diagnosis of lymph node tuberculosis in Djibouti, Republic of Djibouti. Of 197 specimens sampled from 153 patients, 123 were from 95 tuberculous patients. The sensitivity and specificity of MTD were 93 and 100%, respectively. The sensitivity of culture was 89%. PMID:15583341

Evaluation of the Alexon-Trend ProSpecT Campylobacter Microplate Assay

PubMed Central

Tolcin, Rita; LaSalvia, Margaret M.; Kirkley, Barbara A.; Vetter, Emily A.; Cockerill, Franklin R.; Procop, Gary W.

2000-01-01

We evaluated stool specimens known to contain or be free of Campylobacter by traditional culture, using the ProSpecT Campylobacter microplate assay (Alexon-Trend, Ramsey, Minn.). This rapid enzyme immunoassay for the detection of Campylobacter-specific antigens demonstrated 96% sensitivity and 99% specificity and is an acceptable alternative method of Campylobacter detection. PMID:11015419

Cost effectiveness of OptiMal® rapid diagnostic test for malaria in remote areas of the Amazon Region, Brazil

PubMed Central

2010-01-01

Background In areas with limited structure in place for microscopy diagnosis, rapid diagnostic tests (RDT) have been demonstrated to be effective. Method The cost-effectiveness of the Optimal® and thick smear microscopy was estimated and compared. Data were collected on remote areas of 12 municipalities in the Brazilian Amazon. Data sources included the National Malaria Control Programme of the Ministry of Health, the National Healthcare System reimbursement table, hospitalization records, primary data collected from the municipalities, and scientific literature. The perspective was that of the Brazilian public health system, the analytical horizon was from the start of fever until the diagnostic results provided to patient and the temporal reference was that of year 2006. The results were expressed in costs per adequately diagnosed cases in 2006 U.S. dollars. Sensitivity analysis was performed considering key model parameters. Results In the case base scenario, considering 92% and 95% sensitivity for thick smear microscopy to Plasmodium falciparum and Plasmodium vivax, respectively, and 100% specificity for both species, thick smear microscopy is more costly and more effective, with an incremental cost estimated at US$549.9 per adequately diagnosed case. In sensitivity analysis, when sensitivity and specificity of microscopy for P. vivax were 0.90 and 0.98, respectively, and when its sensitivity for P. falciparum was 0.83, the RDT was more cost-effective than microscopy. Conclusion Microscopy is more cost-effective than OptiMal® in these remote areas if high accuracy of microscopy is maintained in the field. Decision regarding use of rapid tests for diagnosis of malaria in these areas depends on current microscopy accuracy in the field. PMID:20937094

Development of a reverse transcription loop-mediated isothermal amplification assay for the detection of vesicular stomatitis New Jersey virus: use of rapid molecular assay to differentiate between vesicular disease viruses

USDA-ARS?s Scientific Manuscript database

Vesicular stomatitis (VS) is endemic in Central America and northern regions of South America. Sporadic outbreaks of VS can occur in cattle and pigs where the clinical presentation can be similar to foot-and-mouth disease (FMD). There is therefore a pressing need for rapid, sensitive and specific d...

Evaluation of OXA-48 K-Se T: an immunochromatographic assay for rapid detection of OXA-48-producing Enterobacteriaceae.

PubMed

Fernández, Javier; Fleites, Ana; Rodcio, María Rosario; Vazquez, Fernando

2016-05-01

The OXA-48 K-Se T, a new immunochromatographic assay for rapid detection of OXA-48-producing Enterobacteriaceae, has been evaluated in a Spanish Hospital during a 3-month period. A collection of 100 Enterobacteriaceae including 79 isolates producing OXA-48 has been tested. Sensitivity and specificity of 100% were obtained. Copyright © 2016 Elsevier Inc. All rights reserved.

A novel assay for detecting canine parvovirus using a quartz crystal microbalance biosensor.

PubMed

Kim, Yong Kwan; Lim, Seong-In; Choi, Sarah; Cho, In-Soo; Park, Eun-Hye; An, Dong-Jun

2015-07-01

Rapid and accurate diagnosis is crucial to reduce both the shedding and clinical signs of canine parvovirus (CPV). The quartz crystal microbalance (QCM) is a new tool for measuring frequency changes associated with antigen-antibody interactions. In this study, the QCM biosensor and ProLinker™ B were used to rapidly diagnosis CPV infection. ProLinker™ B enables antibodies to be attached to a gold-coated quartz surface in a regular pattern and in the correct orientation for antigen binding. Receiver operating characteristics (ROC) curves were used to set a cut-off value using reference CPVs (two groups: one CPV-positive and one CPV-negative). The ROC curves overlapped and the point of intersection was used as the cut-off value. A QCM biosensor with a cut-off value of -205 Hz showed 95.4% (104/109) sensitivity and 98.0% (149/152) specificity when used to test 261 field fecal samples compared to PCR. In conclusion, the QCM biosensor described herein is eminently suitable for the rapid diagnosis of CPV infection with high sensitivity and specificity. Therefore, it is a promising analytical tool that will be useful for clinical diagnosis, which requires rapid and reliable analyses. Copyright © 2015 Elsevier B.V. All rights reserved.

Comparative Evaluation of Tubex TF (Inhibition Magnetic Binding Immunoassay) for Typhoid Fever in Endemic Area.

PubMed

Khanna, Ashish; Khanna, Menka; Gill, Karamjit Singh

2015-11-01

Typhoid fever remains a significant health problem in endemic countries like India. Various serological tests for the diagnosis of typhoid fever are available commercially. We assessed the usefulness of rapid test based on magnetic particle separation to detect Immunoglobulin against Salmonella typhi O9 lipopolysaccharide. Aim of this study was to compare the sensitivity and specificity of widal test, typhidot and tubex TF test for the diagnosis of typhoid fever in an endemic country like India. Serum samples collected from 50 patients of typhoid fever, 50 patients of non typhoid fever and 100 normal healthy individuals residing in Amritsar were subjected to widal test, typhidot test and tubex TF test as per manufacturer's instructions. Data collected was assessed to find sensitivity and specificity of these tests in an endemic area. Significant widal test results were found positive in 68% of patients of typhoid fever and only 4% of non typhoid fever patients. Typhidot (IgM or IgG) was positive in 72% of typhoid fever patients and 10% and 6% in non typhoid fever and normal healthy individuals respectively. Tubex TF showed higher sensitivity of 76% and specificity of 96-99% which was higher than typhidot and comparable to widal test. This was the first evaluation of rapid tubex TF test in northern India. In countries which can afford high cost of test, tubex TF should be recommended for the diagnosis in acute stage of the disease in clinical setting. However, there is urgent need for a highly specific and sensitive test for the diagnosis of typhoid fever in clinical settings in endemic areas.

Oral rapid test: an alternative to traditional HIV screening in Chile

PubMed Central

Irarrazábal, Lisette Paola; Ferrer, Lilian; Cianelli, Rosina; Lara, Loreto; Reed, Reiley; Levy, Judith; Pérez, Carlos

2016-01-01

Objective To compare the sensitivity and specificity of an Oral Rapid Test (ORT) to that of the Enzyme-Linked Immunosorbent Assay (ELISA) for HIV testing in Santiago, Chile; to track the number of study participants returning for ELISA testing results; and to analyze the participants’ perceptions of the ORT compared to the ELISA. Methods A total of 497 people were recruited in Santiago, Chile: 153 had previously tested positive for HIV, and 344 were of unknown status. Participants were tested for HIV using both the ELISA and the ORT to examine and compare specificity and sensitivity. Qualitative data were collected from 22 participants to compare perceptions of the testing experience with ORT versus ELISA. Results The ELISA reported 184 (37%) of the 497 participants as being “positive” for HIV antibodies; the ORT showed 181 (36.4%) as being “reactive” for HIV. The ORT showed a sensitivity of 98.4% (95.7%–99.9%, 95% Confidence Interval) and specificity of 100%. The Kappa test produced K = 0.983 (P < 0.0001). Of the 344 participants whose HIV status was unknown at the start of the study, 55 failed to return for their ELISA results. Participants positively perceived ORT as having reduced both waiting time and anxiety over obtaining their test results. ORT oral swabbing appeared more practical and less invasive than drawing blood for the ELISA. Conclusions The ORT and ELISA were statistically equal in specificity and sensitivity. ORT provides quicker results, potentially ensuring that more people receive them, and does not require handling of or exposure to potentially hazardous blood products. PMID:23939368

Rapid antigen detection tests for malaria diagnosis in severely ill Papua New Guinean children: a comparative study using Bayesian latent class models.

PubMed

Manning, Laurens; Laman, Moses; Rosanas-Urgell, Anna; Turlach, Berwin; Aipit, Susan; Bona, Cathy; Warrell, Jonathan; Siba, Peter; Mueller, Ivo; Davis, Timothy M E

2012-01-01

Although rapid diagnostic tests (RDTs) have practical advantages over light microscopy (LM) and good sensitivity in severe falciparum malaria in Africa, their utility where severe non-falciparum malaria occurs is unknown. LM, RDTs and polymerase chain reaction (PCR)-based methods have limitations, and thus conventional comparative malaria diagnostic studies employ imperfect gold standards. We assessed whether, using Bayesian latent class models (LCMs) which do not require a reference method, RDTs could safely direct initial anti-infective therapy in severe ill children from an area of hyperendemic transmission of both Plasmodium falciparum and P. vivax. We studied 797 Papua New Guinean children hospitalized with well-characterized severe illness for whom LM, RDT and nested PCR (nPCR) results were available. For any severe malaria, the estimated prevalence was 47.5% with RDTs exhibiting similar sensitivity and negative predictive value (NPV) to nPCR (≥96.0%). LM was the least sensitive test (87.4%) and had the lowest NPV (89.7%), but had the highest specificity (99.1%) and positive predictive value (98.9%). For severe falciparum malaria (prevalence 42.9%), the findings were similar. For non-falciparum severe malaria (prevalence 6.9%), no test had the WHO-recommended sensitivity and specificity of >95% and >90%, respectively. RDTs were the least sensitive (69.6%) and had the lowest NPV (96.7%). RDTs appear a valuable point-of-care test that is at least equivalent to LM in diagnosing severe falciparum malaria in this epidemiologic situation. None of the tests had the required sensitivity/specificity for severe non-falciparum malaria but the number of false-negative RDTs in this group was small.

Diagnostic Sensitivity and Specificity of Dark Adaptometry for Detection of Age-Related Macular Degeneration

PubMed Central

Jackson, Gregory R.; Scott, Ingrid U.; Kim, Ivana K.; Quillen, David A.; Iannaccone, Alessandro; Edwards, John G.

2014-01-01

Purpose. Difficulty with night vision is a common complaint of patients with age-related macular degeneration (AMD). Consistent with this complaint, dark adaptation (DA) is substantially impaired in these patients. Because of the severity of the deficit, measurement of DA has been suggested as a means for the diagnosis of AMD. Previous methods for measurement of DA were time intensive (>30 minutes), which made them unsuitable for clinical use. This study evaluated a rapid DA test (≤6.5 minutes) for the detection of AMD. Methods. Dark adaptation was measured by using the AdaptDx dark adaptometer in two groups: subjects with normal retinal health and subjects with AMD. Subjects were assigned to their group by clinical examination and grading of fundus photographs. Subjects were classified as having DA consistent with normal retinal health (rod intercept ≤ 6.5 minutes) or having dark adaptation consistent with AMD (rod intercept > 6.5 minutes). Results. The eligible sample for analysis included 21 normal adults and 127 AMD patients. The rapid test was found to have a diagnostic sensitivity of 90.6% (P < 0.001) and specificity of 90.5% (P < 0.027). Thus, abnormal DA was detected in 115 of 127 AMD patients, and normal DA was found in 19 of 21 normal adults. Conclusions. The high diagnostic sensitivity and specificity compared favorably to long-duration research methods for the measurement of DA, and slit lamp biomicroscopy performed by a retina specialist. These results suggest that a rapid DA test is useful for the detection of AMD. PMID:24550363

Population screening for coeliac disease in primary care by district nurses using a rapid antibody test: diagnostic accuracy and feasibility study

PubMed Central

2007-01-01

Objective To evaluate the feasibility and diagnostic accuracy of screening for coeliac disease by rapid detection of IgA antibodies to tissue transglutaminase performed in primary care. Design District nurses screened 6 year old children using rapid antibody testing of finger prick blood. They also collected capillary blood samples for laboratory determination of IgA and IgG antibodies to endomysium and IgA antibodies to tissue transglutaminase. Children with positive rapid test results were directly sent for biopsy of the small intestine. Setting Primary care in Jász-Nagykun-Szolnok county, Hungary. Participants 2690 children (77% of 6 year olds living in the county) and 120 nurses. Main outcome measures Positivity for antibodies to endomysium or transglutaminase in the laboratory and coeliac disease confirmed at biopsy. Results 37 children (1.4%, 95% confidence interval 0.9% to 1.8%) had biopsy confirmed coeliac disease. Only five of these children had been diagnosed clinically before screening. Rapid testing had a 78.1% sensitivity (70.0% to 89.3%) and 100% specificity (88.4% to 100%) for a final diagnosis of coeliac disease by biopsy. Sensitivity was 65.1% (50.2% to 77.6%) and specificity was 100% (99.8% to 100%) compared with combined results of IgA and IgG laboratory tests. Trained laboratory workers detected 30 of the 31 newly diagnosed IgA competent patients with the rapid test kit used blindly. Median time to biopsy after a positive rapid test result was significantly shorter (20 days, range 4-148) than after a positive laboratory result (142 days, 70-256; P<0.001). Children with coeliac disease detected at screening were smaller and had worse health status than their peers but they improved on a gluten-free diet. Conclusions A simple rapid antibody test enabled primary care nurses to detect patients with coeliac disease in the community who were not picked up in clinical care. Extra training is needed to improve sensitivity. PMID:18063612

Evaluation of a rapid immunochromatographic test for the detection of low burden Dirofilaria immitis (heartworm) in dogs and cats.

PubMed

Genchi, Marco; Mangia, Carlo; Ferrari, Nicola; Loukeri, Sofia

2018-01-01

The performance of a rapid immunochromatographic test for the detection of Dirofilaria immitis antigens (Speed Diro™; BVT-Virbac, France) was assessed in 49 experimentally infected dogs and in 244 naturally infected animals; 142 dogs and 102 cats. In experimentally infected dogs, Speed Diro™ showed a sensitivity of 90.9% in dogs infected with one adult female worm and 100% in dogs infected with more than one female worm. Specificity was 100%. For naturally infected dogs, the Knott test and PetChek ® HTWM PF served as reference methods for microfilaremia and antigenemia, respectively. All microfilaemic dogs (55/142) were positive with Speed Diro™. Importantly, none of the 21 dogs infected with D. repens were positive. The results of Speed Diro™ for the detection of antigenemia were compared with two in-house tests, SNAP ® HTWM and Witness ® Dirofilaria, and all three tests were 100% specific and sensitive in comparison to PetChek ® HTWM PF. For the evaluation of feline samples, 102 cats were examined by echocardiography. Sera from 87 heartworm-infected cats were tested by Speed Diro™ and SNAP ® HTWM. The results of Speed Diro™ were equivalent to SNAP ® HTWM, with a sensitivity of 98.9% and a specificity of 100%.

Nested-PCR and a new ELISA-based NovaLisa test kit for malaria diagnosis in an endemic area of Thailand.

PubMed

Thongdee, Pimwan; Chaijaroenkul, Wanna; Kuesap, Jiraporn; Na-Bangchang, Kesara

2014-08-01

Microscopy is considered as the gold standard for malaria diagnosis although its wide application is limited by the requirement of highly experienced microscopists. PCR and serological tests provide efficient diagnostic performance and have been applied for malaria diagnosis and research. The aim of this study was to investigate the diagnostic performance of nested PCR and a recently developed an ELISA-based new rapid diagnosis test (RDT), NovaLisa test kit, for diagnosis of malaria infection, using microscopic method as the gold standard. The performance of nested-PCR as a malaria diagnostic tool is excellent with respect to its high accuracy, sensitivity, specificity, and ability to discriminate Plasmodium species. The sensitivity and specificity of nested-PCR compared with the microscopic method for detection of Plasmodium falciparum, Plasmodium vivax, and P. falciparum/P. vivax mixed infection were 71.4 vs 100%, 100 vs 98.7%, and 100 vs 95.0%, respectively. The sensitivity and specificity of the ELISA-based NovaLisa test kit compared with the microscopic method for detection of Plasmodium genus were 89.0 vs 91.6%, respectively. NovaLisa test kit provided comparable diagnostic performance. Its relatively low cost, simplicity, and rapidity enables large scale field application.

Specific detection of Angiostrongylus cantonensis in the snail Achatina fulica using a loop-mediated isothermal amplification (LAMP) assay.

PubMed

Liu, Chun-Yan; Song, Hui-Qun; Zhang, Ren-Li; Chen, Mu-Xin; Xu, Min-Jun; Ai, Lin; Chen, Xiao-Guang; Zhan, Xi-Mei; Liang, Shao-Hui; Yuan, Zi-Guo; Lin, Rui-Qing; Zhu, Xing-Quan

2011-08-01

Angiostrongylus cantonensis, a rat lungworm, can cause eosinophilic meningitis and angiostrongyliasis in humans following ingestion of contaminated foods or intermediate/paratenic hosts with infective larvae. The snail Achatina fulica is one of the important intermediate hosts of A. cantonensis and is commonly eaten by humans in some countries. In the present study, we developed a loop-mediated isothermal amplification (LAMP) method for the specific detection of A. cantonensis in Ac. fulica. Primers for LAMP were designed based on the first internal transcribed spacer (ITS-1) of nuclear ribosomal DNA (rDNA) of A. cantonensis. Specificity tests showed that only the products of A. cantonensis were detected when DNA samples of A. cantonensis and the heterologous control samples Anisakis simplex s.s, Trichuris trichiura, Toxocara canis, Trichinella spiralis and Ascaris lumbricoides were amplified by LAMP. Sensitivity evaluation indicated that the LAMP assay is 10 times more sensitive than the conventional polymerase chain reaction (PCR) assay. The established LAMP assay is rapid, inexpensive and easy to be performed. It can be used in clinical applications for rapid and sensitive detection of A. cantonensis in snails, which has implications for the effective control of angiostrongyliasis. Copyright © 2011 Elsevier Ltd. All rights reserved.

A Multiplexed Diagnostic Platform for Point-of-Care Pathogen Detection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Regan, J F; Letant, S E; Adams, K L

2008-02-04

We developed an automated point-of-care diagnostic instrument that is capable of analyzing nasal swab samples for the presence of respiratory diseases. This robust instrument, called FluIDx, performs autonomous multiplexed RT-PCR reactions that are analyzed by microsphere xMAP technology. We evaluated the performance of FluIDx, in comparison rapid tests specific for influenza and respiratory syncytial virus, in a clinical study performed at the UC Davis Medical Center. The clinical study included samples positive for RSV (n = 71), influenza A (n = 16), influenza B (n = 4), adenovirus (n = 5), parainfluenza virus (n = 2), and 44 negative samples,more » according to a composite reference method. FluIDx and the rapid tests detected 85.9% and 62.0% of the RSV positive samples, respectively. Similar sensitivities were recorded for the influenza B samples; whereas the influenza A samples were poorly detected, likely due to the utilization of an influenza A signature that did not accurately match currently circulating influenza A strains. Data for all pathogens were compiled and indicate that FluIDx is more sensitive than the rapid tests, detecting 74.2% (95% C.I. of 64.7-81.9%) of the positive samples in comparison to 53.6% (95% C.I. of 43.7-63.2%) for the rapid tests. The higher sensitivity of FluIDx was partially offset by a lower specificity, 77.3% versus 100.0%. Overall, these data suggest automated flow-through PCR-based instruments that perform multiplexed assays can successfully screen clinical samples for infectious diseases.« less

Use of Tethered Enzymes as a Platform Technology for Rapid Analyte Detection

PubMed Central

Cohen, Roy; Lata, James P.; Lee, Yurim; Hernández, Jean C. Cruz; Nishimura, Nozomi; Schaffer, Chris B.; Mukai, Chinatsu; Nelson, Jacquelyn L.; Brangman, Sharon A.; Agrawal, Yash; Travis, Alexander J.

2015-01-01

Background Rapid diagnosis for time-sensitive illnesses such as stroke, cardiac arrest, and septic shock is essential for successful treatment. Much attention has therefore focused on new strategies for rapid and objective diagnosis, such as Point-of-Care Tests (PoCT) for blood biomarkers. Here we use a biomimicry-based approach to demonstrate a new diagnostic platform, based on enzymes tethered to nanoparticles (NPs). As proof of principle, we use oriented immobilization of pyruvate kinase (PK) and luciferase (Luc) on silica NPs to achieve rapid and sensitive detection of neuron-specific enolase (NSE), a clinically relevant biomarker for multiple diseases ranging from acute brain injuries to lung cancer. We hypothesize that an approach capitalizing on the speed and catalytic nature of enzymatic reactions would enable fast and sensitive biomarker detection, suitable for PoCT devices. Methods and findings We performed in-vitro, animal model, and human subject studies. First, the efficiency of coupled enzyme activities when tethered to NPs versus when in solution was tested, demonstrating a highly sensitive and rapid detection of physiological and pathological concentrations of NSE. Next, in rat stroke models the enzyme-based assay was able in minutes to show a statistically significant increase in NSE levels in samples taken 1 hour before and 0, 1, 3 and 6 hours after occlusion of the distal middle cerebral artery. Finally, using the tethered enzyme assay for detection of NSE in samples from 20 geriatric human patients, we show that our data match well (r = 0.815) with the current gold standard for biomarker detection, ELISA—with a major difference being that we achieve detection in 10 minutes as opposed to the several hours required for traditional ELISA. Conclusions Oriented enzyme immobilization conferred more efficient coupled activity, and thus higher assay sensitivity, than non-tethered enzymes. Together, our findings provide proof of concept for using oriented immobilization of active enzymes on NPs as the basis for a highly rapid and sensitive biomarker detection platform. This addresses a key challenge in developing a PoCT platform for time sensitive and difficult to diagnose pathologies. PMID:26605916

A novel one-step Helicobacter pylori saliva antigen test.

PubMed

Yang, Bi-Ling; Yeh, Chun; Kwong, Wei-Gang; Lee, Shou-Dong

2015-02-01

A rapid, reliable, and sufficiently accurate test for diagnosing Helicobacter pylori infection is required for screening dyspeptic patients before a referral for endoscopy. The purpose of this article is two-fold: first, to evaluate the accuracy of a one-step H. pylori saliva antigen (HPS) test; and second, to compare noninvasive and invasive H. pylori tests in Taiwanese population. A total of 104 consecutive dyspeptic patients admitted for gastroenterology into the outpatient department underwent a one-step HPS test, rapid urease test, histology, and (13)C-urea breath test (13)C-UBT (proto C-13 urea kit). The accuracy of the HPS test was compared with a gold standard defined by at least two positive H. pylori test results from three H. pylori tests (histology, rapid urease test, and (13)C-UBT). The 104 patients eligible for analysis (mean age: 58 years, range 22-87 years), 21 (20%) were gold standard positive. Among them, the positive of the one-step H. pylori saliva Ag test, rapid urease test, (13)C-UBT, histology were (52; 50%), (17; 16%), (27; 25%) and (22; 21%) respectively. The sensitivity and specificity of the HPS tests, rapid urease test, (13)C-UBTs, and histology were 71.43% and 55.42%, 76.19% and 98.80%, 100% and 92.77%, and 85.71% and 95.18%, respectively, relative to the gold standard. The one-step HPS test exhibited a sensitivity of 71.43%, nearly equivalent to that of the rapid urea test. The one-step HPS test exhibited a high sensitivity and low specificity compared with the other tests, indicating that it is not sufficiently accurate for use in a clinical setting for diagnosing H. pylori infection. However, the test is simple to use (requiring only a saliva sample), inexpensive, and noninvasive in its application, and thus appealing for use in population-based prevalence surveys of the epidemiology of H. pylori infection. Copyright © 2014. Published by Elsevier Taiwan.

Synthesis of molecularly imprinted dye-silica nanocomposites with high selectivity and sensitivity: Fluorescent imprinted sensor for rapid and efficient detection of τ-fluvalinate in vodka.

PubMed

Wang, Yunyun; Wang, Jixiang; Cheng, Rujia; Sun, Lin; Dai, Xiaohui; Yan, Yongsheng

2018-04-01

An imprinted fluorescent sensor was fabricated based on SiO 2 nanoparticles encapsulated with a molecularly imprinted polymer containing allyl fluorescein. High fluorine cypermethirin as template molecules, methyl methacrylate as functional monomer, and allyl fluorescein as optical materials synthesized a core-shell fluorescent molecular imprinted sensor, which showed a high and rapid sensitivity and selectivity for the detection of τ-fluvalinate. The sensor presented appreciable sensitivity with a limit of 13.251 nM, rapid detection that reached to equilibrium within 3 min, great linear relationship in the relevant concentration range from 0 to 150 nM, and excellent selectivity over structural analogues. In addition, the fluorescent sensor demonstrated desirable regeneration ability (eight cycling operations). The molecularly imprinted polymers ensured specificity, while the fluorescent dyes provided the stabile sensitivity. Finally, an effective application of the sensor was implemented by the detection of τ-fluvalinate in real samples from vodka. The molecularly imprinted fluorescent sensor showed a promising potential in environmental monitoring and food safety. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Uncomplicated malaria in children: The place of rapid diagnostic test.

PubMed

Elechi, Hassan Abdullahi; Rabasa, Adamu Ibrahim; Bashir, Muhammad Faruk; Gofama, Mustapha Modu; Ibrahim, Halima Abubakar; Askira, Umoru Muhammed

2015-01-01

Malaria has remained a major cause of morbidity and mortality among the under-five children in Nigeria. Prompt and accurate diagnosis of malaria is necessary in controlling this high burden and preventing unnecessary use of anti-malarial drugs. Malaria rapid diagnostic test (MRDT) offers the hope of achieving this goal. However, the performance of these kits among the most vulnerable age group to malaria is inadequate. In this cross-sectional study, 433 out-patients, aged <5 years with fever or history of fever were enrolled. Each candidate was tested for malaria parasitaemia using ACON; malaria pf. Thick and thin films were also prepared from the same finger prick blood for each candidate. Malaria rapid diagnostic test had sensitivity of 8.3%, specificity of 100%, positive predictive value (PPV) of 100% and negative predictive value (NPV) of 74%. The sensitivity of MRDT increased with increasing age. This effect of age on sensitivity was statistically significant (P = 0.007). Similarly parasite density had significant effect on the sensitivity of MRDT (P = <0.001). Histidine-rich protein-2 based MRDT is not a reliable mean of diagnosing malaria in the under-five age children with acute uncomplicated malaria.

Loop-mediated isothermal amplification (LAMP) method for detection of genetically modified maize T25.

PubMed

Xu, Junyi; Zheng, Qiuyue; Yu, Ling; Liu, Ran; Zhao, Xin; Wang, Gang; Wang, Qinghua; Cao, Jijuan

2013-11-01

The loop-mediated isothermal amplification (LAMP) assay indicates a potential and valuable means for genetically modified organism (GMO) detection especially for its rapidity, simplicity, and low cost. We developed and evaluated the specificity and sensitivity of the LAMP method for rapid detection of the genetically modified (GM) maize T25. A set of six specific primers was successfully designed to recognize six distinct sequences on the target gene, including a pair of inner primers, a pair of outer primers, and a pair of loop primers. The optimum reaction temperature and time were verified to be 65°C and 45 min, respectively. The detection limit of this LAMP assay was 5 g kg(-1) GMO component. Comparative experiments showed that the LAMP assay was a simple, rapid, accurate, and specific method for detecting the GM maize T25.

Loop-mediated isothermal amplification (LAMP) method for detection of genetically modified maize T25

PubMed Central

Xu, Junyi; Zheng, Qiuyue; Yu, Ling; Liu, Ran; Zhao, Xin; Wang, Gang; Wang, Qinghua; Cao, Jijuan

2013-01-01

The loop-mediated isothermal amplification (LAMP) assay indicates a potential and valuable means for genetically modified organism (GMO) detection especially for its rapidity, simplicity, and low cost. We developed and evaluated the specificity and sensitivity of the LAMP method for rapid detection of the genetically modified (GM) maize T25. A set of six specific primers was successfully designed to recognize six distinct sequences on the target gene, including a pair of inner primers, a pair of outer primers, and a pair of loop primers. The optimum reaction temperature and time were verified to be 65°C and 45 min, respectively. The detection limit of this LAMP assay was 5 g kg−1 GMO component. Comparative experiments showed that the LAMP assay was a simple, rapid, accurate, and specific method for detecting the GM maize T25. PMID:24804053

A vertically aligned carbon nanotube-based impedance sensing biosensor for rapid and high sensitive detection of cancer cells.

PubMed

Abdolahad, Mohammad; Taghinejad, Mohammad; Taghinejad, Hossein; Janmaleki, Mohsen; Mohajerzadeh, Shams

2012-03-21

A novel vertically aligned carbon nanotube based electrical cell impedance sensing biosensor (CNT-ECIS) was demonstrated for the first time as a more rapid, sensitive and specific device for the detection of cancer cells. This biosensor is based on the fast entrapment of cancer cells on vertically aligned carbon nanotube arrays and leads to mechanical and electrical interactions between CNT tips and entrapped cell membranes, changing the impedance of the biosensor. CNT-ECIS was fabricated through a photolithography process on Ni/SiO(2)/Si layers. Carbon nanotube arrays have been grown on 9 nm thick patterned Ni microelectrodes by DC-PECVD. SW48 colon cancer cells were passed over the surface of CNT covered electrodes to be specifically entrapped on elastic nanotube beams. CNT arrays act as both adhesive and conductive agents and impedance changes occurred as fast as 30 s (for whole entrapment and signaling processes). CNT-ECIS detected the cancer cells with the concentration as low as 4000 cells cm(-2) on its surface and a sensitivity of 1.7 × 10(-3)Ω cm(2). Time and cell efficiency factor (TEF and CEF) parameters were defined which describe the sensor's rapidness and resolution, respectively. TEF and CEF of CNT-ECIS were much higher than other cell based electrical biosensors which are compared in this paper.

Establishment of reverse transcription loop-mediated isothermal amplification for rapid detection and differentiation of canine distemper virus infected and vaccinated animals.

PubMed

Liu, Da-Fei; Liu, Chun-Guo; Tian, Jin; Jiang, Yi-Tong; Zhang, Xiao-Zhan; Chai, Hong-Liang; Yang, Tian-Kuo; Yin, Xiu-Chen; Zhang, Hong-Ying; Liu, Ming; Hua, Yu-Ping; Qu, Lian-Dong

2015-06-01

Although widespread vaccination against canine distemper virus (CDV) has been conducted for many decades, several canine distemper outbreaks in vaccinated animals have been reported frequently. In order to detect and differentiate the wild-type and vaccine strains of the CDV from the vaccinated animals, a novel reverse transcription loop-mediated isothermal amplification (RT-LAMP) method was developed. A set of four primers-two internal and two external-were designed to target the H gene for the specific detection of wild-type CDV variants. The CDV-H RT-LAMP assay rapidly amplified the target gene, within 60 min, using a water bath held at a constant temperature of 65°C. The assay was 100-fold more sensitive than conventional RT-PCR, with a detection limit of 10(-1)TCID50ml(-1). The system showed a preference for wild-type CDV, and exhibited less sensitivity to canine parvovirus, canine adenovirus type 1 and type 2, canine coronavirus, and canine parainfluenza virus. The assay was validated using 102 clinical samples obtained from vaccinated dog farms, and the results were comparable to a multiplex nested RT-PCR assay. The specific CDV-H RT-LAMP assay provides a simple, rapid, and sensitive tool for the detection of canines infected with wild-type CDV from canines vaccinated with attenuated vaccine. Copyright © 2015 Elsevier B.V. All rights reserved.

Multicountry Prospective Clinical Evaluation of Two Enzyme-Linked Immunosorbent Assays and Two Rapid Diagnostic Tests for Diagnosing Dengue Fever

PubMed Central

Dauner, Allison L.; Valks, Andrea; Forshey, Brett M.; Long, Kanya C.; Thaisomboonsuk, Butsaya; Sierra, Gloria; Picos, Victor; Talmage, Sara; Morrison, Amy C.; Halsey, Eric S.; Comach, Guillermo; Yasuda, Chadwick; Loeffelholz, Michael; Jarman, Richard G.; Fernandez, Stefan; An, Ung Sam; Kochel, Tadeusz J.; Jasper, Louis E.; Wu, Shuenn-Jue L.

2015-01-01

We evaluated four dengue diagnostic devices from Alere, including the SD Bioline Dengue Duo (nonstructural [NS] 1 Ag and IgG/IgM), the Panbio Dengue Duo Cassette (IgM/IgG) rapid diagnostic tests (RDTs), and the Panbio dengue IgM and IgG capture enzyme-linked immunosorbent assays (ELISAs) in a prospective, controlled, multicenter study in Peru, Venezuela, Cambodia, and the United States, using samples from 1,021 febrile individuals. Archived, well-characterized samples from an additional 135 febrile individuals from Thailand were also used. Reference testing was performed on all samples using an algorithm involving virus isolation, in-house IgM and IgG capture ELISAs, and plaque reduction neutralization tests (PRNT) to determine the infection status of the individual. The primary endpoints were the clinical sensitivities and specificities of these devices. The SD Bioline Dengue Duo had an overall sensitivity of 87.3% (95% confidence interval [CI], 84.1 to 90.2%) and specificity of 86.8% (95% CI, 83.9 to 89.3%) during the first 14 days post-symptom onset (p.s.o.). The Panbio Dengue Duo Cassette demonstrated a sensitivity of 92.1% (87.8 to 95.2%) and specificity of 62.2% (54.5 to 69.5%) during days 4 to 14 p.s.o. The Panbio IgM capture ELISA had a sensitivity of 87.6% (82.7 to 91.4%) and specificity of 88.1% (82.2 to 92.6%) during days 4 to 14 p.s.o. Finally, the Panbio IgG capture ELISA had a sensitivity of 69.6% (62.1 to 76.4%) and a specificity of 88.4% (82.6 to 92.8%) during days 4 to 14 p.s.o. for identification of secondary dengue infections. This multicountry prospective study resulted in reliable real-world performance data that will facilitate data-driven laboratory test choices for managing patient care during dengue outbreaks. PMID:25588659

Rapid detection of all known ebolavirus species by reverse transcription-loop-mediated isothermal amplification (RT-LAMP).

PubMed

Oloniniyi, Olamide K; Kurosaki, Yohei; Miyamoto, Hiroko; Takada, Ayato; Yasuda, Jiro

2017-08-01

Ebola virus disease (EVD), a highly virulent infectious disease caused by ebolaviruses, has a fatality rate of 25-90%. Without a licensed chemotherapeutic agent or vaccine for the treatment and prevention of EVD, control of outbreaks requires accurate and rapid diagnosis of cases. In this study, five sets of six oligonucleotide primers targeting the nucleoprotein gene were designed for specific identification of each of the five ebolavirus species using reverse transcription-loop mediated isothermal amplification (RT-LAMP) assay. The detection limits of the ebolavirus species-specific primer sets were evaluated using in vitro transcribed RNAs. The detection limit of species-specific RT-LAMP assays for Zaire ebolavirus, Sudan ebolavirus, Taï Forest ebolavirus, and Bundibugyo ebolavirus was 256 copies/reaction, while the detection limit for Reston ebolavirus was 64 copies/reaction, and the detection time for each of the RT-LAMP assays was 13.3±3.0, 19.8±4.6, 14.3±0.6, 16.1±4.7, and 19.8±2.4min (mean±SD), respectively. The sensitivity of the species-specific RT-LAMP assays were similar to that of the established RT-PCR and quantitative RT-PCR assays for diagnosis of EVD and are suitable for field or point-of-care diagnosis. The RT-LAMP assays were specific for the detection of the respective species of ebolavirus with no cross reaction with other species of ebolavirus and other viral hemorrhagic fever viruses such as Marburg virus, Lassa fever virus, and Dengue virus. The species-specific RT-LAMP assays developed in this study are rapid, sensitive, and specific and could be useful in case of an EVD outbreak. Copyright © 2017 Elsevier B.V. All rights reserved.

Evaluation of the Kodak Surecell Chlamydia test for the laboratory diagnosis of adult inclusion conjunctivitis.

PubMed

Tantisira, J G; Kowalski, R P; Gordon, Y J

1995-07-01

The Kodak Surecell Chlamydia test, a rapid enzyme immunoassay, has been reported to be highly sensitive (93%) and specific (96%) for detecting chlamydial lipopolysaccharide antigen in conjunctival specimens from infants, but has not been evaluated previously in adult conjunctival specimens. This study was designed to determine the efficacy of the Kodak Surecell Chlamydia test for the laboratory diagnosis of adult inclusion conjunctivitis. Twenty Chlamydia culture-positive conjunctival specimens from adults (true-positives) and 20 true-negative specimens were tested with the Kodak Surecell Chlamydia test. The Kodak Surecell Chlamydia test was 40% (8/20) sensitive, 100% (20/20) specific, and 70% (28/40) efficient. This study indicates that the Kodak Surecell Chlamydia test, though highly specific, is less sensitive in its ability to diagnose chlamydial conjunctivitis in adults than has been reported previously in infants.

Highly sensitive and rapid bacteria detection using molecular beacon-Au nanoparticles hybrid nanoprobes.

PubMed

Cao, Jing; Feng, Chao; Liu, Yan; Wang, Shouyu; Liu, Fei

2014-07-15

Since many diseases are caused by pathogenic bacterial infections, accurate and rapid detection of pathogenic bacteria is in urgent need to timely apply appropriate treatments and to reduce economic costs. To end this, we designed molecular beacon-Au nanoparticle hybrid nanoprobes to improve the bacterial detection efficiency and sensitivity. Here, we show that the designed molecular beacon modified Au nanoparticles could specifically recognize synthetic DNAs targets and can readily detect targets in clinical samples. Moreover, the hybrid nanoprobes can recognize Escherichia coli within an hour at a concentration of 10(2) cfu/ml, which is 1000-folds sensitive than using molecular beacon directly. Our results show that the molecular beacon-Au nanoparticle hybrid nanoprobes have great potential in medical and biological applications. Copyright © 2014 Elsevier B.V. All rights reserved.

Utility of feline coronavirus antibody tests.

PubMed

Addie, Diane D; le Poder, Sophie; Burr, Paul; Decaro, Nicola; Graham, Elizabeth; Hofmann-Lehmann, Regina; Jarrett, Oswald; McDonald, Michael; Meli, Marina L

2015-02-01

Eight different tests for antibodies to feline coronavirus (FCoV) were evaluated for attributes that are important in situations in veterinary practice. We compared four indirect immunofluorescent antibody tests (IFAT), one enzyme-linked immunosorbent assay (ELISA) (FCoV Immunocomb; Biogal) and three rapid immunochromatographic (RIM) tests against a panel of samples designated by consensus as positive or negative. Specificity was 100% for all but the two IFATs based on transmissible gastroenteritis virus (TGEV), at 83.3% and 97.5%. The IFAT and ELISA tests were best for obtaining an antibody titre and for working in the presence of virus. The RIM tests were the best for obtaining a result quickly (10-15 mins); of these, the Speed F-Corona was the most sensitive, at 92.4%, followed by FASTest feline infectious peritonitis (FIP; 84.6%) and Anigen Rapid FCoV antibody test (64.1%). Sensitivity was 100% for the ELISA, one FCoV IFAT and one TGEV IFAT; and 98.2% for a second TGEV IFA and 96.1% for a second FCoV IFAT. All tests worked with effusions, even when only blood products were stipulated in the instruction manual. The ELISA and Anigen RIM tests were best for small quantities of sample. The most appropriate FCoV antibody test to use depends on the reason for testing: in excluding a diagnosis of FIP, sensitivity, specificity, small sample quantity, rapidity and ability to work in the presence of virus all matter. For FCoV screening, speed and sensitivity are important, and for FCoV elimination antibody titre is essential. © ISFM and AAFP 2014.

Rapid antigen tests for dengue virus serotypes and Zika virus in patient serum.

PubMed

Bosch, Irene; de Puig, Helena; Hiley, Megan; Carré-Camps, Marc; Perdomo-Celis, Federico; Narváez, Carlos F; Salgado, Doris M; Senthoor, Dewahar; O'Grady, Madeline; Phillips, Elizabeth; Durbin, Ann; Fandos, Diana; Miyazaki, Hikaru; Yen, Chun-Wan; Gélvez-Ramírez, Margarita; Warke, Rajas V; Ribeiro, Lucas S; Teixeira, Mauro M; Almeida, Roque P; Muñóz-Medina, José E; Ludert, Juan E; Nogueira, Mauricio L; Colombo, Tatiana E; Terzian, Ana C B; Bozza, Patricia T; Calheiros, Andrea S; Vieira, Yasmine R; Barbosa-Lima, Giselle; Vizzoni, Alexandre; Cerbino-Neto, José; Bozza, Fernando A; Souza, Thiago M L; Trugilho, Monique R O; de Filippis, Ana M B; de Sequeira, Patricia C; Marques, Ernesto T A; Magalhaes, Tereza; Díaz, Francisco J; Restrepo, Berta N; Marín, Katerine; Mattar, Salim; Olson, Daniel; Asturias, Edwin J; Lucera, Mark; Singla, Mohit; Medigeshi, Guruprasad R; de Bosch, Norma; Tam, Justina; Gómez-Márquez, Jose; Clavet, Charles; Villar, Luis; Hamad-Schifferli, Kimberly; Gehrke, Lee

2017-09-27

The recent Zika virus (ZIKV) outbreak demonstrates that cost-effective clinical diagnostics are urgently needed to detect and distinguish viral infections to improve patient care. Unlike dengue virus (DENV), ZIKV infections during pregnancy correlate with severe birth defects, including microcephaly and neurological disorders. Because ZIKV and DENV are related flaviviruses, their homologous proteins and nucleic acids can cause cross-reactions and false-positive results in molecular, antigenic, and serologic diagnostics. We report the characterization of monoclonal antibody pairs that have been translated into rapid immunochromatography tests to specifically detect the viral nonstructural 1 (NS1) protein antigen and distinguish the four DENV serotypes (DENV1-4) and ZIKV without cross-reaction. To complement visual test analysis and remove user subjectivity in reading test results, we used image processing and data analysis for data capture and test result quantification. Using a 30-μl serum sample, the sensitivity and specificity values of the DENV1-4 tests and the pan-DENV test, which detects all four dengue serotypes, ranged from 0.76 to 1.00. Sensitivity/specificity for the ZIKV rapid test was 0.81/0.86, respectively, using a 150-μl serum input. Serum ZIKV NS1 protein concentrations were about 10-fold lower than corresponding DENV NS1 concentrations in infected patients; moreover, ZIKV NS1 protein was not detected in polymerase chain reaction-positive patient urine samples. Our rapid immunochromatography approach and reagents have immediate application in differential clinical diagnosis of acute ZIKV and DENV cases, and the platform can be applied toward developing rapid antigen diagnostics for emerging viruses. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Rapid antigen detection test for group A streptococcus in children with pharyngitis.

PubMed

Cohen, Jérémie F; Bertille, Nathalie; Cohen, Robert; Chalumeau, Martin

2016-07-04

Group A streptococcus (GAS) accounts for 20% to 40% of cases of pharyngitis in children; the remaining cases are caused by viruses. Compared with throat culture, rapid antigen detection tests (RADTs) offer diagnosis at the point of care (within five to 10 minutes). To determine the diagnostic accuracy of RADTs for diagnosing GAS in children with pharyngitis. To assess the relative diagnostic accuracy of the two major types of RADTs (enzyme immunoassays (EIA) and optical immunoassays (OIA)) by indirect and direct comparison. We searched CENTRAL, MEDLINE, EMBASE, Web of Science, CDSR, DARE, MEDION and TRIP (January 1980 to July 2015). We also conducted related citations tracking via PubMed, handsearched reference lists of included studies and relevant review articles, and screened all articles citing included studies via Google Scholar. We included studies that compared RADT for GAS pharyngitis with throat culture on a blood agar plate in a microbiology laboratory in children seen in ambulatory care. Two review authors independently screened titles and abstracts for relevance, assessed full texts for inclusion, and carried out data extraction and quality assessment using the QUADAS-2 tool. We used bivariate meta-analysis to estimate summary sensitivity and specificity, and to investigate heterogeneity across studies. We compared the accuracy of EIA and OIA tests using indirect and direct evidence. We included 98 unique studies in the review (116 test evaluations; 101,121 participants). The overall methodological quality of included studies was poor, mainly because many studies were at high risk of bias regarding patient selection and the reference standard used (in 73% and 43% of test evaluations, respectively). In studies in which all participants underwent both RADT and throat culture (105 test evaluations; 58,244 participants; median prevalence of participants with GAS was 29.5%), RADT had a summary sensitivity of 85.6%; 95% confidence interval (CI) 83.3 to 87.6 and a summary specificity of 95.4%; 95% CI 94.5 to 96.2. There was substantial heterogeneity in sensitivity across studies; specificity was more stable. There was no evidence of a trade-off between sensitivity and specificity. Heterogeneity in accuracy was not explained by study-level characteristics such as whether an enrichment broth was used before plating, mean age and clinical severity of participants, and GAS prevalence. The sensitivity of EIA and OIA tests was comparable (summary sensitivity 85.4% versus 86.2%). Sensitivity analyses showed that summary estimates of sensitivity and specificity were stable in low risk of bias studies. In a population of 1000 children with a GAS prevalence of 30%, 43 patients with GAS will be missed. Whether or not RADT can be used as a stand-alone test to rule out GAS will depend mainly on the epidemiological context. The sensitivity of EIA and OIA tests seems comparable. RADT specificity is sufficiently high to ensure against unnecessary use of antibiotics. Based on these results, we would expect that amongst 100 children with strep throat, 86 would be correctly detected with the rapid test while 14 would be missed and not receive antibiotic treatment.

Development of an isothermal recombinase polymerase amplification assay for rapid detection of pseudorabies virus.

PubMed

Yang, Yang; Qin, Xiaodong; Zhang, Wei; Li, Zhiyong; Zhang, Shuaijun; Li, Yanmin; Zhang, Zhidong

2017-06-01

Recombinase polymerase amplification assays using real-time fluorescent detection (real-time RPA assay) and lateral flow dipstick (RPA LFD assay) were developed targeting the gD gene of pseudorabies virus (PRV). Both assays were performed at 39 °C within 20 min. The sensitivity of the real-time RPA assay and the RPA LFD assay was 100 copies per reaction and 160 copies per reaction, respectively. Both assays did not detect DNAs from other virus or PRV negative samples. Therefore, the developed RPA assays provide a rapid, simple, sensitive and specific alternative tool for detection of PRV. Copyright © 2017. Published by Elsevier Ltd.

Prospective Comparison of a New Chromogenic Medium, MRSASelect, to CHROMagar MRSA and Mannitol-Salt Medium Supplemented with Oxacillin or Cefoxitin for Detection of Methicillin-Resistant Staphylococcus aureus

PubMed Central

Stoakes, Luba; Reyes, Romina; Daniel, Janis; Lennox, Gwen; John, Michael A.; Lannigan, Robert; Hussain, Zafar

2006-01-01

MRSASelect agar was compared to CHROMagar, mannitol-salt agar with oxacillin, and mannitol-salt agar with cefoxitin (MSA-CFOX) for the isolation of methicillin-resistant Staphylococcus aureus (MRSA). The sensitivities and specificities were 97.3% and 99.8%, 82.9% and 99.1%, 80.2% and 79%, and 99.1% and 84.8%, respectively. MSA-CFOX and MRSASelect had a high sensitivity. MRSASelect, however, was more specific and proved to be a more reliable and rapid medium for the detection of MRSA. PMID:16455933

Malaria rapid diagnostic tests in elimination settings—can they find the last parasite?

PubMed Central

McMorrow, M. L.; Aidoo, M.; Kachur, S. P.

2016-01-01

Rapid diagnostic tests (RDTs) for malaria have improved the availability of parasite-based diagnosis throughout the malaria-endemic world. Accurate malaria diagnosis is essential for malaria case management, surveillance, and elimination. RDTs are inexpensive, simple to perform, and provide results in 15–20 min. Despite high sensitivity and specificity for Plasmodium falciparum infections, RDTs have several limitations that may reduce their utility in low-transmission settings: they do not reliably detect low-density parasitaemia (≤200 parasites/μL), many are less sensitive for Plasmodium vivax infections, and their ability to detect Plasmodium ovale and Plasmodium malariae is unknown. Therefore, in elimination settings, alternative tools with higher sensitivity for low-density infections (e.g. nucleic acid-based tests) are required to complement field diagnostics, and new highly sensitive and specific field-appropriate tests must be developed to ensure accurate diagnosis of symptomatic and asymptomatic carriers. As malaria transmission declines, the proportion of low-density infections among symptomatic and asymptomatic persons is likely to increase, which may limit the utility of RDTs. Monitoring malaria in elimination settings will probably depend on the use of more than one diagnostic tool in clinical-care and surveillance activities, and the combination of tools utilized will need to be informed by regular monitoring of test performance through effective quality assurance. PMID:21910780

A luminescent hybridoma-based biosensor for rapid detection of V. cholerae upon induction of calcium signaling pathway.

PubMed

Zamani, Parichehr; Sajedi, Reza H; Hosseinkhani, Saman; Zeinoddini, Mehdi; Bakhshi, Bita

2016-05-15

In this study, a hybridoma based biosensor was developed for rapid, sensitive and selective detection of Vibrio cholerae O1 which converts the antibody-antigen binding to bioluminescence light. After investigation on hybridoma performance, the biosensor was constructed by transfecting specific hybridoma cells with aequorin reporter gene and the bioluminescence activities of stable biosensor were measured. The sensitivity of biosensor was as few as 50 CFU/ml and it showed no responses to other entric bacteria. Moreover, the response time of biosensor was estimated in 7th second which means this method is considerably faster than many available detection assays. In addition, this biosensor was successfully applied to V. cholerae detection in environmental samples with no significant loss in sensitivity, demonstrating our proposed biosensor provides a sensitive and reliable method for detection of V. cholerae in natural samples. The application of whole hybridoma cell directly as a sensing element in biosensor construction which mentioned for the first time in present study suggests that hybridoma cells could provide a valuable tool for future studies in both basic and diagnostic sciences and could be considered as a fast and specific sensing element for detection of other pathogens in different applications. Copyright © 2015 Elsevier B.V. All rights reserved.

Development of an immunomagnetic separation-polymerase chain reaction (IMS-PCR) assay specific for Enterocytozoon bieneusi in water samples.

PubMed

Sorel, N; Guillot, E; Thellier, M; Accoceberry, I; Datry, A; Mesnard-Rouiller, L; Miégeville, M

2003-01-01

Microsporidia have become widely recognized as important human pathogens. Among Microsporidia, Enterocytozoon bieneusi is responsible for severe gastrointestinal disease. To date, no current therapy has been proven effective. Their mode of transmission and environmental occurrence are poorly documented because of the lack of detection methods that are both species-specific and sensitive. In this study, we developed a sensitive and specific molecular method to detect E. bieneusi spores in water samples. The molecular assay combined immunomagnetic separation (IMS) and polymerase chain reaction (PCR) amplification to detect E. bieneusi spores. A comparison was made of IMS magnetic beads coated with two different monoclonal antibodies, one specific for the Encephalitozoon genus that cross-reacts with E. bieneusi and the other specific only for the E. bieneusi species itself. Immunotech beads coated with the antibody specific for E. bieneusi were found to be the most effective combination. The highly specific IMS-PCR assay developed in this study provides a rapid and sensitive means of screening water samples for the presence of E. bieneusi spores.

Rapid genetic typing of diarrheagenic Escherichia coli using a two-tube modified molecular beacon based multiplex real-time PCR assay and its clinical application

PubMed Central

2014-01-01

Background Diarrheagenic Escherichia coli (DEC), including Enterotoxigenic E.coli (ETEC), Enteroaggregative E.coli (EAEC), Enteropathogenic E.coli (EPEC), Enterohemolysin E.coli (EHEC) and Enteroinvasive E.coli (EIEC) causes diarrhea or hemolytic uremic syndromes among infants and travelers around the world. A rapid, reliable and repeatable method is urgent for identifying DEC so as to provide the reference for responding to diarrheal disease outbreak and the treatment of the diarrheal patients associated with DEC. Methods In this study, specific primers and modified molecular beacon probes of nine specific virulence genes, whose 5′end were added with homo tail sequence, were designed; and a two-tube modified molecular beacon based multiplex real–time PCR (rtPCR) assay for the identification of five Escherichia coli pathotypes, including ETEC, EAEC, EPEC, EHEC and EIEC was developed and optimized. Totally 102 bacterial strains, including 52 reference bacterial strains and 50 clinical strains were detected to confirm whether the target genes selected were specific. Then detection limits of the assay were tested. Lastly, the assay was applied to the detection of 11860 clinical samples to evaluate the specificity and sensitivity of the developed assay compared with the conventional PCR. Results The target genes were 100% specific as assessed on 102 bacterial strains since no cross-reactions were observed. The detection limits ranged from 88 CFU/mL (EHEC) to 880 CFU/mL (EPEC). Compared with the conventional PCR, the specificity and sensitivity of the multiplex rtPCR was 100% and over 99%, respectively. The coefficient of variation (CV) for each target gene ranged from 0.45% to 1.53%. 171 positive clinical samples were mostly identified as ETEC (n = 111, 64.9%) and EPEC (n = 38, 22.2%), which were the dominating pathotypes of DEC strains. Conclusion The developed multiplex rtPCR assay for the identification of DEC was high sensitive and specific and could be applied to the rapid identification of DEC in clinical and public health laboratories. PMID:25023669

Robust detection of rare species using environmental DNA: The importance of primer specificity

Treesearch

Taylor M. Wilcox; Kevin S. McKelvey; Michael K. Young; Stephen F. Jane; Winsor H. Lowe; Andrew R. Whiteley; Michael K. Schwartz

2013-01-01

Environmental DNA (eDNA) is being rapidly adopted as a tool to detect rare animals. Quantitative PCR (qPCR) using probebased chemistries may represent a particularly powerful tool because of the method's sensitivity, specificity, and potential to quantify target DNA. However, there has been little work understanding the performance of these assays in the presence...

Polymeric LabChip Real-Time PCR as a Point-of-Care-Potential Diagnostic Tool for Rapid Detection of Influenza A/H1N1 Virus in Human Clinical Specimens

PubMed Central

Song, Hyun-Ok; Kim, Je-Hyoung; Ryu, Ho-Sun; Lee, Dong-Hoon; Kim, Sun-Jin; Kim, Deog-Joong; Suh, In Bum; Choi, Du Young; In, Kwang-Ho; Kim, Sung-Woo; Park, Hyun

2012-01-01

It is clinically important to be able to detect influenza A/H1N1 virus using a fast, portable, and accurate system that has high specificity and sensitivity. To achieve this goal, it is necessary to develop a highly specific primer set that recognizes only influenza A viral genes and a rapid real-time PCR system that can detect even a single copy of the viral gene. In this study, we developed and validated a novel fluidic chip-type real-time PCR (LabChip real-time PCR) system that is sensitive and specific for the detection of influenza A/H1N1, including the pandemic influenza strain A/H1N1 of 2009. This LabChip real-time PCR system has several remarkable features: (1) It allows rapid quantitative analysis, requiring only 15 min to perform 30 cycles of real-time PCR. (2) It is portable, with a weight of only 5.5 kg. (3) The reaction cost is low, since it uses disposable plastic chips. (4) Its high efficiency is equivalent to that of commercially available tube-type real-time PCR systems. The developed disposable LabChip is an economic, heat-transferable, light-transparent, and easy-to-fabricate polymeric chip compared to conventional silicon- or glass-based labchip. In addition, our LabChip has large surface-to-volume ratios in micro channels that are required for overcoming time consumed for temperature control during real-time PCR. The efficiency of the LabChip real-time PCR system was confirmed using novel primer sets specifically targeted to the hemagglutinin (HA) gene of influenza A/H1N1 and clinical specimens. Eighty-five human clinical swab samples were tested using the LabChip real-time PCR. The results demonstrated 100% sensitivity and specificity, showing 72 positive and 13 negative cases. These results were identical to those from a tube-type real-time PCR system. This indicates that the novel LabChip real-time PCR may be an ultra-fast, quantitative, point-of-care-potential diagnostic tool for influenza A/H1N1 with a high sensitivity and specificity. PMID:23285281

Direct detection of methicillin-resistant Staphylococcus aureus from blood cultures using an immunochromatographic immunoassay-based MRSA rapid kit for the detection of penicillin-binding protein 2a.

PubMed

Shin, Kyeong Seob; Song, Hyung Geun; Kim, Haejung; Yoon, Sangsun; Hong, Seung Bok; Koo, Sun Hoe; Kim, Jimyung; Kim, Jongwan; Roh, Kyoung Ho

2010-07-01

Using an EZ-Step MRSA rapid kit, a novel screening test for methicillin-resistant Staphylococcus aureus (MRSA) that detects penicillin-binding protein 2a, 34 of 36 MRSA-positive clinical blood culture samples were positive on direct testing (sensitivity, 94.4%), whereas 21 of 21 methicillin-susceptible S. aureus-positive samples were negative (specificity, 100%).

Development of a rapid diagnostic assay for the detection of tomato chlorotic dwarf viroid based on isothermal reverse-transcription-recombinase polymerase amplification.

PubMed

Hammond, Rosemarie W; Zhang, Shulu

2016-10-01

A molecular diagnostic assay utilizing reverse transcription-recombinase polymerase amplification (RT-RPA) at an isothermal constant temperature of 39°C and target-specific primers and probe were developed for the rapid, sensitive, and specific detection of tomato chlorotic dwarf viroid (TCDVd) in infected leaf and seed tissues. The performance of the AmplifyRP(®) Acceler8™ RT-RPA diagnostic assay, utilizing a lateral flow strip contained within an amplicon detection chamber, was evaluated and the results were compared with a standard RT-PCR assay. The AmplifyRP(®) Acceler8™ assay was specific for TCDVd in leaf and seed tissues, its sensitivity was comparable to conventional RT-PCR in leaf tissues, and it does not require extensive sample purification, specialized equipment, or technical expertise. This is the first report utilizing an RT-RPA assay to detect viroids and the assay can be used both in the laboratory and in the field for TCDVd detection. Published by Elsevier B.V.

Rapid HIV-1 testing during labor: a multicenter study.

PubMed

Bulterys, Marc; Jamieson, Denise J; O'Sullivan, Mary Jo; Cohen, Mardge H; Maupin, Robert; Nesheim, Steven; Webber, Mayris P; Van Dyke, Russell; Wiener, Jeffrey; Branson, Bernard M

2004-07-14

Timely testing of women in labor with undocumented human immunodeficiency virus (HIV) status could enable immediate provision of antiretroviral prophylaxis. To determine the feasibility and acceptance of rapid HIV testing among women in labor and to assess rapid HIV assay performance. The Mother-Infant Rapid Intervention At Delivery (MIRIAD) study implemented 24-hour counseling and voluntary rapid HIV testing for women in labor at 16 US hospitals from November 16, 2001, through November 15, 2003. A rapid HIV-1 antibody test for whole blood was used. Acceptance of HIV testing; sensitivity, specificity, and predictive value of the rapid test; time from blood collection to patient notification of results. There were 91,707 visits to the labor and delivery units in the study, 7381 of which were by eligible women without documentation of HIV testing. Of these, 5744 (78%) women were approached for rapid HIV testing and 4849 (84%) consented. HIV-1 test results were positive for 34 women (prevalence = 7/1000). Sensitivity and specificity of the rapid test were 100% and 99.9%, respectively; positive predictive value was 90% compared with 76% for enzyme immunoassay (EIA). Factors independently associated with higher test acceptance included younger age, being black or Hispanic, gestational age less than 32 weeks, and having had no prenatal care. Lower acceptance was associated with being admitted between 4 pm and midnight, particularly on Friday nights, but this may be explained in part by fewer available personnel. Median time from blood collection to patient notification of result was 66 minutes (interquartile range, 45-120 minutes), compared with 28 hours for EIA (P<.001). Rapid HIV testing is feasible and delivers accurate and timely test results for women in labor. It provides HIV-positive women prompt access to intrapartum and neonatal antiretroviral prophylaxis, proven to reduce perinatal HIV transmission, and may be particularly applicable to higher-risk populations.

Rapid tests for the diagnosis of visceral leishmaniasis in patients with suspected disease

PubMed Central

Boelaert, Marleen; Verdonck, Kristien; Menten, Joris; Sunyoto, Temmy; van Griensven, Johan; Chappuis, Francois; Rijal, Suman

2014-01-01

Background The diagnosis of visceral leishmaniasis (VL) in patients with fever and a large spleen relies on showing Leishmania parasites in tissue samples and on serological tests. Parasitological techniques are invasive, require sophisticated laboratories, consume time, or lack accuracy. Recently, rapid diagnostic tests that are easy to perform have become available. Objectives To determine the diagnostic accuracy of rapid tests for diagnosing VL in patients with suspected disease presenting at health services in endemic areas. Search methods We searched MEDLINE, EMBASE, LILACS, CIDG SR, CENTRAL, SCI-expanded, Medion, Arif, CCT, and the WHO trials register on 3 December 2013, without applying language or date limits. Selection criteria This review includes original, phase III, diagnostic accuracy studies of rapid tests in patients clinically suspected to have VL. As reference standards, we accepted: (1) direct smear or culture of spleen aspirate; (2) composite reference standard based on one or more of the following: parasitology, serology, or response to treatment; and (3) latent class analysis. Data collection and analysis Two review authors independently extracted data and assessed quality of included studies using the QUADAS-2 tool. Discrepancies were resolved by a third author. We carried out a meta-analysis to estimate sensitivity and specificity of rapid tests, using a bivariate normal model with a complementary log-log link function. We analysed each index test separately. As possible sources of heterogeneity, we explored: geographical area, commercial brand of index test, type of reference standard, disease prevalence, study size, and risk of bias (QUADAS-2). We also undertook a sensitivity analysis to assess the influence of imperfect reference standards. Main results Twenty-four studies containing information about five index tests (rK39 immunochromatographic test (ICT), KAtex latex agglutination test in urine, FAST agglutination test, rK26 ICT, and rKE16 ICT) recruiting 4271 participants (2605 with VL) were included. We carried out a meta-analysis for the rK39 ICT (including 18 studies; 3622 participants) and the latex agglutination test (six studies; 1374 participants). The results showed considerable heterogeneity. For the rK39 ICT, the overall sensitivity was 91.9% (95% confidence interval (95% CI) 84.8 to 96.5) and the specificity 92.4% (95% CI 85.6 to 96.8). The sensitivity was lower in East Africa (85.3%; 95% CI 74.5 to 93.2) than in the Indian subcontinent (97.0%; 95% CI 90.0 to 99.5). For the latex agglutination test, overall sensitivity was 63.6% (95% CI 40.9 to 85.6) and specificity 92.9% (95% CI 76.7 to 99.2). Authors' conclusions The rK39 ICT shows high sensitivity and specificity for the diagnosis of visceral leishmaniasis in patients with febrile splenomegaly and no previous history of the disease, but the sensitivity is notably lower in east Africa than in the Indian subcontinent. Other rapid tests lack accuracy, validation, or both. PLAIN LANGUAGE SUMMARY Rapid diagnostic tests for visceral leishmaniasis Visceral leishmaniasis (or kala-azar) is caused by a parasite, results in fever, a large spleen and other health problems, occuring in India, Bangladesh and Nepal, east Africa, the Mediterranean region and Brazil. Without treatment people die, and proper treatment can result in cure, so diagnosis is important. Many of the tests that are used to determine if a person has visceral leishmaniasis are complicated, costly, painful and sometimes dangerous for the patients. Now rapid diagnostic tests that are safe and easy to perform are available. This Cochrane review describes how accurate these rapid diagnostic tests are for diagnosing visceral leishmaniasis. We summarize those studies that evaluated the rapid tests in people who, according to their physicians, could have the disease. We only included studies in which the researchers had used established methods to distinguish the people with visceral leishmaniasis from those who did not have the disease. We found 24 studies, which contained information about five different rapid tests. A total of 4271 people participated in these studies. One of the rapid tests (called the rK39 immunochromatographic test) gave correct, positive results in 92% of the people with visceral leishmaniasis and it gave correct, negative results in 92% of the people who did not have the disease. This test worked better in India and Nepal than in east Africa. In India and Nepal, it gave correct, positive results in 97% of the people with the disease. In east Africa, it gave correct, positive results in only 85% of the people with the disease. A second rapid test (called latex agglutination test) gave correct, positive results in 64% of the people with the disease and it gave correct, negative results in 93% of the people without the disease. For the other rapid tests evaluated, there are too few studies to know how accurate they are. PMID:24947503

Comparison of nonstructural protein-1 antigen detection by rapid and enzyme-linked immunosorbent assay test and its correlation with polymerase chain reaction for early diagnosis of dengue

PubMed Central

Gaikwad, Seema; Sawant, Sandhya S.; Shastri, Jayanthi S.

2017-01-01

INTRODUCTION: Early diagnosis of dengue is important for appropriate clinical management and vector control. Different serological tests based on the principle of immunochromatography and enzyme-linked immunosorbent assay (ELISA) are commonly used for detection of antigen and antibodies of dengue virus. The performance of these tests depends on the sensitivity and specificity. Hence, the study was undertaken to compare nonstructural protein-1 (NS1) antigen detection by rapid and ELISA with real-time polymerase chain reaction (RT-PCR) for diagnosis of dengue. MATERIALS AND METHODS: Prospective laboratory study was carried out on sera samples (n = 200) from clinically suspected cases of dengue. The sera samples were subjected for NS1 antigen detection test by rapid test, NS1 ELISA, and RT-PCR. The results of rapid and ELISA tests were compared with real Time PCR. RESULTS: The sensitivity, specificity, positive, and negative predictive value of rapid dengue NS1 antigen test were 81.5%, 66.7%, 78.2%, and 71.1%, respectively whereas that of NS1 ELISA were 89.9%, 100%, 100%, and 94%, respectively. Concordance of Rapid NS1 and NS1 ELISA with PCR was 75.5% and 94%. DISCUSSION AND CONCLUSION: NS1 antigen ELISA can be implemented in diagnostic laboratories for diagnosis of dengue in the acute phase of illness. The test also has great potential value for use in epidemic situations, as it could facilitate the early screening of patients and limit disease expansion. PMID:28706387

Development of a rapid diagnostic method for identification of Staphylococcus aureus and antimicrobial resistance in positive blood culture bottles using a PCR-DNA-chromatography method.

PubMed

Ohshiro, Takeya; Miyagi, Chihiro; Tamaki, Yoshikazu; Mizuno, Takuya; Ezaki, Takayuki

2016-06-01

Blood culturing and the rapid reporting of results are essential for infectious disease clinics to obtain bacterial information that can affect patient prognosis. When gram-positive coccoid cells are observed in blood culture bottles, it is important to determine whether the strain is Staphylococcus aureus and whether the strain has resistance genes, such as mecA and blaZ, for proper antibiotic selection. Previous work led to the development of a PCR method that is useful for rapid identification of bacterial species and antimicrobial susceptibility. However, that method has not yet been adopted in community hospitals due to the high cost and methodological complexity. We report here the development of a quick PCR and DNA-chromatography test, based on single-tag hybridization chromatography, that permits detection of S. aureus and the mecA and blaZ genes; results can be obtained within 1 h for positive blood culture bottles. We evaluated this method using 42 clinical isolates. Detection of S. aureus and the resistance genes by the PCR-DNA-chromatography method was compared with that obtained via the conventional identification method and actual antimicrobial susceptibility testing. Our method had a sensitivity of 97.0% and a specificity of 100% for the identification of the bacterial species. For the detection of the mecA gene of S. aureus, the sensitivity was 100% and the specificity was 95.2%. For the detection of the blaZ gene of S. aureus, the sensitivity was 100% and the specificity was 88.9%. The speed and simplicity of this PCR-DNA-chromatography method suggest that our method will facilitate rapid diagnoses. Copyright © 2016 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Venous glucose, serum lactate and base deficit as biochemical predictors of mortality in patients with polytrauma.

PubMed

Saad, Sameh; Mohamed, Naglaa; Moghazy, Amr; Ellabban, Gouda; El-Kamash, Soliman

2016-01-01

The trauma and injury severity score (TRISS) and Acute Physiology and Chronic Health Evaluation IV (APACHE IV) are accurate but complex. This study aimed to compare venous glucose, levels of serum lactate, and base deficit in polytraumatized patients as simple parameters to predict the mortality in these patients versus (TRISS) and (APACHE IV). This was a comparative cross-sectional study of 282 patients with polytrauma presented to the Emergency Department (ED). The best cut off value of TRISS probability of survival score for prediction of mortality among poly-traumatized patients was ≤90. APACHE IV demonstrated 67% sensitivity and 95% specificity at 95% CI at cut off point 99. The best cutoff value of Random Blood Sugar was >140 mg/dl, with 89% sensitivity, 49% specificity; base deficit was less than -5.6 with 64% sensitivity, 93% specificity; lactate was >2.6 mmol/L with 92%, sensitivity, 42% specificity. Venous glucose, serum lactate and base deficit are easy and rapid biochemical predictors of mortality in patients with polytrauma. These predictors could be used as TRISS and APACHE IV in predicting mortality.

Blinded study determination of high sensitivity and specificity microchip electrophoresis–SSCP/HA to detect mutations in the p53 gene

PubMed Central

Hestekin, Christa N.; Lin, Jennifer S.; Senderowicz, Lionel; Jakupciak, John P.; O’Connell, Catherine; Rademaker, Alfred; Barron, Annelise E.

2012-01-01

Knowledge of the genetic changes that lead to disease has grown and continues to grow at a rapid pace. However, there is a need for clinical devices that can be used routinely to translate this knowledge into the treatment of patients. Use in a clinical setting requires high sensitivity and specificity (>97%) in order to prevent misdiagnoses. Single strand conformational polymorphism (SSCP) and heteroduplex analysis (HA) are two DNA-based, complementary methods for mutation detection that are inexpensive and relatively easy to implement. However, both methods are most commonly detected by slab gel electrophoresis, which can be labor-intensive, time-consuming, and often the methods are unable to produce high sensitivity and specificity without the use of multiple analysis conditions. Here we demonstrate the first blinded study using microchip electrophoresis-SSCP/HA. We demonstrate the ability of microchip electrophoresis-SSCP/HA to detect with 98% sensitivity and specificity >100 samples from the p53 gene exons 5–9 in a blinded study in an analysis time of less than 10 minutes. PMID:22002021

Multi-colored immunochromatography using nanobeads for rapid and sensitive typing of seasonal influenza viruses.

PubMed

Sakurai, Akira; Takayama, Katsuyoshi; Nomura, Namiko; Yamamoto, Naoki; Sakoda, Yoshihiro; Kobayashi, Yukuharu; Kida, Hiroshi; Shibasaki, Futoshi

2014-12-01

Immunochromatography (IC) is an antigen-detection assay that plays an important role in the rapid diagnosis of influenza viruses because of its rapid turnaround and ease of use. Despite the usefulness of IC, the limit of detection of common IC kits is as high as 10(3)-10(4) plaque forming units (pfu) per reaction, resulting in their limited sensitivities. Early diagnosis within 24h would provide more appropriate timing of treatment. In this study, a multi-colored NanoAct™ bead IC was established to detect seasonal influenza viruses. This method has approximately 10-fold higher sensitivity than that of colloidal gold or colored latex bead IC assays, and does not require specific instruments. More notably, NanoAct™ bead IC can distinguish influenza A and B viruses from clinical samples with a straightforward readout composed of colored lines. Our results will provide new strategies for the diagnosis, treatment, and a chance to survey of influenza viruses in developing countries and in the field research. Copyright © 2014 Elsevier B.V. All rights reserved.

Rapid and sensitive detection of Lily symptomless virus by reverse transcription loop-mediated isothermal amplification.

PubMed

He, Xiangfeng; Xue, Fei; Xu, Shufa; Wang, Wenhe

2016-12-01

Lily symptomless virus (LSV) is one of the most prevalent viruses that infect lily plants worldwide. A rapid and sensitive reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for detection of LSV, using two primer pairs that specifically amplified the conserved sequence of LSV coat protein. The optimum reaction conditions were as follows: 4mM MgCl 2 and 0.8M betaine with incubation at 64°C for 30min. The limit of detection of LSV from infected lily leaves was 10-fold higher for RT-LAMP than for conventional RT-PCR. Moreover, RT-LAMP detected LSV in not only symptomatic, but also in symptomless tissues of infected plants. These findings indicate that our RT-LAMP method for LSV can serve as a low-cost, simple, and rapid alternative to conventional detection assays. Copyright © 2016 Elsevier B.V. All rights reserved.

Rapid detection of infectious hypodermal and hematopoietic necrosis virus (IHHNV) by real-time, isothermal recombinase polymerase amplification assay.

PubMed

Xia, Xiaoming; Yu, Yongxin; Hu, Linghao; Weidmann, Manfred; Pan, Yingjie; Yan, Shuling; Wang, Yongjie

2015-04-01

Infectious hypodermal and hematopoietic necrosis virus (IHHNV) causes mortality or runt deformity syndrome in penaeid shrimps and is responsible for significant economic losses in the shrimp aquaculture industry. Here, we describe a novel real-time isothermal recombinase polymerase amplification (RPA) assay developed for IHHNV detection. Using IHHNV plasmid standards and DNA samples from a variety of organisms, we evaluated the ability of the IHHNV-RPA assay to detect IHHNV based on analysis of its sensitivity, specificity, rapidity, and reproducibility. Probit analysis of eight independent experimental replicates indicated satisfactory performance of the RPA assay, which is sufficiently sensitive to detect as few as 4 copies of the IHHNV genome within 7 min at 39 °C with 95 % reliability. Therefore, this rapid RPA method has great potential for applications, either in field use or as a point of care diagnostic technique.

Development of a human-specific B. thetaiotaomicron IMS ...

EPA Pesticide Factsheets

Immunomagnetic separation/adenosine triphosphate (IMS/ATP) assays utilize paramagnetic beads and target-specific antibodies to isolate target organisms. Following isolation, adenosine tri-phosphate (ATP) is extracted from the target population and quantified. An inversely-coupled (Inv-IMS/ATP)assay for detection of Bacteroides thetaiotaomicron was developed and applied for rapid detection of human-associated fecal contamination in surface waters in Baja California. Specificity of the assay was tested against challenge solutions of varying concentrations of dog, gull, horse and chicken feces, and a field validation survey of coastal and WWTP effluent water quality in Rosarito and Enseneda, Baja California was conducted. Inv IMS/ATP measurements made shown to be specific and sensitive to human fecal contamination. At test concentrations of less than 1000 MPN ENT/100 mL, sensitivity and specificity of the assay both exceeded 80%. Moreover, the Inv-IMS/ATP assay yielded measurements of viable B. thetaiotaomicron that were comparable to the HF183 human marker in complex surface waters impacted with both wastewater and runoff, and the Inv-IMS/ATP assay was able to effectively differentiate between surface waters impacted with adequately and inadequately treated wastewater. The Inv-IMS/ATP assays shows promise for rapid evaluation of recreational water quality in areas where access to more expensive methods is limited and in areas where water quality in unpredicta

Seasonal performance of a malaria rapid diagnosis test at community health clinics in a malaria-hyperendemic region of Burkina Faso

PubMed Central

2012-01-01

Backgound Treatment of confirmed malaria patients with Artemisinin-based Combination Therapy (ACT) at remote areas is the goal of many anti-malaria programs. Introduction of effective and affordable malaria Rapid Diagnosis Test (RDT) in remote areas could be an alternative tool for malaria case management. This study aimed to assess performance of the OptiMAL dipstick for rapid malaria diagnosis in children under five. Methods Malaria symptomatic and asymptomatic children were recruited in a passive manner in two community clinics (CCs). Malaria diagnosis by microscopy and RDT were performed. Performance of the tests was determined. Results RDT showed similar ability (61.2%) to accurately diagnose malaria as microscopy (61.1%). OptiMAL showed a high level of sensitivity and specificity, compared with microscopy, during both transmission seasons (high & low), with a sensitivity of 92.9% vs. 74.9% and a specificity of 77.2% vs. 87.5%. Conclusion By improving the performance of the test through accurate and continuous quality control of the device in the field, OptiMAL could be suitable for use at CCs for the management and control of malaria. PMID:22647557

Evaluation of a monoclonal antibody-based rapid immunochromatographic test for direct detection of rabies virus in the brain of humans and animals.

PubMed

Ahmed, Kamruddin; Wimalaratne, Omala; Dahal, Narapati; Khawplod, Pakamatz; Nanayakkara, Susilakanthi; Rinzin, Karma; Perera, Devika; Karunanayake, Dushantha; Matsumoto, Takashi; Nishizono, Akira

2012-04-01

Rabies diagnosis uses a direct fluorescent antibody test (FAT) that is difficult, costly, and time-consuming, and requires trained personnel. We developed a rapid immunochromatographic test (RICT) for the diagnosis of rabies. The efficacy of the RICT was compared with that of the FAT. Brain samples were collected from humans, dogs, cats, and other animals in Sri Lanka (n = 248), Bhutan (n = 27), and Thailand (n = 228). The sensitivity (0.74-0.95), specificity (0.98-1.0), positive predictive value (0.98-1.0), negative predictive value (0.75-0.97), accuracy (0.91-0.98), and kappa measure of agreement (0.79-0.93) were all satisfactory for animal samples and samples preserved in 50% glycerol saline solution. Because the RICT showed high sensitivity but low specificity with human brain samples, it is unsuitable for confirming rabies in humans. No amino acid substitutions were found in the antibody attachment sites of the nucleoprotein gene with FAT-positive, RICT-negative samples. The RICT is reliable, user friendly, rapid, robust, and can be used in laboratories with a modest infrastructure.

Highly sensitive and specific colorimetric detection of cancer cells via dual-aptamer target binding strategy.

PubMed

Wang, Kun; Fan, Daoqing; Liu, Yaqing; Wang, Erkang

2015-11-15

Simple, rapid, sensitive and specific detection of cancer cells is of great importance for early and accurate cancer diagnostics and therapy. By coupling nanotechnology and dual-aptamer target binding strategies, we developed a colorimetric assay for visually detecting cancer cells with high sensitivity and specificity. The nanotechnology including high catalytic activity of PtAuNP and magnetic separation & concentration plays a vital role on the signal amplification and improvement of detection sensitivity. The color change caused by small amount of target cancer cells (10 cells/mL) can be clearly distinguished by naked eyes. The dual-aptamer target binding strategy guarantees the detection specificity that large amount of non-cancer cells and different cancer cells (10(4) cells/mL) cannot cause obvious color change. A detection limit as low as 10 cells/mL with detection linear range from 10 to 10(5) cells/mL was reached according to the experimental detections in phosphate buffer solution as well as serum sample. The developed enzyme-free and cost effective colorimetric assay is simple and no need of instrument while still provides excellent sensitivity, specificity and repeatability, having potential application on point-of-care cancer diagnosis. Copyright © 2015 Elsevier B.V. All rights reserved.

Rapid Debris Analysis Project Task 3 Final Report - Sensitivity of Fallout to Source Parameters, Near-Detonation Environment Material Properties, Topography, and Meteorology

DOE Office of Scientific and Technical Information (OSTI.GOV)

Goldstein, Peter

2014-01-24

This report describes the sensitivity of predicted nuclear fallout to a variety of model input parameters, including yield, height of burst, particle and activity size distribution parameters, wind speed, wind direction, topography, and precipitation. We investigate sensitivity over a wide but plausible range of model input parameters. In addition, we investigate a specific example with a relatively narrow range to illustrate the potential for evaluating uncertainties in predictions when there are more precise constraints on model parameters.

CONDUCTING-POLYMER NANOWIRE IMMUNOSENSOR ARRAYS FOR MICROBIAL PATHOGENS

EPA Science Inventory

The lack of methods for routine rapid and sensitive detection and quantification of specific pathogens has limited the amount of information available on their occurrence in drinking water and other environmental samples. The nanowire biosensor arrays developed in this study w...

Rapid colorimetric detection of Zika virus from serum and urine specimens by reverse transcription loop-mediated isothermal amplification (RT-LAMP)

PubMed Central

Biggerstaff, Brad J.; Tanner, Nathan A.; Lauterbach, Molly; Lanciotti, Robert S.

2017-01-01

Zika virus (ZIKV) has emerged as a major global public health concern in the last two years due to its link as a causative agent of human birth defects. Its rapid expansion into the Western Hemisphere as well as the ability to be transmitted from mother to fetus, through sexual transmission and possibly through blood transfusions has increased the need for a rapid and expansive public health response to this unprecedented epidemic. A non-invasive and rapid ZIKV diagnostic screening assay that can be performed in a clinical setting throughout pregnancy is vital for prenatal care of women living in areas of the world where exposure to the virus is possible. To meet this need we have developed a sensitive and specific reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) assay to detect ZIKV RNA in urine and serum with a simple visual detection. RT-LAMP results were shown to have a limit of detection 10-fold higher than qRT-PCR. As little as 1.2 RNA copies/μl was detected by RT-LAMP from a panel of 178 diagnostic specimens. The assay was shown to be highly specific for ZIKV RNA when tested with diagnostic specimens positive for dengue virus (DENV) and chikungunya virus (CHIKV). The assay described here illustrates the potential for a fast, reliable, sensitive and specific assay for the detection of ZIKV from urine or serum that can be performed in a clinical or field setting with minimal equipment and technological expertise. PMID:28945787

Rapid colorimetric detection of Zika virus from serum and urine specimens by reverse transcription loop-mediated isothermal amplification (RT-LAMP).

PubMed

Calvert, Amanda E; Biggerstaff, Brad J; Tanner, Nathan A; Lauterbach, Molly; Lanciotti, Robert S

2017-01-01

Zika virus (ZIKV) has emerged as a major global public health concern in the last two years due to its link as a causative agent of human birth defects. Its rapid expansion into the Western Hemisphere as well as the ability to be transmitted from mother to fetus, through sexual transmission and possibly through blood transfusions has increased the need for a rapid and expansive public health response to this unprecedented epidemic. A non-invasive and rapid ZIKV diagnostic screening assay that can be performed in a clinical setting throughout pregnancy is vital for prenatal care of women living in areas of the world where exposure to the virus is possible. To meet this need we have developed a sensitive and specific reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) assay to detect ZIKV RNA in urine and serum with a simple visual detection. RT-LAMP results were shown to have a limit of detection 10-fold higher than qRT-PCR. As little as 1.2 RNA copies/μl was detected by RT-LAMP from a panel of 178 diagnostic specimens. The assay was shown to be highly specific for ZIKV RNA when tested with diagnostic specimens positive for dengue virus (DENV) and chikungunya virus (CHIKV). The assay described here illustrates the potential for a fast, reliable, sensitive and specific assay for the detection of ZIKV from urine or serum that can be performed in a clinical or field setting with minimal equipment and technological expertise.

Accuracy of dengue clinical diagnosis with and without NS1 antigen rapid test: Comparison between human and Bayesian network model decision.

PubMed

Sa-Ngamuang, Chaitawat; Haddawy, Peter; Luvira, Viravarn; Piyaphanee, Watcharapong; Iamsirithaworn, Sopon; Lawpoolsri, Saranath

2018-06-18

Differentiating dengue patients from other acute febrile illness patients is a great challenge among physicians. Several dengue diagnosis methods are recommended by WHO. The application of specific laboratory tests is still limited due to high cost, lack of equipment, and uncertain validity. Therefore, clinical diagnosis remains a common practice especially in resource limited settings. Bayesian networks have been shown to be a useful tool for diagnostic decision support. This study aimed to construct Bayesian network models using basic demographic, clinical, and laboratory profiles of acute febrile illness patients to diagnose dengue. Data of 397 acute undifferentiated febrile illness patients who visited the fever clinic of the Bangkok Hospital for Tropical Diseases, Thailand, were used for model construction and validation. The two best final models were selected: one with and one without NS1 rapid test result. The diagnostic accuracy of the models was compared with that of physicians on the same set of patients. The Bayesian network models provided good diagnostic accuracy of dengue infection, with ROC AUC of 0.80 and 0.75 for models with and without NS1 rapid test result, respectively. The models had approximately 80% specificity and 70% sensitivity, similar to the diagnostic accuracy of the hospital's fellows in infectious disease. Including information on NS1 rapid test improved the specificity, but reduced the sensitivity, both in model and physician diagnoses. The Bayesian network model developed in this study could be useful to assist physicians in diagnosing dengue, particularly in regions where experienced physicians and laboratory confirmation tests are limited.

Visual Detection of Canine Parvovirus Based on Loop-Mediated Isothermal Amplification Combined with Enzyme-Linked Immunosorbent Assay and with Lateral Flow Dipstick

PubMed Central

SUN, Yu-Ling; YEN, Chon-Ho; TU, Ching-Fu

2013-01-01

ABSTRACT Loop-mediated isothermal amplification (LAMP) combined with enzyme-linked immunosorbent assay (LAMP–ELISA) and with lateral flow dipstick (LAMP–LFD) are rapid, sensitive and specific methods for the visual detection of clinical pathogens. In this study, LAMP–ELISA and LAMP–LFD were developed for the visual detection of canine parvovirus (CPV). For LAMP, a set of four primers (biotin-labeled forward inner primers) was designed to specifically amplify a region of the VP2 gene of CPV. The optimum time and temperature for LAMP were 60 min and 65°C, respectively. The specific capture oligonucleotide probes, biotin-labeled CPV probe for LAMP–ELISA and fluorescein isothiocyanate-labeled CPV probe for LAMP–LFD were also designed for hybridization with LAMP amplicons on streptavidin-coated wells and LFD strips, respectively. For the comparison of detection sensitivity, conventional PCR and LAMP for CPV detection were also performed. The CPV detection limits by PCR, PCR–ELISA, LAMP, LAMP–ELISA and LAMP–LFD were 102, 102, 10−1, 10−1 and 10−1 TCID50/ml, respectively. In tests using artificially contaminated dog fecal samples, the samples with CPV inoculation levels of ≥1 TCID50/ml gave positive results by both LAMP–ELISA and LAMP–LFD. Our data indicated that both LAMP–ELISA and LAMP–LFD are promising as rapid, sensitive and specific methods for an efficient diagnosis of CPV infection. PMID:24334855

Diagnostic performance of influenza viruses and RSV rapid antigen detection tests in children in tertiary care.

PubMed

Moesker, F M; van Kampen, J J A; Aron, G; Schutten, M; van de Vijver, D A M C; Koopmans, M P G; Osterhaus, A D M E; Fraaij, P L A

2016-06-01

Rapid antigen detection tests (RADTs) are increasingly used to detect influenza viruses and respiratory syncytial virus (RSV). However, their sensitivity and specificity are a matter of debate, challenging their clinical usefulness. Comparing diagnostic performances of BinaxNow Influenza AB(®) (BNI) and BinaxNow RSV(®) (BNR), to those of real-time reverse transcriptase PCR (RT-PCR), virus isolation and direct immunofluorescence (D-IF) in paediatric patients. Between November 2005 and September 2013, 521 nasal washings from symptomatic children (age <5 years) attending our tertiary care centre were tested, with a combination of the respective assays using RT-PCR as gold standard. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of BNI were 69% (confidence interval [CI] [51-83]), 96% [94-97], 55% [39-70] and 98% [96-99] respectively. Of eleven false-negative samples, RT-PCR Ct-values were higher than all RT-PCR positive test results (27 vs 22, p=0.012). Of twenty false-positive samples, none were culture positive and two tested positive in D-IF. Sensitivity, specificity, PPV and NPV for BNR were 79% [73-85], 98% [96-99], 97% [93-99] and 88% [84-91]. Of the 42 false-negative samples the median Ct-value was higher than that of all RT-PCR positive samples (31 vs 23, p<0.0001). Five false-positive samples were detected. Three of these tested positive for RSV in virus isolation and D-IF. RADTs have a high specificity with BNR being superior to BNI. However, their relative low sensitivity limits their usefulness for clinical decision making in a tertiary care paediatric hospital. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Diagnostic value of animal-side antibody assays for rapid detection of Mycobacterium bovis or Mycobacterium microti infection in South American camelids.

PubMed

Lyashchenko, Konstantin P; Greenwald, Rena; Esfandiari, Javan; Rhodes, Shelley; Dean, Gillian; de la Rua-Domenech, Ricardo; Meylan, Mireille; Vordermeier, H Martin; Zanolari, Patrik

2011-12-01

Tuberculosis (TB) in South American camelids (SAC) is caused by Mycobacterium bovis or Mycobacterium microti. Two serological methods, rapid testing (RT) and the dual-path platform (DPP) assay, were evaluated using naturally infected SAC. The study population included 156 alpacas and 175 llamas in Great Britain, Switzerland, and the United States. TB due to M. bovis (n = 44) or M. microti (n = 8) in 35 alpacas and 17 llamas was diagnosed by gross pathology examination and culture. Control animals were from herds with no TB history. The RT and the DPP assay showed sensitivities of 71% and 74%, respectively, for alpacas, while the sensitivity for llamas was 77% for both assays. The specificity of the DPP assay (98%) was higher than that of RT (94%) for llamas; the specificities of the two assays were identical (98%) for alpacas. When the two antibody tests were combined, the parallel-testing interpretation (applied when either assay produced a positive result) enhanced the sensitivities of antibody detection to 89% for alpacas and 88% for llamas but at the cost of lower specificities (97% and 93%, respectively), whereas the serial-testing interpretation (applied when both assays produced a positive result) maximized the specificity to 100% for both SAC species, although the sensitivities were 57% for alpacas and 65% for llamas. Over 95% of the animals with evidence of TB failed to produce skin test reactions, thus confirming concerns about the validity of this method for testing SAC. The findings suggest that serological assays may offer a more accurate and practical alternative for antemortem detection of camelid TB.

Diagnostic Value of Animal-Side Antibody Assays for Rapid Detection of Mycobacterium bovis or Mycobacterium microti Infection in South American Camelids▿

PubMed Central

Lyashchenko, Konstantin P.; Greenwald, Rena; Esfandiari, Javan; Rhodes, Shelley; Dean, Gillian; de la Rua-Domenech, Ricardo; Meylan, Mireille; Vordermeier, HMartin; Zanolari, Patrik

2011-01-01

Tuberculosis (TB) in South American camelids (SAC) is caused by Mycobacterium bovis or Mycobacterium microti. Two serological methods, rapid testing (RT) and the dual-path platform (DPP) assay, were evaluated using naturally infected SAC. The study population included 156 alpacas and 175 llamas in Great Britain, Switzerland, and the United States. TB due to M. bovis (n = 44) or M. microti (n = 8) in 35 alpacas and 17 llamas was diagnosed by gross pathology examination and culture. Control animals were from herds with no TB history. The RT and the DPP assay showed sensitivities of 71% and 74%, respectively, for alpacas, while the sensitivity for llamas was 77% for both assays. The specificity of the DPP assay (98%) was higher than that of RT (94%) for llamas; the specificities of the two assays were identical (98%) for alpacas. When the two antibody tests were combined, the parallel-testing interpretation (applied when either assay produced a positive result) enhanced the sensitivities of antibody detection to 89% for alpacas and 88% for llamas but at the cost of lower specificities (97% and 93%, respectively), whereas the serial-testing interpretation (applied when both assays produced a positive result) maximized the specificity to 100% for both SAC species, although the sensitivities were 57% for alpacas and 65% for llamas. Over 95% of the animals with evidence of TB failed to produce skin test reactions, thus confirming concerns about the validity of this method for testing SAC. The findings suggest that serological assays may offer a more accurate and practical alternative for antemortem detection of camelid TB. PMID:22012976

Highly sensitive fluorescence detection of metastatic lymph nodes of gastric cancer with photo-oxidation of protoporphyrin IX.

PubMed

Koizumi, N; Harada, Y; Beika, M; Minamikawa, T; Yamaoka, Y; Dai, P; Murayama, Y; Yanagisawa, A; Otsuji, E; Tanaka, H; Takamatsu, T

2016-08-01

The establishment of a precise and rapid method to detect metastatic lymph nodes (LNs) is essential to perform less invasive surgery with reduced gastrectomy along with reduced lymph node dissection. We herein describe a novel imaging strategy to detect 5-aminolevulinic acid (5-ALA)-induced protoporphyrin IX (PpIX) fluorescence in excised LNs specifically with reduced effects of tissue autofluorescence based on photo-oxidation of PpIX. We applied the method in a clinical setting, and evaluated its feasibility. To reduce the unfavorable effect of autofluorescence, we focused on photo-oxidation of PpIX: Following light irradiation, PpIX changes into another substance, photo-protoporphyrin, via an oxidative process, which has a different spectral peak, at 675 nm, whereas PpIX has its spectral peak at 635 nm. Based on the unique spectral alteration, fluorescence spectral imaging before and after light irradiation and subsequent originally-developed image processing was performed. Following in vitro study, we applied this method to a total of 662 excised LNs obtained from 30 gastric cancer patients administered 5-ALA preoperatively. Specific visualization of PpIX was achieved in in vitro study. The method allowed highly sensitive detection of metastatic LNs, with sensitivity of 91.9% and specificity of 90.8% in the in vivo clinical trial. Receiver operating characteristic analysis indicated high diagnostic accuracy, with the area under the curve of 0.926. We established a highly sensitive and specific 5-ALA-induced fluorescence imaging method applicable in clinical settings. The novel method has a potential to become a useful tool for intraoperative rapid diagnosis of LN metastasis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Rapid visual detection of Mycobacterium avium subsp. paratuberculosis by recombinase polymerase amplification combined with a lateral flow dipstick

PubMed Central

Zhao, Guimin; Wang, Hongmei; Hou, Peili; He, Chengqiang

2018-01-01

Paratuberculosis (Johne's disease) is a chronic debilitating disease of domestic and wild ruminants. However, widespread point-of-care testing is infrequent due to the lack of a robust method. The isothermal recombinase polymerase amplification (RPA) technique has applied for rapid diagnosis. Herein, RPA combined with a lateral flow dipstick (LFD) assay was developed to estimate DNA from Mycobacterium avium subsp. paratuberculosis. First, analytical specificity and sensitivity of the RPA-nfo primer and probe sets were assessed. The assay successfully detected M. paratuberculosis DNA in 30 min at 39℃ with a detection limit of up to eight copies per reaction, which was equivalent to that of the real-time quantitative polymerase chain reaction (qPCR) assay. The assay was specific, as it did not amplify genomes from five other Mycobacterium spp. or five pathogenic enteric bacteria. Six hundred-twelve clinical samples (320 fecal and 292 serum) were assessed by RPA-LFD, qPCR, and enzyme-linked immunosorbent assay, respectively. The RPA-LFD assay yielded 100% sensitivity, 97.63% specificity, and 98.44% concordance rate with the qPCR results. This is the first report utilizing an RPA-LFD assay to visualize and rapidly detect M. paratuberculosis. Our results show this assay should be a useful method for the diagnosis of paratuberculosis in resource-constrained settings. PMID:29284204

Multiplex PCR for rapid diagnosis and differentiation of pox and pox-like diseases in dromedary Camels.

PubMed

Khalafalla, Abdelmalik I; Al-Busada, Khalid A; El-Sabagh, Ibrahim M

2015-07-07

Pox and pox-like diseases of camels are a group of exanthematous skin conditions that have become increasingly important economically. Three distinct viruses may cause them: camelpox virus (CMLV), camel parapox virus (CPPV) and camelus dromedary papilloma virus (CdPV). These diseases are often difficult to differentiate based on clinical presentation in disease outbreaks. Molecular methods such as PCR targeting species-specific genes have been developed and used to identify these diseases, but not simultaneously in a single tube. Recently, multiplex PCR has gained reputation as a convenient diagnostic method with cost-and timesaving benefits. In the present communication, we describe the development, optimization and validation of a multiplex PCR assay able to detect simultaneously the genome of the three viruses in one single test allowing for rapid and efficient molecular diagnosis. The assay was developed based on the evaluation and combination of published and new primer sets and was validated with viral genomic DNA extracted from known virus strains (n = 14) and DNA extracted from homogenized clinical skin specimens (n = 86). The assay detects correctly the target pathogens by amplification of targeted genes, even in case of co-infection. The method showed high sensitivity, and the specificity was confirmed by PCR-product sequencing. This assay provide rapid, sensitive and specific method for identifying three important viruses in specimens collected from dromedary camels with varying clinical presentations.

Evaluation of the IS6110 PCR assay for the rapid diagnosis of tuberculous meningitis

PubMed Central

Deshpande, Poonam S; Kashyap, Rajpal S; Ramteke, Sonali S; Nagdev, Khushboo J; Purohit, Hemant J; Taori, Girdhar M; Daginawala, Hatim F

2007-01-01

Background Tuberculous meningitis (TBM) is one of the common clinical manifestations of extra-pulmonary tuberculosis. It is difficult to diagnose due to a lack of rapid, sensitive, and specific tests. Newer methods, which are easy and reliable, are required to diagnose TBM at an early stage. Thus our aim was to evaluate the polymerase chain reaction (PCR) technique, using primers directed against the IS6110 gene, for the detection of Mycobacterium tuberculosis in the CSF, for the diagnosis of TBM patients. Methods An in-house IS6110 PCR method using a specific pair of primers designed to amplify the insertion sequence, IS6110, in the M. tuberculosis genome was used to analyze CSF. A total of 80 CSF samples from different groups of patients were studied (confirmed TBM n = 35, clinically suspected TBM n = 16, non-TBM infectious meningitis n = 12, non infectious neurological diseases n = 17). Results PCR gave a sensitivity of 91.4% and specificity of 75.9% for the diagnosis of TBM in patients with TBM confirmed by culture. In 16 clinically diagnosed, but unconfirmed, TBM cases PCR was positive in 10 (62.5%) cases. There were seven (24.1%) PCR-positive cases among the 29 patients with non-TBM and non-infectious neurological disease. Conclusion We conclude that the performance of an in-house IS6110 PCR assay is valuable in the rapid diagnosis of tuberculous meningitis. PMID:17976247

Rapid visual detection of Mycobacterium avium subsp. paratuberculosis by recombinase polymerase amplification combined with a lateral flow dipstick.

PubMed

Zhao, Guimin; Wang, Hongmei; Hou, Peili; He, Chengqiang; He, Hongbin

2018-03-31

Paratuberculosis (Johne's disease) is a chronic debilitating disease of domestic and wild ruminants. However, widespread point-of-care testing is infrequent due to the lack of a robust method. The isothermal recombinase polymerase amplification (RPA) technique has applied for rapid diagnosis. Herein, RPA combined with a lateral flow dipstick (LFD) assay was developed to estimate DNA from Mycobacterium avium subsp. paratuberculosis . First, analytical specificity and sensitivity of the RPA-nfo primer and probe sets were assessed. The assay successfully detected M. paratuberculosis DNA in 30 min at 39°C with a detection limit of up to eight copies per reaction, which was equivalent to that of the real-time quantitative polymerase chain reaction (qPCR) assay. The assay was specific, as it did not amplify genomes from five other Mycobacterium spp. or five pathogenic enteric bacteria. Six hundred-twelve clinical samples (320 fecal and 292 serum) were assessed by RPA-LFD, qPCR, and enzyme-linked immunosorbent assay, respectively. The RPA-LFD assay yielded 100% sensitivity, 97.63% specificity, and 98.44% concordance rate with the qPCR results. This is the first report utilizing an RPA-LFD assay to visualize and rapidly detect M. paratuberculosis . Our results show this assay should be a useful method for the diagnosis of paratuberculosis in resource-constrained settings.

A sensitive epitope-blocking ELISA for the detection of Chikungunya virus-specific antibodies in patients.

PubMed

Goh, Lucas Y H; Kam, Yiu-Wing; Metz, Stefan W; Hobson-Peters, Jody; Prow, Natalie A; McCarthy, Suzi; Smith, David W; Pijlman, Gorben P; Ng, Lisa F P; Hall, Roy A

2015-09-15

Chikungunya fever (CHIKF) has re-emerged as an arboviral disease that mimics clinical symptoms of other diseases such as dengue, malaria, as well as other alphavirus-related illnesses leading to problems with definitive diagnosis of the infection. Herein we describe the development and evaluation of a sensitive epitope-blocking ELISA (EB-ELISA) capable of specifically detecting anti-chikungunya virus (CHIKV) antibodies in clinical samples. The assay uses a monoclonal antibody (mAb) that binds an epitope on the E2 protein of CHIKV and does not exhibit cross-reactivity to other related alphaviruses. We also demonstrated the use of recombinant CHIK virus-like particles (VLPs) as a safe alternative antigen to infectious virions in the assay. Based on testing of 60 serum samples from patients in the acute or convalescent phase of CHIKV infection, the EB-ELISA provided us with 100% sensitivity, and exhibited 98.5% specificity when Ross River virus (RRV)- or Barmah Forest virus (BFV)-immune serum samples were included. This assay meets the public health demands of a rapid, robust, sensitive and specific, yet simple assay for specifically diagnosing CHIK-infections in humans. Copyright © 2015. Published by Elsevier B.V.

Rapid ABO genotyping by high-speed droplet allele-specific PCR using crude samples.

PubMed

Taira, Chiaki; Matsuda, Kazuyuki; Takeichi, Naoya; Furukawa, Satomi; Sugano, Mitsutoshi; Uehara, Takeshi; Okumura, Nobuo; Honda, Takayuki

2018-01-01

ABO genotyping has common tools for personal identification of forensic and transplantation field. We developed a new method based on a droplet allele-specific PCR (droplet-AS-PCR) that enabled rapid PCR amplification. We attempted rapid ABO genotyping using crude DNA isolated from dried blood and buccal cells. We designed allele-specific primers for three SNPs (at nucleotides 261, 526, and 803) in exons 6 and 7 of the ABO gene. We pretreated dried blood and buccal cells with proteinase K, and obtained crude DNAs without DNA purification. Droplet-AS-PCR allowed specific amplification of the SNPs at the three loci using crude DNA, with results similar to those for DNA extracted from fresh peripheral blood. The sensitivity of the methods was 5%-10%. The genotyping of extracted DNA and crude DNA were completed within 8 and 9 minutes, respectively. The genotypes determined by the droplet-AS-PCR method were always consistent with those obtained by direct sequencing. The droplet-AS-PCR method enabled rapid and specific amplification of three SNPs of the ABO gene from crude DNA treated with proteinase K. ABO genotyping by the droplet-AS-PCR has the potential to be applied to various fields including a forensic medicine and transplantation medical care. © 2017 Wiley Periodicals, Inc.

Development and testing of a rapid diagnostic test for bubonic and pneumonic plague.

PubMed

Chanteau, Suzanne; Rahalison, Lila; Ralafiarisoa, Lalao; Foulon, Jeanine; Ratsitorahina, Mahery; Ratsifasoamanana, Lala; Carniel, Elisabeth; Nato, Farida

2003-01-18

Plague is often fatal without prompt and appropriate treatment. It affects mainly poor and remote populations. Late diagnosis is one of the major causes of human death and spread of the disease, since it limits the effectiveness of control measures. We aimed to develop and assess a rapid diagnostic test (RDT) for plague. We developed a test that used monoclonal antibodies to the F1 antigen of Yersinia pestis. Sensitivity and specificity were assessed with a range of bacterial cultures and clinical samples, and compared with findings from available ELISA and bacteriological tests for plague. Samples from patients thought to have plague were tested with the RDT in the laboratory and by health workers in 26 pilot sites in Madagascar. The RDT detected concentrations of F1 antigen as low as 0.5 ng/mL in up to 15 min, and had a shelf life of 21 days at 60 degrees C. Its sensitivity and specificity were both 100%. RDT detected 41.6% and 31% more positive clinical specimens than did bacteriological methods and ELISA, respectively. The agreement rate between tests done at remote centres and in the laboratory was 89.8%. With the combination of bacteriological methods and F1 ELISA as reference standard, the positive and negative predictive values of the RDT were 90.6% and 86.7%, respectively. Our RDT is a specific, sensitive, and reliable test that can easily be done by health workers at the patient's bedside, for the rapid diagnosis of pneumonic and bubonic plague. This test will be of key importance for the control of plague in endemic countries.

Field performance of clinical case definitions for influenza screening during the 2009 pandemic.

PubMed

Chen, Shey-Ying; Chen, Yee-Chun; Chiang, Wen-Chu; Kung, Hsiang-Chi; King, Chwan-Chuen; Lai, Mei-Shu; Chie, Wei-Chu; Chen, Shyr-Chyr; Chen, Wen-Jone; Chang, Shan-Chwen

2012-11-01

The aim of this study was to assess the performance of 3 different influenza-like illness (ILI) case definitions, adopted by the European (European-CDC), USA (USA-CDC), and Taiwan Centers for Disease Prevention and Control (Taiwan-CDC), as screening tools for influenza during the 2009 H1N1 pandemic. From August 15 to 30, 2009, all emergency department patients with clinical symptoms or at epidemiologic risk for influenza were enrolled in an observational cohort study. Influenza diagnosis was established by positive rapid influenza diagnostic test or virus isolation. Sensitivity, specificity, positive predictive value, and negative predictive value of the European-, USA, and Taiwan-CDC ILI case definitions for screening were determined. A total of 870 patients were screened during the study period. Rapid influenza diagnostic test was positive in 315 patients, 273 (85.6%) of whom had fever duration less than 72 hours. Virus isolation identified 4 more patients with influenza A initially negative by rapid influenza diagnostic test. The mean (SD) age of these 319 patients was 24.3 (18.1) years. Of the 870 screened patients, 670 (77.0%), 476 (54.7%), and 325 (37.4%) met the European-, USA-, and Taiwan-CDC ILI case definition, respectively. Screening sensitivity was 95%, 77.7%, and 57.7% and specificity was 33.4%, 58.6%, and 74.4%, respectively. Differences in sensitivity and specificity between any 2 of the 3 groups were statistically significant (P < .05). First-line physicians should recognize the advantage and limitation of different ILI case definitions in influenza screening, especially confronted by pandemic or highly pathogenic avian influenza in the future. Copyright © 2012 Elsevier Inc. All rights reserved.

Assessing the reliability of microscopy and rapid diagnostic tests in malaria diagnosis in areas with varying parasite density among older children and adult patients in Nigeria.

PubMed

Ayogu, E E; Ukwe, C V; Nna, E O

2016-01-01

Current malaria control strategies are based on early diagnosis and appropriate treatment of malaria cases. The study aimed at comparing the performance of blood film microscopy and rapid diagnostic test (RDT) in Plasmodium falciparum detection in patients ≥6 years of age. A total of 154 consecutive pyretic patients aged 6-62 years were enrolled, sampled, and tested for malaria using RDT (first response) and microscopy by Giemsa staining. Genomic DNA was extracted after saponin hemolysis and nested polymerase chain reaction (PCR) was used to detect Plasmodium falciparum. The endpoints were sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). Of the 154 patients, 80 (51.9%) had fever of ≥37.5°C. 106 (68.8%) were positive by First response® , 132 (85.7%) by microscopy, and 121 (78.6%) by PCR. The sensitivity, specificity, PPV, and NPV of first response compared to microscopic method were 82.2%, 100.0%, 100.0%, and 34.3%, respectively, while it was 75.4%, 75.0%, 95.3%, and 31.2%, respectively, when compared to PCR. The sensitivity, specificity, PPV, and NPV of the microscopic method compared to PCR were 92.3%, 50.0%, 90.91%, and 54.5%, respectively. There was a significant difference in the performance of RDT and film microscopy methods (P ≤ 0.05). Microscopy performed better and is more reliable than first response (RDT) in areas with low parasite density among patients ≥6 years of age. Rapid diagnostic tests could be useful in aareas with high parasite density as an alternative to smear microscopy.

Rapid and sensitive approach to simultaneous detection of genomes of hepatitis A, B, C, D and E viruses.

PubMed

Kodani, Maja; Mixson-Hayden, Tonya; Drobeniuc, Jan; Kamili, Saleem

2014-10-01

Five viruses have been etiologically associated with viral hepatitis. Nucleic acid testing (NAT) remains the gold standard for diagnosis of viremic stages of infection. NAT methodologies have been developed for all hepatitis viruses; however, a NAT-based assay that can simultaneously detect all five viruses is not available. We designed TaqMan card-based assays for detection of HAV RNA, HBV DNA, HCV RNA, HDV RNA and HEV RNA. The performances of individual assays were evaluated on TaqMan Array Cards (TAC) for detecting five viral genomes simultaneously. Sensitivity and specificity were determined by testing 329 NAT-tested clinical specimens. All NAT-positive samples for HCV (n = 32), HDV (n = 28) and HEV (n = 14) were also found positive in TAC (sensitivity, 100%). Forty-three of 46 HAV-NAT positive samples were also positive in TAC (sensitivity, 94%), while 36 of 39 HBV-NAT positive samples were positive (sensitivity, 92%). No false-positives were detected for HBV (n = 32), HCV (n = 36), HDV (n = 30), and HEV (n = 31) NAT-negative samples (specificity 100%), while 38 of 41 HAV-NAT negative samples were negative by TAC (specificity 93%). TAC assay was concordant with corresponding individual NATs for hepatitis A-E viral genomes and can be used for their detection simultaneously. The TAC assay has potential for use in hepatitis surveillance, for screening of donor specimens and in outbreak situations. Wider availability of TAC-ready assays may allow for customized assays, for improving acute jaundice surveillance and for other purposes for which there is need to identify multiple pathogens rapidly. Published by Elsevier B.V.

Development and performance evaluation of a novel immunofluorescence chromatographic assay for histidine-rich protein 2 of Plasmodium falciparum.

PubMed

Kang, Keren; Dzakah, Emmanuel E; Huang, Yongping; Xie, Mingquan; Luo, Xiaochun; Li, Wenmei; Wang, Jihua

2015-05-30

The low sensitivity and specificity of Plasmodium falciparum diagnostic tests pose a serious health threat to people living in endemic areas. The objective of the study was to develop a rapid assay for the detection of histidine-rich protein 2 (HRP2) of P. falciparum in whole blood by immunofluorescence chromatographic technology. A total of 1163 positive and negative blood samples were screened. The double-antibody sandwich assay was used to establish the kit and its performance was evaluated for sensitivity, specificity, accuracy, precision, stability, and clinical effectiveness. The cut-off level of detection of the kit was 25 parasites/μl. Common interfering substances in human blood specimens, such as bilirubin, triglyceride and cholesterol had no significant effect on HRP2 antigen detection. The precision of the kit was run with different concentration of standard calibrators and the values were less than 10 %. The performance of this diagnostic kit in the detection of the calibrators has shown that a shelf life of about 12 months gives a more reliable result. Among clinical samples tested, the HRP2 test kit and the reference products had good coincidence rate in a parallel experiment and this test kit had a more sensitive detecting level to the target protein than the reference kits used in this study. The specificity and sensitivity for this test were 99.6 % (800/803) and 99.7 % (1160/1163), respectively. A novel HRP2 immunofluorescence detection method was developed in this study. Overall performance evaluation indicated that the kit has a rapid, high sensitivity and on-spot method for detecting P. falciparum.

Streptococcus A in paediatric accident and emergency: are rapid streptococcal tests and clinical examination of any help?

PubMed

Van Limbergen, J; Kalima, P; Taheri, S; Beattie, T F

2006-01-01

Rapid streptococcal tests (RSTs) for streptococcal pharyngitis have made diagnosis at once simpler and more complicated. The American Academy of Pediatrics recommends that all RSTs be confirmed by a follow up throat culture unless local validation has proved the RST to be equally sensitive. To evaluate (a) RST as a single diagnostic tool, compared with RST with or without throat culture; (b) clinical diagnosis and the relative contribution of different symptoms. The study included 213 patients with clinical signs of pharyngitis. Throat swabs were analysed using Quickvue+ Strep A Test; negative RSTs were backed up by throat culture. Thirteen clinical features commonly associated with strep throat were analysed using backward stepwise logistic regression. Positive results (RST or throat culture) were obtained in 33 patients; RST correctly identified 21. Eleven samples were false negative on RST. At a strep throat prevalence of 15.9%, sensitivity of RST was 65.6% (95% CI 46.8% to 81.4%) and specificity 99.4% (96.7% to 99.9%). Sensitivity of clinical diagnosis alone was 57% (34% to 78%) and specificity 71% (61% to 80%). Clinically, only history of sore throat, rash, and pyrexia contributed to the diagnosis of strep throat (p<0.05). The high specificity of RST facilitates early diagnosis of strep throat. However, the low sensitivity of RST does not support its use as a single diagnostic tool. The sensitivity in the present study is markedly different from that reported by the manufacturer. Clinical examination is of limited value in the diagnosis of strep throat. It is important to audit the performance of new diagnostic tests, previously validated in different settings.

Streptococcus A in paediatric accident and emergency: are rapid streptococcal tests and clinical examination of any help?

PubMed Central

Van Limbergen, J; Kalima, P; Taheri, S; Beattie, T F

2006-01-01

Background Rapid streptococcal tests (RSTs) for streptococcal pharyngitis have made diagnosis at once simpler and more complicated. The American Academy of Pediatrics recommends that all RSTs be confirmed by a follow up throat culture unless local validation has proved the RST to be equally sensitive. Aims To evaluate (a) RST as a single diagnostic tool, compared with RST with or without throat culture; (b) clinical diagnosis and the relative contribution of different symptoms. Methods The study included 213 patients with clinical signs of pharyngitis. Throat swabs were analysed using Quickvue+ Strep A Test; negative RSTs were backed up by throat culture. Thirteen clinical features commonly associated with strep throat were analysed using backward stepwise logistic regression. Results Positive results (RST or throat culture) were obtained in 33 patients; RST correctly identified 21. Eleven samples were false negative on RST. At a strep throat prevalence of 15.9%, sensitivity of RST was 65.6% (95% CI 46.8% to 81.4%) and specificity 99.4% (96.7% to 99.9%). Sensitivity of clinical diagnosis alone was 57% (34% to 78%) and specificity 71% (61% to 80%). Clinically, only history of sore throat, rash, and pyrexia contributed to the diagnosis of strep throat (p<0.05). Conclusion The high specificity of RST facilitates early diagnosis of strep throat. However, the low sensitivity of RST does not support its use as a single diagnostic tool. The sensitivity in the present study is markedly different from that reported by the manufacturer. Clinical examination is of limited value in the diagnosis of strep throat. It is important to audit the performance of new diagnostic tests, previously validated in different settings. PMID:16373800

Evaluation of the OnSite (Pf/Pan) rapid diagnostic test for diagnosis of clinical malaria.

PubMed

Mohon, Abu Naser; Elahi, Rubayet; Podder, Milka Patracia; Mohiuddin, Khaja; Hossain, Mohammad Sharif; Khan, Wasif A; Haque, Rashidul; Alam, Mohammad Shafiul

2012-12-12

Accurate diagnosis of malaria is an essential prerequisite for proper treatment and drug resistance monitoring. Microscopy is considered the gold standard for malaria diagnosis but has limitations. ELISA, PCR, and Real Time PCR are also used to diagnose malaria in reference laboratories, although their application at the field level is currently not feasible. Rapid diagnostic tests (RDTs) however, have been brought into field operation and widely adopted in recent days. This study evaluates OnSite (Pf/Pan) antigen test, a new RDT introduced by CTK Biotech Inc, USA for malaria diagnosis in a reference setting. Blood samples were collected from febrile patients referred for malaria diagnosis by clinicians. Subjects were included in this study from two different Upazila Health Complexes (UHCs) situated in two malaria endemic districts of Bangladesh. Microscopy and nested PCR were considered the gold standard in this study. OnSite (Pf/Pan) RDT was performed on preserved whole blood samples. In total, 372 febrile subjects were included in this study. Of these subjects, 229 (61.6%) tested positive for Plasmodium infection detected by microscopy and nested PCR. OnSite (Pf/Pan) RDT was 94.2% sensitive (95% CI, 89.3-97.3) and 99.5% specific (95% CI, 97.4-00.0) for Plasmodium falciparum diagnosis and 97.3% sensitive (95% CI, 90.5-99.7) and 98.7% specific (95% CI, 96.6-99.6) for Plasmodium vivax diagnosis. Sensitivity varied with differential parasite count for both P. falciparum and P. vivax. The highest sensitivity was observed in febrile patients with parasitaemia that ranged from 501-1,000 parasites/μL regardless of the Plasmodium species. The new OnSite (Pf/Pan) RDT is both sensitive and specific for symptomatic malaria diagnosis in standard laboratory conditions.

A rapid and sensitive dot-blot hybridization assay for the detection of citrus exocortis viroid in Citrus medica with digoxigenin-labelled RNA probes.

PubMed

Fonseca, M E; Marcellino, L H; Gander, E

1996-04-05

A rapid and sensitive dot-blot hybridization assay using in vitro-transcribed digoxigenin-labelled RNA probes (riboprobes) was developed aiming at detection of citrus exocortis viroid (CEVd) in crude sap of infected Citrus medica plants. The protocol includes a very quick and simple preparation of RNA extracts from samples using a denaturation step with formaldehyde. From our results, the employment of this step is highly recommended because the hybridization signals in formaldehyde-denatured samples were significantly stronger when compared with that of extracts without formaldehyde treatment. The assay was found to be sensitive enough to detect 0.1 ng of purified CEVd RNA and was able to detect viroid in 0.2 mg of symptomatic Citrus medica leaves. The use of riboprobes also allowed hybridization under high temperature conditions, avoiding non-specific background.

Diagnostic accuracy of tests to detect Hepatitis C antibody: a meta-analysis and review of the literature.

PubMed

Tang, Weiming; Chen, Wen; Amini, Ali; Boeras, Debi; Falconer, Jane; Kelly, Helen; Peeling, Rosanna; Varsaneux, Olivia; Tucker, Joseph D; Easterbrook, Philippa

2017-11-01

Although direct-acting antivirals can achieve sustained virological response rates greater than 90% in Hepatitis C Virus (HCV) infected persons, at present the majority of HCV-infected individuals remain undiagnosed and therefore untreated. While there are a wide range of HCV serological tests available, there is a lack of formal assessment of their diagnostic performance. We undertook a systematic review and meta-analysis to evaluate he diagnostic accuracy of available rapid diagnostic tests (RDT) and laboratory based EIA assays in detecting antibodies to HCV. We used the PRISMA checklist and Cochrane guidance to develop our search protocol. The search strategy was registered in PROSPERO (CRD42015023567). The search focused on hepatitis C, diagnostic tests, and diagnostic accuracy within eight databases (MEDLINE, EMBASE, the Cochrane Central Register of Controlled Trials, Science Citation Index Expanded, Conference Proceedings Citation Index-Science, SCOPUS, Literatura Latino-Americana e do Caribe em Ciências da Saúde and WHO Global Index Medicus. Studies were included if they evaluated an assay to determine the sensitivity and specificity of HCV antibody (HCV Ab) in humans. Two reviewers independently extracted data and performed a quality assessment of the studies using the QUADAS tool. We pooled test estimates using the DerSimonian-Laird method, by using the software R and RevMan. 5.3. A total of 52 studies were identified that included 52,673 unique test measurements. Based on five studies, the pooled sensitivity and specificity of HCV Ab rapid diagnostic tests (RDTs) were 98% (95% CI 98-100%) and 100% (95% CI 100-100%) compared to an enzyme immunoassay (EIA) reference standard. High HCV Ab RDTs sensitivity and specificity were observed across screening populations (general population, high risk populations, and hospital patients) using different reference standards (EIA, nucleic acid testing, immunoblot). There were insufficient studies to undertake subanalyses based on HIV co-infection. Oral HCV Ab RDTs also had excellent sensitivity and specificity compared to blood reference tests, respectively at 94% (95% CI 93-96%) and 100% (95% CI 100-100%). Among studies that assessed individual oral RDTs, the eight studies revealed that OraQuick ADVANCE® had a slightly higher sensitivity (98%, 95% CI 97-98%) compared to the other oral brands (pooled sensitivity: 88%, 95% CI 84-92%). RDTs, including oral tests, have excellent sensitivity and specificity compared to laboratory-based methods for HCV antibody detection across a wide range of settings. Oral HCV Ab RDTs had good sensitivity and specificity compared to blood reference standards.

Preservation of Specific Protein Signaling States Using Heat Based Stabilizor System.

PubMed

Borén, Mats

2017-01-01

The ability to adequately measure the phosphorylation state of a protein has major biological as well as clinical relevance. Due to its variable nature, reversible protein phosphorylations are sensitive to changes in the tissue environment. Stabilizor TM T1 is a system for rapid inactivation of enzymatic activity in biological samples. Enzyme inactivation is accomplished using thermal denaturation in a rapid, homogeneous, and reproducible fashion without the need of added chemical inhibitors. Using pCREB(Ser133) as a model system, the applicability of the Stabilizor system to preserve a rapidly lost phosphorylation is shown.

Performance Assessment of the CapitalBio Mycobacterium Identification Array System for Identification of Mycobacteria

PubMed Central

Liu, Jingbo; Yan, Zihe; Han, Min; Han, Zhijun; Jin, Lingjie; Zhao, Yanlin

2012-01-01

The CapitalBio Mycobacterium identification microarray system is a rapid system for the detection of Mycobacterium tuberculosis. The performance of this system was assessed with 24 reference strains, 486 Mycobacterium tuberculosis clinical isolates, and 40 clinical samples and then compared to the “gold standard” of DNA sequencing. The CapitalBio Mycobacterium identification microarray system showed highly concordant identification results of 100% and 98.4% for Mycobacterium tuberculosis complex (MTC) and nontuberculous mycobacteria (NTM), respectively. The sensitivity and specificity of the CapitalBio Mycobacterium identification array for identification of Mycobacterium tuberculosis isolates were 99.6% and 100%, respectively, for direct detection and identification of clinical samples, and the overall sensitivity was 52.5%. It was 100% for sputum, 16.7% for pleural fluid, and 10% for bronchoalveolar lavage fluid, respectively. The total assay was completed in 6 h, including DNA extraction, PCR, and hybridization. The results of this study confirm the utility of this system for the rapid identification of mycobacteria and suggest that the CapitalBio Mycobacterium identification array is a molecular diagnostic technique with high sensitivity and specificity that has the capacity to quickly identify most mycobacteria. PMID:22090408

Recombinase polymerase and enzyme-linked immunosorbent assay as a DNA amplification-detection strategy for food analysis.

PubMed

Santiago-Felipe, S; Tortajada-Genaro, L A; Puchades, R; Maquieira, A

2014-02-06

Polymerase chain reaction in conjunction with enzyme-linked immunosorbent assay (PCR-ELISA) is a well-established technique that provides a suitable rapid, sensitive, and selective method for a broad range of applications. However, the need for precise rapid temperature cycling of PCR is an important drawback that can be overcome by employing isothermal amplification reactions such as recombinase polymerase amplification (RPA). The RPA-ELISA combination is proposed for amplification at a low, constant temperature (40°C) in a short time (40 min), for the hybridisation of labelled products to specific 5'-biotinylated probes/streptavidin in coated microtiter plates at room temperature, and for detection by colorimetric immunoassay. RPA-ELISA was applied to screen common safety threats in foodstuffs, such as allergens (hazelnut, peanut, soybean, tomato, and maize), genetically modified organisms (P35S and TNOS), pathogenic bacteria (Salmonella sp. and Cronobacter sp.), and fungi (Fusarium sp.). Satisfactory sensitivity and reproducibility results were achieved for all the targets. The RPA-ELISA technique does away with thermocycling and provides a suitable sensitive, specific, and cost-effective method for routine applications, and proves particularly useful for resource-limited settings. Copyright © 2013 Elsevier B.V. All rights reserved.

Development of a serotype colloidal gold strip using monoclonal antibody for rapid detection type Asia1 foot-and-mouth disease

PubMed Central

2011-01-01

Background In this study, we developed a rapid, one step colloid gold strip (CGS) capable of specifically detecting type Asia1 foot-and-mouth disease virus (FMDV). We have produced two monoclonal antibodies (mAb) to type Asia1 FMD (named 1B8 and 5E2). On the test strip, the purified 1B8 labelled with the colloidal gold was used as the detector, and the purified 5E2 and goat anti-mouse antibodies were wrapped onto nitrocellulose (NC) membranes as the test and the control line, respectively. The rapid colloidal gold stereotype diagnostic strip was housed in a plastic case. Results In specificity and sensitivity assay, there was no cross-reaction of the antigen with the other type of FMD and SVDV. The detection sensitivity was found to be as high as 10-5 dilution of Asia1/JSL/05 (1 × 107.2TCID50/50 μL). There was excellent agreement between the results obtained by CGS and reverse indirect hemagglutination assay (RIHA), and the agreement can reach to 98.75%. Conclusion We developed colloidal gold strips that have good qualities and does not require specialized equipment or technicians. This method provided a feasible, convenient, rapid, and effective for detecting type Asia1 FMDV in the fields. PMID:21880157

Simultaneous point-of-care detection of anemia and sickle cell disease in Tanzania: the RAPID study.

PubMed

Smart, Luke R; Ambrose, Emmanuela E; Raphael, Kevin C; Hokororo, Adolfine; Kamugisha, Erasmus; Tyburski, Erika A; Lam, Wilbur A; Ware, Russell E; McGann, Patrick T

2018-02-01

Both anemia and sickle cell disease (SCD) are highly prevalent across sub-Saharan Africa, and limited resources exist to diagnose these conditions quickly and accurately. The development of simple, inexpensive, and accurate point-of-care (POC) assays represents an important advance for global hematology, one that could facilitate timely and life-saving medical interventions. In this prospective study, Robust Assays for Point-of-care Identification of Disease (RAPID), we simultaneously evaluated a POC immunoassay (Sickle SCAN™) to diagnose SCD and a first-generation POC color-based assay to detect anemia. Performed at Bugando Medical Center in Mwanza, Tanzania, RAPID tested 752 participants (age 1 day to 20 years) in four busy clinical locations. With minimally trained medical staff, the SCD POC assay diagnosed SCD with 98.1% sensitivity and 91.1% specificity. The hemoglobin POC assay had 83.2% sensitivity and 74.5% specificity for detection of severe anemia (Hb ≤ 7 g/dL). Interobserver agreement was excellent for both POC assays (r = 0.95-0.96). Results for the hemoglobin POC assay have informed the second-generation assay design to be more suitable for low-resource settings. RAPID provides practical feasibility data regarding two novel POC assays for the diagnosis of anemia and SCD in real-world field evaluations and documents the utility and potential impact of these POC assays for sub-Saharan Africa.

Development of a serotype colloidal gold strip using monoclonal antibody for rapid detection type Asia1 foot-and-mouth disease.

PubMed

Lin, Tong; Shao, Jun-jun; Du, Jun-zheng; Cong, Guo-zheng; Gao, Shan-dian; Chang, Huiyun

2011-09-01

In this study, we developed a rapid, one step colloid gold strip (CGS) capable of specifically detecting type Asia1 foot-and-mouth disease virus (FMDV). We have produced two monoclonal antibodies (mAb) to type Asia1 FMD (named 1B8 and 5E2). On the test strip, the purified 1B8 labelled with the colloidal gold was used as the detector, and the purified 5E2 and goat anti-mouse antibodies were wrapped onto nitrocellulose (NC) membranes as the test and the control line, respectively. The rapid colloidal gold stereotype diagnostic strip was housed in a plastic case. In specificity and sensitivity assay, there was no cross-reaction of the antigen with the other type of FMD and SVDV. The detection sensitivity was found to be as high as 10(-5) dilution of Asia1/JSL/05 (1 × 10(7.2)TCID(50)/50 μL). There was excellent agreement between the results obtained by CGS and reverse indirect hemagglutination assay (RIHA), and the agreement can reach to 98.75%. We developed colloidal gold strips that have good qualities and does not require specialized equipment or technicians. This method provided a feasible, convenient, rapid, and effective for detecting type Asia1 FMDV in the fields.

RNAi screen for rapid therapeutic target identification in leukemia patients

PubMed Central

Tyner, Jeffrey W.; Deininger, Michael W.; Loriaux, Marc M.; Chang, Bill H.; Gotlib, Jason R.; Willis, Stephanie G.; Erickson, Heidi; Kovacsovics, Tibor; O'Hare, Thomas; Heinrich, Michael C.; Druker, Brian J.

2009-01-01

Targeted therapy has vastly improved outcomes in certain types of cancer. Extension of this paradigm across a broad spectrum of malignancies will require an efficient method to determine the molecular vulnerabilities of cancerous cells. Improvements in sequencing technology will soon enable high-throughput sequencing of entire genomes of cancer patients; however, determining the relevance of identified sequence variants will require complementary functional analyses. Here, we report an RNAi-assisted protein target identification (RAPID) technology that individually assesses targeting of each member of the tyrosine kinase gene family. We demonstrate that RAPID screening of primary leukemia cells from 30 patients identifies targets that are critical to survival of the malignant cells from 10 of these individuals. We identify known, activating mutations in JAK2 and K-RAS, as well as patient-specific sensitivity to down-regulation of FLT1, CSF1R, PDGFR, ROR1, EPHA4/5, JAK1/3, LMTK3, LYN, FYN, PTK2B, and N-RAS. We also describe a previously undescribed, somatic, activating mutation in the thrombopoietin receptor that is sensitive to down-stream pharmacologic inhibition. Hence, the RAPID technique can quickly identify molecular vulnerabilities in malignant cells. Combination of this technique with whole-genome sequencing will represent an ideal tool for oncogenic target identification such that specific therapies can be matched with individual patients. PMID:19433805

The accuracy of HIV rapid testing in integrated bio-behavioral surveys of men who have sex with men across 5 Provinces in South Africa

PubMed Central

Kufa, Tendesayi; Lane, Tim; Manyuchi, Albert; Singh, Beverley; Isdahl, Zachary; Osmand, Thomas; Grasso, Mike; Struthers, Helen; McIntyre, James; Chipeta, Zawadi; Puren, Adrian

2017-01-01

Abstract We describe the accuracy of serial rapid HIV testing among men who have sex with men (MSM) in South Africa and discuss the implications for HIV testing and prevention. This was a cross-sectional survey conducted at five stand-alone facilities from five provinces. Demographic, behavioral, and clinical data were collected. Dried blood spots were obtained for HIV-related testing. Participants were offered rapid HIV testing using 2 rapid diagnostic tests (RDTs) in series. In the laboratory, reference HIV testing was conducted using a third-generation enzyme immunoassay (EIA) and a fourth-generation EIA as confirmatory. Accuracy, sensitivity, specificity, positive predictive value, negative predictive value, false-positive, and false-negative rates were determined. Between August 2015 and July 2016, 2503 participants were enrolled. Of these, 2343 were tested by RDT on site with a further 2137 (91.2%) having definitive results on both RDT and EIA. Sensitivity, specificity, positive predictive value, negative predictive value, false-positive rates, and false-negative rates were 92.6% [95% confidence interval (95% CI) 89.6–94.8], 99.4% (95% CI 98.9–99.7), 97.4% (95% CI 95.2–98.6), 98.3% (95% CI 97.6–98.8), 0.6% (95% CI 0.3–1.1), and 7.4% (95% CI 5.2–10.4), respectively. False negatives were similar to true positives with respect to virological profiles. Overall accuracy of the RDT algorithm was high, but sensitivity was lower than expected. Post-HIV test counseling should include discussions of possible false-negative results and the need for retesting among HIV negatives. PMID:28700474

[Microbiology--laboratory examinations for bacterias].

PubMed

Hen, Renjun; Imafuku, Yuji; Yoshida, Hiroshi

2002-11-01

As it has been required to identify pathogenic microbes in shorter times, simple and rapid methods have been developed and used. Here, we summarized the present situation of rapid diagnostic testing in clinical microbiology in Japan, and also presented our results on PBP2' detection. The rapid test kits available in Japan for E. coli, Helicobacter pylori, Salmonella, Streptococcus and Staphylococcus aureus were described. Rapid examination methods are based mainly on immunologic reactions, which included slide agglutination using latex particle, immunochromatography and ELISA. Times required for the identification are 10 to 15 minutes. Moreover, rapid test kits employing PCR are also marketed. Further, we evaluated MRSA-LA "Seiken" which is a rapid detection kit for PBP2' produced by MRSA. The test was shown to be highly sensitive and specific. For the rapid identification of pathogenic microbes, simple and rapid test kits described here will be used more in clinical diagnosis.

Detection and prevention of mycoplasma hominis infection

DOEpatents

DelVecchio, Vito G.; Gallia, Gary L.; McCleskey, Ferne K.

1997-01-21

The present invention is directed to a rapid and sensitive method for detecting Mycoplasma hominis using M. hominis-specific probes, oligonucleotides or antibodies. In particular a target sequence can be amplified by in vitro nucleic acid amplification techniques, detected by nucleic acid hybridization using the subject probes and oligonucleotides or detected by immunoassay using M. hominis-specific antibodies. M. hominis-specific nucleic acids which do not recognize or hybridize to genomic nucleic acid of other Mycoplasma species are also provided.

Performance of rapid tests and algorithms for HIV screening in Abidjan, Ivory Coast.

PubMed

Loukou, Y G; Cabran, M A; Yessé, Zinzendorf Nanga; Adouko, B M O; Lathro, S J; Agbessi-Kouassi, K B T

2014-01-01

Seven rapid diagnosis tests (RDTs) of HIV were evaluated by a panel group who collected serum samples from patients in Abidjan (HIV-1 = 203, HIV-2 = 25, HIV-dual = 25, HIV = 305). Kit performances were recorded after the reference techniques (enzyme-linked immunosorbent assay). The following RDTs showed a sensitivity of 100% and a specificity higher than 99%: Determine, Oraquick, SD Bioline, BCP, and Stat-Pak. These kits were used to establish infection screening strategies. The combination with 2 or 3 of these tests in series or parallel algorithms showed that series combinations with 2 tests (Oraquick and Bioline) and 3 tests (Determine, BCP, and Stat-Pak) gave the best performances (sensitivity, specificity, positive predictive value, and negative predictive value of 100%). However, the combination with 2 tests appeared to be more onerous than the combination with 3 tests. The combination with Determine, BCP, and Stat-Pak tests serving as a tiebreaker could be an alternative to the HIV/AIDS serological screening in Abidjan.

RAPID DIAGNOSIS OF PNEUMOCOCCAL PNEUMONIA AMONG HIV-INFECTED ADULTS WITH URINE ANTIGEN DETECTION

PubMed Central

Boulware, David R; Daley, Charles L.; Merrifield, Cynthia; Hopewell, Philip C.; Janoff, Edward N.

2007-01-01

Objectives Streptococcus pneumoniae is the leading cause of bacterial pneumonia and associated bacteremia during HIV infection. Rapid diagnostic assays may limit inappropriate therapy. Methods Clinical signs and symptoms and sera and urine were collected prospectively from 70 adults with pneumococcal pneumonia, including 47 with HIV co-infection. Pneumococcal C-polysaccharide antigen was detected in urine using the Binax® immunochromatographic test (ICT). A systematic review of 24 published studies was conducted. Results Clinical symptoms, signs, and laboratory parameters except leukocytosis, were similar in HIV-infected and HIV-seronegative pneumonia. The performance of the urine antigen ICT was independent of HIV-status (sensitivity 81%, specificity 98%, positive (PPV) and negative predictive values (NPV) 98%, and 82%, respectively). The sensitivity of sputum Gram’s stain was 58% [34/59] with sputum unable to be provided by 16%. The CRP response was identical in HIV-infected (mean ± SD) 133 ± 88 vs. seronegative 135 ± 104 mg/L (p=0.9). In the systematic review, the ICT performance revealed 74% sensitivity (95% CI: 72% to 77%) and 94% specificity (95% CI: 93% to 95%). Urine antigen testing increases etiologic diagnosis by 23% (Range: 10% –59%) when testing adults with community acquired pneumonia of unknown etiology. Conclusions Urinary antigen detection provides a credible rapid diagnostic test for pneumococcal pneumonia regardless of HIV-status. CRP response to acute infection is similar in HIV co-infection and increases diagnostic certainty. PMID:17692384

Comparison of the diagnostic accuracy of a rapid immunochromatographic test and the rapid plasma reagin test for antenatal syphilis screening in Mozambique.

PubMed Central

Montoya, Pablo J.; Lukehart, Sheila A.; Brentlinger, Paula E.; Blanco, Ana J.; Floriano, Florencia; Sairosse, Josefa; Gloyd, Stephen

2006-01-01

OBJECTIVE: Programmes to control syphilis in developing countries are hampered by a lack of laboratory services, delayed diagnosis, and doubts about current screening methods. We aimed to compare the diagnostic accuracy of an immunochromatographic strip (ICS) test and the rapid plasma reagin (RPR) test with the combined gold standard (RPR, Treponema pallidum haemagglutination assay and direct immunofluorescence stain done at a reference laboratory) for the detection of syphilis in pregnancy. METHODS: We included test results from 4789 women attending their first antenatal visit at one of six health facilities in Sofala Province, central Mozambique. We compared diagnostic accuracy (sensitivity, specificity, and positive and negative predictive values) of ICS and RPR done at the health facilities and ICS performed at the reference laboratory. We also made subgroup comparisons by human immunodeficiency virus (HIV) and malaria status. FINDINGS: For active syphilis, the sensitivity of the ICS was 95.3% at the reference laboratory, and 84.1% at the health facility. The sensitivity of the RPR at the health facility was 70.7%. Specificity and positive and negative predictive values showed a similar pattern. The ICS outperformed RPR in all comparisons (P<0.001). CONCLUSION: The diagnostic accuracy of the ICS compared favourably with that of the gold standard. The use of the ICS in Mozambique and similar settings may improve the diagnosis of syphilis in health facilities, both with and without laboratories. PMID:16501726

Evaluation of the Clearview® Malaria pLDH Malaria Rapid Diagnostic Test in a non-endemic setting.

PubMed

Houzé, Sandrine; Hubert, Véronique; Cohen, Dorit Pessler; Rivetz, Baruch; Le Bras, Jacques

2011-09-27

Malaria Rapid Diagnostic Tests (RDTs) are widely used to diagnose malaria. The present study evaluated a new RDT, the Clearview® Malaria pLDH test targeting the pan-Plasmodium antigen lactate dehydrogenase (pLDH). The Clearview® Malaria pLDH test was evaluated on fresh samples obtained in returned international travellers using microscopy corrected by PCR as the reference method. Included samples were Plasmodium falciparum (139), Plasmodium vivax (22), Plasmodium ovale (20), Plasmodium malariae (7), and 102 negative. Overall sensitivity for the detection of Plasmodium spp was 93.2%. For P. falciparum, the sensitivity was 98.6%; for P. vivax, P. ovale and P. malariae, overall sensitivities were 90.9%, 60.0% and 85.7% respectively. For P. falciparum and for P. vivax, the sensitivities increased to 100% at parasite densities above 100/μl. The specificity was 100%. The test was easily to perform and the result was stable for at least 1 hour. The Clearview® Malaria pLDH was efficient for the diagnosis of malaria. The test was very sensitive for P. falciparum and P. vivax detection. The sensitivities for P. ovale and P. malariae were better than other RDTs.

Validity of Race, Ethnicity, and National Origin in Population-based Cancer Registries and Rapid Case Ascertainment Enhanced With a Spanish Surname List.

PubMed

Clarke, Lisa C; Rull, Rudolph P; Ayanian, John Z; Boer, Robert; Deapen, Dennis; West, Dee W; Kahn, Katherine L

2016-01-01

Accurate information regarding race, ethnicity, and national origins is critical for identifying disparities in the cancer burden. To examine the use of a Spanish surname list to improve the quality of race-related information obtained from rapid case ascertainment (RCA) and to estimate the accuracy of race-related information obtained from cancer registry records collected by routine reporting. Self-reported survey responses of 3954 participants from California enrolled in the Cancer Care Outcomes Research and Surveillance Consortium. Sensitivity, specificity, positive predictive value, and percent agreement. We used logistic regression to identify predictors of underreporting and overreporting of a race/ethnicity. Use of the Spanish surname list increased the sensitivity of RCA for Latino ethnicity from 37% to 83%. Sensitivity for cancer registry records collected by routine reporting was ≥95% for whites, blacks, and Asians, and specificity was high for all groups (86%-100%). However, patterns of misclassification by race/ethnicity were found that could lead to biased cancer statistics for specific race/ethnicities. Discordance between self-reported and registry-reported race/ethnicity was more likely for women, Latinos, and Asians. Methods to improve race and ethnicity data, such as using Spanish surnames in RCA and instituting data collection guidelines for hospitals, are needed to ensure minorities are accurately represented in clinical and epidemiological research.

Clostridium difficile infection diagnostics - evaluation of the C. DIFF Quik Chek Complete assay, a rapid enzyme immunoassay for detection of toxigenic C. difficile in clinical stool samples.

PubMed

Johansson, Karin; Karlsson, Hanna; Norén, Torbjörn

2016-11-01

Diagnostic testing for Clostridium difficile infection (CDI) has, in recent years, seen the introduction of rapid dual-EIA (enzyme immunoassay) tests combining species-specific glutamate dehydrogenase (GDH) with toxin A/B. In a prospective study, we compared the C. DIFF Quik Chek Complete test to a combination of selective culture (SC) and loop-mediated isothermal amplification (LAMP) of the toxin A gene. Of 419 specimens, 68 were positive in SC including 62 positive in LAMP (14.7%). The combined EIA yielded 82 GDH positives of which 47 were confirmed toxin A/B positive (11%) corresponding to a sensitivity and specificity of 94% for GDH EIA compared to SC and for toxin A/B EIA a sensitivity of 71% and a specificity of 99% compared to LAMP. Twenty different PCR ribotypes were evenly distributed except for UK 081 where only 25% were toxin A/B positive compared to LAMP. We propose a primary use of a combined GDH toxin A/B EIA permitting a sensitive 1-h result of 379 of 419 (90%, all negatives plus GDH and toxin EIA positives) referred specimens. The remaining 10% being GDH positive should be tested for toxin A/B gene on the same day and positive results left to a final decision by the physician. © 2016 APMIS. Published by John Wiley & Sons Ltd.

A Novel Malaria Pf/Pv Ab Rapid Diagnostic Test Using a Differential Diagnostic Marker Identified by Network Biology.

PubMed

Cho, Sung Jin; Lee, Jihoo; Lee, Hyun Jae; Jo, Hyun-Young; Sinniah, Mangalam; Kim, Hak-Yong; Chong, Chom-Kyu; Song, Hyun-Ok

2016-01-01

Rapid diagnostic tests (RDTs) can detect anti-malaria antibodies in human blood. As they can detect parasite infection at the low parasite density, they are useful in endemic areas where light infection and/or re-infection of parasites are common. Thus, malaria antibody tests can be used for screening bloods in blood banks to prevent transfusion-transmitted malaria (TTM), an emerging problem in malaria endemic areas. However, only a few malaria antibody tests are available in the microwell-based assay format and these are not suitable for field application. A novel malaria antibody (Ab)-based RDT using a differential diagnostic marker for falciparum and vivax malaria was developed as a suitable high-throughput assay that is sensitive and practical for blood screening. The marker, merozoite surface protein 1 (MSP1) was discovered by generation of a Plasmodium-specific network and the hierarchical organization of modularity in the network. Clinical evaluation revealed that the novel Malaria Pf/Pv Ab RDT shows improved sensitivity (98%) and specificity (99.7%) compared with the performance of a commercial kit, SD BioLine Malaria P.f/P.v (95.1% sensitivity and 99.1% specificity). The novel Malaria Pf/Pv Ab RDT has potential for use as a cost-effective blood-screening tool for malaria and in turn, reduces TTM risk in endemic areas.

A sensitive and innovative detection method for rapid C-reactive proteins analysis based on a micro-fluxgate sensor system

PubMed Central

Yang, Zhen; Zhi, Shaotao; Feng, Zhu; Lei, Chong; Zhou, Yong

2018-01-01

A sensitive and innovative assay system based on a micro-MEMS-fluxgate sensor and immunomagnetic beads-labels was developed for the rapid analysis of C-reactive proteins (CRP). The fluxgate sensor presented in this study was fabricated through standard micro-electro-mechanical system technology. A multi-loop magnetic core made of Fe-based amorphous ribbon was employed as the sensing element, and 3-D solenoid copper coils were used to control the sensing core. Antibody-conjugated immunomagnetic microbeads were strategically utilized as signal tags to label the CRP via the specific conjugation of CRP to polyclonal CRP antibodies. Separate Au film substrates were applied as immunoplatforms to immobilize CRP-beads labels through classical sandwich assays. Detection and quantification of the CRP at different concentrations were implemented by detecting the stray field of CRP labeled magnetic beads using the newly-developed micro-fluxgate sensor. The resulting system exhibited the required sensitivity, stability, reproducibility, and selectivity. A detection limit as low as 0.002 μg/mL CRP with a linearity range from 0.002 μg/mL to 10 μg/mL was achieved, and this suggested that the proposed biosystem possesses high sensitivity. In addition to the extremely low detection limit, the proposed method can be easily manipulated and possesses a quick response time. The response time of our sensor was less than 5 s, and the entire detection period for CRP analysis can be completed in less than 30 min using the current method. Given the detection performance and other advantages such as miniaturization, excellent stability and specificity, the proposed biosensor can be considered as a potential candidate for the rapid analysis of CRP, especially for point-of-care platforms. PMID:29601593

Evaluation of simple rapid HIV assays and development of national rapid HIV test algorithms in Dar es Salaam, Tanzania

PubMed Central

2009-01-01

Background Suitable algorithms based on a combination of two or more simple rapid HIV assays have been shown to have a diagnostic accuracy comparable to double enzyme-linked immunosorbent assay (ELISA) or double ELISA with Western Blot strategies. The aims of this study were to evaluate the performance of five simple rapid HIV assays using whole blood samples from HIV-infected patients, pregnant women, voluntary counseling and testing attendees and blood donors, and to formulate an alternative confirmatory strategy based on rapid HIV testing algorithms suitable for use in Tanzania. Methods Five rapid HIV assays: Determine™ HIV-1/2 (Inverness Medical), SD Bioline HIV 1/2 3.0 (Standard Diagnostics Inc.), First Response HIV Card 1–2.0 (PMC Medical India Pvt Ltd), HIV1/2 Stat-Pak Dipstick (Chembio Diagnostic System, Inc) and Uni-Gold™ HIV-1/2 (Trinity Biotech) were evaluated between June and September 2006 using 1433 whole blood samples from hospital patients, pregnant women, voluntary counseling and testing attendees and blood donors. All samples that were reactive on all or any of the five rapid assays and 10% of non-reactive samples were tested on a confirmatory Inno-Lia HIV I/II immunoblot assay (Immunogenetics). Results Three hundred and ninety samples were confirmed HIV-1 antibody positive, while 1043 were HIV negative. The sensitivity at initial testing of Determine, SD Bioline and Uni-Gold™ was 100% (95% CI; 99.1–100) while First Response and Stat-Pak had sensitivity of 99.5% (95% CI; 98.2–99.9) and 97.7% (95% CI; 95.7–98.9), respectively, which increased to 100% (95% CI; 99.1–100) on repeat testing. The initial specificity of the Uni-Gold™ assay was 100% (95% CI; 99.6–100) while specificities were 99.6% (95% CI; 99–99.9), 99.4% (95% CI; 98.8–99.7), 99.6% (95% CI; 99–99.9) and 99.8% (95% CI; 99.3–99.9) for Determine, SD Bioline, First Response and Stat-Pak assays, respectively. There was no any sample which was concordantly false positive in Uni-Gold™, Determine and SD Bioline assays. Conclusion An alternative confirmatory HIV testing strategy based on initial testing on either SD Bioline or Determine assays followed by testing of reactive samples on the Determine or SD Bioline gave 100% sensitivity (95% CI; 99.1–100) and 100% specificity (95% CI; 96–99.1) with Uni-Gold™ as tiebreaker for discordant results. PMID:19226452

Evaluation of simple rapid HIV assays and development of national rapid HIV test algorithms in Dar es Salaam, Tanzania.

PubMed

Lyamuya, Eligius F; Aboud, Said; Urassa, Willy K; Sufi, Jaffer; Mbwana, Judica; Ndugulile, Faustin; Massambu, Charles

2009-02-18

Suitable algorithms based on a combination of two or more simple rapid HIV assays have been shown to have a diagnostic accuracy comparable to double enzyme-linked immunosorbent assay (ELISA) or double ELISA with Western Blot strategies. The aims of this study were to evaluate the performance of five simple rapid HIV assays using whole blood samples from HIV-infected patients, pregnant women, voluntary counseling and testing attendees and blood donors, and to formulate an alternative confirmatory strategy based on rapid HIV testing algorithms suitable for use in Tanzania. Five rapid HIV assays: Determine HIV-1/2 (Inverness Medical), SD Bioline HIV 1/2 3.0 (Standard Diagnostics Inc.), First Response HIV Card 1-2.0 (PMC Medical India Pvt Ltd), HIV1/2 Stat-Pak Dipstick (Chembio Diagnostic System, Inc) and Uni-Gold HIV-1/2 (Trinity Biotech) were evaluated between June and September 2006 using 1433 whole blood samples from hospital patients, pregnant women, voluntary counseling and testing attendees and blood donors. All samples that were reactive on all or any of the five rapid assays and 10% of non-reactive samples were tested on a confirmatory Inno-Lia HIV I/II immunoblot assay (Immunogenetics). Three hundred and ninety samples were confirmed HIV-1 antibody positive, while 1043 were HIV negative. The sensitivity at initial testing of Determine, SD Bioline and Uni-Gold was 100% (95% CI; 99.1-100) while First Response and Stat-Pak had sensitivity of 99.5% (95% CI; 98.2-99.9) and 97.7% (95% CI; 95.7-98.9), respectively, which increased to 100% (95% CI; 99.1-100) on repeat testing. The initial specificity of the Uni-Gold assay was 100% (95% CI; 99.6-100) while specificities were 99.6% (95% CI; 99-99.9), 99.4% (95% CI; 98.8-99.7), 99.6% (95% CI; 99-99.9) and 99.8% (95% CI; 99.3-99.9) for Determine, SD Bioline, First Response and Stat-Pak assays, respectively. There was no any sample which was concordantly false positive in Uni-Gold, Determine and SD Bioline assays. An alternative confirmatory HIV testing strategy based on initial testing on either SD Bioline or Determine assays followed by testing of reactive samples on the Determine or SD Bioline gave 100% sensitivity (95% CI; 99.1-100) and 100% specificity (95% CI; 96-99.1) with Uni-Gold as tiebreaker for discordant results.

Rapid direct methods for enumeration of specific, active bacteria in water and biofilms

NASA Technical Reports Server (NTRS)

McFeters, G. A.; Pyle, B. H.; Lisle, J. T.; Broadaway, S. C.

1999-01-01

Conventional methods for detecting indicator and pathogenic bacteria in water may underestimate the actual population due to sublethal environmental injury, inability of the target bacteria to take up nutrients and other physiological factors which reduce bacterial culturability. Rapid and direct methods are needed to more accurately detect and enumerate active bacteria. Such a methodological advance would provide greater sensitivity in assessing the microbiological safety of water and food. The principle goal of this presentation is to describe novel approaches we have formulated for the rapid and simultaneous detection of bacteria plus the determination of their physiological activity in water and other environmental samples. The present version of our method involves the concentration of organisms by membrane filtration or immunomagnetic separation and combines an intracellular fluorochrome (CTC) for assessment of respiratory activity plus fluorescent-labelled antibody detection of specific bacteria. This approach has also been successfully used to demonstrate spatial and temporal heterogeneities of physiological activities in biofilms when coupled with cryosectioning. Candidate physiological stains include those capable of determining respiratory activity, membrane potential, membrane integrity, growth rate and cellular enzymatic activities. Results obtained thus far indicate that immunomagnetic separation can provide a high degree of sensitivity in the recovery of seeded target bacteria (Escherichia coli O157:H7) in water and hamburger. The captured and stained target bacteria are then enumerated by either conventional fluorescence microscopy or ChemScan(R), a new instrument that is very sensitive and rapid. The ChemScan(R) laser scanning instrument (Chemunex, Paris, France) provides the detection of individual fluorescently labelled bacterial cells using three emission channels in less than 5 min. A high degree of correlation has been demonstrated between results obtained with the ChemScan and traditional plate counts of mixed natural bacterial populations in water. The continuing evolution of these methods will be valuable in the rapid and accurate analysis of environmental samples.

Performance characteristics of an automated latex immunoturbidimetric assay [HemosIL® HIT-Ab(PF4-H)] for the diagnosis of immune heparin-induced thrombocytopenia.

PubMed

Warkentin, Theodore E; Sheppard, Jo-Ann I; Linkins, Lori-Ann; Arnold, Donald M; Nazy, Ishac

2017-05-01

Heparin-induced thrombocytopenia (HIT) is a prothrombotic drug reaction caused by platelet-activating anti-PF4/heparin antibodies. Given time-sensitive treatment considerations, a rapid and accurate laboratory test for HIT antibodies is needed. To determine operating characteristics for the HemosIL ® HIT-Ab (PF4/H) , a rapid, on-demand, fully-automated, latex immunoturbidimetric assay (LIA), for diagnosis of HIT. We evaluated LIA sensitivity, specificity, negative (NPV) and positive predictive value (PPV), negative (LR-) and positive likelihood ratio (LR+), using citrated-plasma from 429 patients (prospective cohort study of 4Ts scoring; HIT, n=31), and from consecutive HIT patients (n=125), using reference standard serotonin-release assay (SRA). Comparators included two PF4-dependent enzyme-immunoassays (EIAs). We used stratum-specific likelihood ratios (SSLRs) to determine how differing magnitudes of LIA-positivity influenced post-test probability of HIT. LIA operating characteristics were: sensitivity=97.4% (152/156); specificity=94.0% (374/398); PPV=55.6% (30/54); and NPV=99.7% (374/375). At manufacturers' cutoffs, LIA specificity and PPV were superior to the EIAs. Although a negative LIA pointed strongly against HIT (LR-, 0.034), the post-test probability was ~2% with high 4Ts score. The LIA's LR+ was high (16.0), with SSLRs rising substantially with greater LIA-positivity: 5.7 (1.0-4.9U/mL), 31 (5.0-15.9U/mL), and 128 (≥16U/mL). A LIA-positive result (at 1.0 cutoff) indicated at least 24% HIT probability (low 4Ts score), rising to 90% with high 4Ts score. Although approximately 1 in 40 SRA-positive patients tested LIA-negative, the LIA's high NPV and PPV indicate that this rapid assay is useful for the diagnostic evaluation of HIT, including in low pre-test situations. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

The Rapid-Heat LAMPellet Method: A Potential Diagnostic Method for Human Urogenital Schistosomiasis

PubMed Central

Carranza-Rodríguez, Cristina; Pérez-Arellano, José Luis; Vicente, Belén; López-Abán, Julio; Muro, Antonio

2015-01-01

Background Urogenital schistosomiasis due to Schistosoma haematobium is a serious underestimated public health problem affecting 112 million people - particularly in sub-Saharan Africa. Microscopic examination of urine samples to detect parasite eggs still remains as definitive diagnosis. This work was focussed on developing a novel loop-mediated isothermal amplification (LAMP) assay for detection of S. haematobium DNA in human urine samples as a high-throughput, simple, accurate and affordable diagnostic tool to use in diagnosis of urogenital schistosomiasis. Methodology/Principal Findings A LAMP assay targeting a species specific sequence of S. haematobium ribosomal intergenic spacer was designed. The effectiveness of our LAMP was assessed in a number of patients´ urine samples with microscopy confirmed S. haematobium infection. For potentially large-scale application in field conditions, different DNA extraction methods, including a commercial kit, a modified NaOH extraction method and a rapid heating method were tested using small volumes of urine fractions (whole urine, supernatants and pellets). The heating of pellets from clinical samples was the most efficient method to obtain good-quality DNA detectable by LAMP. The detection limit of our LAMP was 1 fg/µL of S. haematobium DNA in urine samples. When testing all patients´ urine samples included in our study, diagnostic parameters for sensitivity and specificity were calculated for LAMP assay, 100% sensitivity (95% CI: 81.32%-100%) and 86.67% specificity (95% CI: 75.40%-94.05%), and also for microscopy detection of eggs in urine samples, 69.23% sensitivity (95% CI: 48.21% -85.63%) and 100% specificity (95% CI: 93.08%-100%). Conclusions/Significance We have developed and evaluated, for the first time, a LAMP assay for detection of S. haematobium DNA in heated pellets from patients´ urine samples using no complicated requirement procedure for DNA extraction. The procedure has been named the Rapid-Heat LAMPellet method and has the potential to be developed further as a field diagnostic tool for use in urogenital schistosomiasis-endemic areas. PMID:26230990

Performance of C-reactive protein and procalcitonin to distinguish viral from bacterial and malarial causes of fever in Southeast Asia.

PubMed

Lubell, Yoel; Blacksell, Stuart D; Dunachie, Susanna; Tanganuchitcharnchai, Ampai; Althaus, Thomas; Watthanaworawit, Wanitda; Paris, Daniel H; Mayxay, Mayfong; Peto, Thomas J; Dondorp, Arjen M; White, Nicholas J; Day, Nicholas P J; Nosten, François; Newton, Paul N; Turner, Paul

2015-11-11

Poor targeting of antimicrobial drugs contributes to the millions of deaths each year from malaria, pneumonia, and other tropical infectious diseases. While malaria rapid diagnostic tests have improved use of antimalarial drugs, there are no similar tests to guide the use of antibiotics in undifferentiated fevers. In this study we estimate the diagnostic accuracy of two well established biomarkers of bacterial infection, procalcitonin and C-reactive protein (CRP) in discriminating between common viral and bacterial infections in malaria endemic settings of Southeast Asia. Serum procalcitonin and CRP levels were measured in stored serum samples from febrile patients enrolled in three prospective studies conducted in Cambodia, Laos and, Thailand. Of the 1372 patients with a microbiologically confirmed diagnosis, 1105 had a single viral, bacterial or malarial infection. Procalcitonin and CRP levels were compared amongst these aetiological groups and their sensitivity and specificity in distinguishing bacterial infections and bacteraemias from viral infections were estimated using standard thresholds. Serum concentrations of both biomarkers were significantly higher in bacterial infections and malaria than in viral infections. The AUROC for CRP in discriminating between bacterial and viral infections was 0.83 (0.81-0.86) compared with 0.74 (0.71-0.77) for procalcitonin (p < 0.0001). This relative advantage was evident in all sites and when stratifying patients by age and admission status. For CRP at a threshold of 10 mg/L, the sensitivity of detecting bacterial infections was 95% with a specificity of 49%. At a threshold of 20 mg/L sensitivity was 86% with a specificity of 67%. For procalcitonin at a low threshold of 0.1 ng/mL the sensitivity was 90% with a specificity of 39%. At a higher threshold of 0.5 ng/ul sensitivity was 60% with a specificity of 76%. In samples from febrile patients with mono-infections from rural settings in Southeast Asia, CRP was a highly sensitive and moderately specific biomarker for discriminating between viral and bacterial infections. Use of a CRP rapid test in peripheral health settings could potentially be a simple and affordable measure to better identify patients in need of antibacterial treatment and part of a global strategy to combat the emergence of antibiotic resistance.

PCR-Restriction Fragment Length Polymorphism for Rapid, Low-Cost Identification of Isoniazid-Resistant Mycobacterium tuberculosis▿

PubMed Central

Caws, Maxine; Tho, Dau Quang; Duy, Phan Minh; Lan, Nguyen Thi Ngoc; Hoa, Dai Viet; Torok, Mili Estee; Chau, Tran Thi Hong; Van Vinh Chau, Nguyen; Chinh, Nguyen Tran; Farrar, Jeremy

2007-01-01

PCR-restriction fragment length poymorphism (PCR-RFLP) is a simple, robust technique for the rapid identification of isoniazid-resistant Mycobacterium tuberculosis. One hundred consecutive isolates from a Vietnamese tuberculosis hospital were tested by MspA1I PCR-RFLP for the detection of isoniazid-resistant katG_315 mutants. The test had a sensitivity of 80% and a specificity of 100% against conventional phenotypic drug susceptibility testing. The positive and negative predictive values were 1 and 0.86, respectively. None of the discrepant isolates had mutant katG_315 codons by sequencing. The test is cheap (less than $1.50 per test), specific, and suitable for the rapid identification of isoniazid resistance in regions with a high prevalence of katG_315 mutants among isoniazid-resistant M. tuberculosis isolates. PMID:17428939

PCR/LDR/universal array platforms for the diagnosis of infectious disease.

PubMed

Pingle, Maneesh; Rundell, Mark; Das, Sanchita; Golightly, Linnie M; Barany, Francis

2010-01-01

Infectious diseases account for between 14 and 17 million deaths worldwide each year. Accurate and rapid diagnosis of bacterial, fungal, viral, and parasitic infections is therefore essential to reduce the morbidity and mortality associated with these diseases. Classical microbiological and serological methods have long served as the gold standard for diagnosis but are increasingly being replaced by molecular diagnostic methods that demonstrate increased sensitivity and specificity and provide an identification of the etiologic agent in a shorter period of time. PCR/LDR coupled with universal array detection provides a highly sensitive and specific platform for the detection and identification of bacterial and viral infections.

PCR/LDR/Universal Array Platforms for the Diagnosis of Infectious Disease

PubMed Central

Pingle, Maneesh; Rundell, Mark; Das, Sanchita; Golightly, Linnie M.; Barany, Francis

2015-01-01

Infectious diseases account for between 14 and 17 million deaths worldwide each year. Accurate and rapid diagnosis of bacterial, fungal, viral, and parasitic infections is therefore essential to reduce the morbidity and mortality associated with these diseases. Classical microbiological and serological methods have long served as the gold standard for diagnosis but are increasingly being replaced by molecular diagnostic methods that demonstrate increased sensitivity and specificity and provide an identification of the etiologic agent in a shorter period of time. PCR/LDR coupled with universal array detection provides a highly sensitive and specific platform for the detection and identification of bacterial and viral infections. PMID:20217576

Simple and rapid method for the detection of Filobasidiella neoformans in a probiotic dairy product by using loop-mediated isothermal amplification.

PubMed

Ishikawa, Hiroshi; Kasahara, Kohei; Sato, Sumie; Shimakawa, Yasuhisa; Watanabe, Koichi

2014-05-16

Yeast contamination is a serious problem in the food industry and a major cause of food spoilage. Several yeasts, such as Filobasidiella neoformans, which cause cryptococcosis in humans, are also opportunistic pathogens, so a simple and rapid method for monitoring yeast contamination in food is essential. Here, we developed a simple and rapid method that utilizes loop-mediated isothermal amplification (LAMP) for the detection of F. neoformans. A set of five specific LAMP primers was designed that targeted the 5.8S-26S rDNA internal transcribed spacer 2 region of F. neoformans, and the primer set's specificity was confirmed. In a pure culture of F. neoformans, the LAMP assay had a lower sensitivity threshold of 10(2)cells/mL at a runtime of 60min. In a probiotic dairy product artificially contaminated with F. neoformans, the LAMP assay also had a lower sensitivity threshold of 10(2)cells/mL, which was comparable to the sensitivity of a quantitative PCR (qPCR) assay. We also developed a simple two-step method for the extraction of DNA from a probiotic dairy product that can be performed within 15min. This method involves initial protease treatment of the test sample at 45°C for 3min followed by boiling at 100°C for 5min under alkaline conditions. In a probiotic dairy product artificially contaminated with F. neoformans, analysis by means of our novel DNA extraction method followed by LAMP with our specific primer set had a lower sensitivity threshold of 10(3)cells/mL at a runtime of 60min. In contrast, use of our novel method of DNA extraction followed by qPCR assay had a lower sensitivity threshold of only 10(5)cells/mL at a runtime of 3 to 4h. Therefore, unlike the PCR assay, our LAMP assay can be used to quickly evaluate yeast contamination and is sensitive even for crude samples containing bacteria or background impurities. Our study provides a powerful tool for the primary screening of large numbers of food samples for yeast contamination. Copyright © 2014 Elsevier B.V. All rights reserved.

In vitro evaluation of five rapid antigen detection tests for group A beta-haemolytic streptococcal sore throat infections.

PubMed

Lasseter, Gemma M; McNulty, Cliodna A M; Richard Hobbs, F D; Mant, David; Little, Paul

2009-12-01

Using accurate and easy to use rapid antigen detection tests (RADTs) to identify group A beta-haemolytic Streptococci (GABHS) sore throat infections could reduce unnecessary antibiotic prescribing and antimicrobial resistance. Although there is no international consensus on the use of RADTs, these kits have been widely adopted in Finland, France and the USA. Yet in the UK, the Clinical Knowledge Summaries, that provide the main online guidance for GPs, discourage RADTs use, citing their poor sensitivity and inability to impact on prescribing decisions in acute sore throat infections. The purpose of this study was to evaluate the ease of use and in vitro accuracy (sensitivity and specificity) of the five most commonly used RADTs in Europe (OSOM Ultra, Quickvue Dipstick, Streptatest, Clearview Exact Strep A and IMI Test Pack). To ensure the RADTs were evaluated objectively, a standardized in vitro method using known concentrations of GABHS was used to remove the inherent biases associated with clinical studies. The IMI Test Pack was the easiest RADT to use overall. The ability to detect all positive GABHS (sensitivity) varied considerably between kits from 95% [95% confidence interval (CI): 88-98%], for the IMI Test Pack and OSOM, to 62% (95% CI: 51-72%) for Clearview, at the highest GABHS concentration. None of the RADTs gave any false-positive results with commensal flora-they were 100% specific. The IMI Test Pack is most suitable for use in primary care, as it had high sensitivity, high specificity and was easy to use.

A global comparative evaluation of commercial immunochromatographic rapid diagnostic tests for visceral leishmaniasis.

PubMed

Cunningham, Jane; Hasker, Epco; Das, Pradeep; El Safi, Sayda; Goto, Hiro; Mondal, Dinesh; Mbuchi, Margaret; Mukhtar, Maowia; Rabello, Ana; Rijal, Suman; Sundar, Shyam; Wasunna, Monique; Adams, Emily; Menten, Joris; Peeling, Rosanna; Boelaert, Marleen

2012-11-15

Poor access to diagnosis stymies control of visceral leishmaniasis (VL). Antibody-detecting rapid diagnostic tests (RDTs) can be performed in peripheral health settings. However, there are many brands available and published reports of variable accuracy. Commercial VL RDTs containing bound rK39 or rKE16 antigen were evaluated using archived human sera from confirmed VL cases (n = 750) and endemic non-VL controls (n = 754) in the Indian subcontinent (ISC), Brazil, and East Africa to assess sensitivity and specificity with 95% confidence intervals. A subset of RDTs were also evaluated after 60 days' heat incubation (37°C, 45°C). Interlot and interobserver variability was assessed. All test brands performed well against ISC panels (sensitivity range, 92.8%-100%; specificity range, 96%-100%); however, sensitivity was lower against Brazil and East African panels (61.5%-91% and 36.8%-87.2%, respectively). Specificity was consistently > 95% in Brazil and ranged between 90.8% and 98% in East Africa. Performance of some products was adversely affected by high temperatures. Agreement between lots and readers was good to excellent (κ > 0.73-0.99). Diagnostic accuracy of VL RDTs varies between the major endemic regions. Many tests performed well and showed good heat stability in the ISC; however, reduced sensitivity against Brazilian and East African panels suggests that in these regions, used alone, several RDTs are inadequate for excluding a VL diagnosis. More research is needed to assess ease of use and to compare performance using whole blood instead of serum and in patients coinfected with human immunodeficiency virus.

A novel highly sensitive, rapid and safe Rift Valley fever virus neutralization test.

PubMed

Wichgers Schreur, Paul J; Paweska, Janusz T; Kant, Jet; Kortekaas, Jeroen

2017-10-01

Antibodies specific for Rift Valley fever virus (RVFV) can be detected by diverse methods, including ezyme-linked immunosortbent assay (ELISA) and virus neutralization test (VNT). The VNT is superior in sensitivity and specificity and is therefore considered the gold standard serological assay. Classical VNTs make use of virulent RVFV and therefore have to be performed in biosafety level 3 laboratories. Here, we report the development of a novel VNT that is based on an avirulent RVFV expressing the enhanced green fluorescent protein (eGFP), which can be performed safely outside level 3 biocontainment facilities. Evaluation with a broad panel of experimental sera and field sera demonstrated that this novel VNT is faster and more sensitive than the classical VNT. Copyright © 2017 Elsevier B.V. All rights reserved.

Rapid quantification of viable Legionella in nuclear cooling tower waters using filter cultivation, fluorescent in situ hybridization and solid-phase cytometry.

PubMed

Baudart, J; Guillaume, C; Mercier, A; Lebaron, P; Binet, M

2015-05-01

To develop a rapid and sensitive method to quantify viable Legionella spp. in cooling tower water samples. A rapid, culture-based method capable of quantifying as few as 600 Legionella microcolonies per litre within 2 days in industrial waters was developed. The method combines a short cultivation step of microcolonies on GVPC agar plate, specific detection of Legionella cells by a fluorescent in situ hybridization (FISH) approach, and a sensitive enumeration using a solid-phase cytometer. Following optimization of the cultivation conditions, the qualitative and quantitative performance of the method was assessed and the method was applied to 262 nuclear power plant cooling water samples. The performance of this method was in accordance with the culture method (NF-T 90-431) for Legionella enumeration. The rapid detection of viable Legionella in water is a major concern to the effective monitoring of this pathogenic bacterium in the main water sources involved in the transmission of legionellosis infection (Legionnaires' disease). The new method proposed here appears to be a robust, efficient and innovative means for rapidly quantifying cultivable Legionella in cooling tower water samples within 48 h. © 2015 The Society for Applied Microbiology.

Fluorescence polarization immunoassays for rapid, accurate and sensitive determination of mycotoxins

USDA-ARS?s Scientific Manuscript database

Fluorescence polarization immunoassay (FPIA) is a type of homogeneous assay. For low molecular weight antigens, such as mycotoxins, it is based on the competition between an unlabeled antigen and its fluorescent-labeled derivative (tracer) for an antigen-specific antibody. The antigen content is det...

Rapid detection of multidrug-resistant Mycobacterium tuberculosis using the malachite green decolourisation assay

PubMed Central

Coban, Ahmet Yilmaz; Uzun, Meltem

2013-01-01

Early detection of drug resistance in Mycobacterium tuberculosis isolates allows for earlier and more effective treatment of patients. The aim of this study was to investigate the performance of the malachite green decolourisation assay (MGDA) in detecting isoniazid (INH) and rifampicin (RIF) resistance in M. tuberculosis clinical isolates. Fifty M. tuberculosis isolates, including 19 multidrug-resistant, eight INH-resistant and 23 INH and RIF-susceptible samples, were tested. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and agreement of the assay for INH were 92.5%, 91.3%, 92.5%, 91.3% and 92%, respectively. Similarly, the sensitivity, specificity, PPV, NPV and agreement of the assay for RIF were 94.7%, 100%, 100%, 96.8% and 98%, respectively. There was a major discrepancy in the tests of two isolates, as they were sensitive to INH by the MGDA test, but resistant by the reference method. There was a minor discrepancy in the tests of two additional isolates, as they were sensitive to INH by the reference method, but resistant by the MGDA test. The drug susceptibility test results were obtained within eight-nine days. In conclusion, the MGDA test is a reliable and accurate method for the rapid detection of INH and RIF resistance compared with the reference method and the MGDA test additionally requires less time to obtain results. PMID:24402143

Rapid assessment of tetanus vaccine-induced immunity in Bangladesh and the Gambia.

PubMed

Ramakrishnan, Girija; Wright, Marcia; Alam, Masud; Naylor, Caitlin; Kabir, Mamun; Zerin, Ayesha; Ferdous, Tahsin; Pedersen, Karl; Hennig, Branwen J; Donowitz, Jeffrey R; Wegmuller, Rita; Haque, Rashidul; Petri, William A; Herbein, Joel; Gilchrist, Carol A

2017-03-01

We have developed recombinant fragment C based rapid point of care dipstick devices to assess tetanus immunization status using plasma or whole blood. The devices demonstrated specificity of 0.90 and sensitivity of 0.90 (whole blood)/0.94 (plasma) at field sites in Bangladesh and The Gambia when compared to a commercial ELISA with the immune cut-off titer set as ≥0.1IU/mL. Copyright © 2016 Elsevier Inc. All rights reserved.

Multi-Country Evaluation of the Sensitivity and Specificity of Two Commercially-Available NS1 ELISA Assays for Dengue Diagnosis

PubMed Central

Guzman, Maria G.; Jaenisch, Thomas; Gaczkowski, Roger; Ty Hang, Vo Thi; Sekaran, Shamala Devi; Kroeger, Axel; Vazquez, Susana; Ruiz, Didye; Martinez, Eric; Mercado, Juan C.; Balmaseda, Angel; Harris, Eva; Dimano, Efren; Leano, Prisca Susan A.; Yoksan, Sutee; Villegas, Elci; Benduzu, Herminia; Villalobos, Iris; Farrar, Jeremy; Simmons, Cameron P.

2010-01-01

Background Early diagnosis of dengue can assist patient triage and management and prevent unnecessary treatments and interventions. Commercially available assays that detect the dengue virus protein NS1 in the plasma/serum of patients offers the possibility of early and rapid diagnosis. Methodology/Principal Findings The sensitivity and specificity of the Pan-E Dengue Early ELISA and the Platelia™ Dengue NS1 Ag assays were compared against a reference diagnosis in 1385 patients in 6 countries in Asia and the Americas. Platelia was more sensitive (66%) than Pan-E (52%) in confirmed dengue cases. Sensitivity varied by geographic region, with both assays generally being more sensitive in patients from SE Asia than the Americas. Both kits were more sensitive for specimens collected within the first few days of illness onset relative to later time points. Pan-E and Platelia were both 100% specific in febrile patients without evidence of acute dengue. In patients with other confirmed diagnoses and healthy blood donors, Platelia was more specific (100%) than Pan-E (90%). For Platelia, when either the NS1 test or the IgM test on the acute sample was positive, the sensitivity versus the reference result was 82% in samples collected in the first four days of fever. NS1 sensitivity was not associated to disease severity (DF or DHF) in the Platelia test, whereas a trend for higher sensitivity in DHF cases was seen in the Pan-E test (however combined with lower overall sensitivity). Conclusions/Significance Collectively, this multi-country study suggests that the best performing NS1 assay (Platelia) had moderate sensitivity (median 64%, range 34–76%) and high specificity (100%) for the diagnosis of dengue. The poor sensitivity of the evaluated assays in some geographical regions suggests further assessments are needed. The combination of NS1 and IgM detection in samples collected in the first few days of fever increased the overall dengue diagnostic sensitivity. PMID:20824173

A rapid immunochromatographic test strip for detecting rabies virus antibody.

PubMed

Wang, Hualei; Feng, Na; Yang, Songtao; Wang, Chengyu; Wang, Tiecheng; Gao, Yuwei; Su, Jianqing; Zheng, Xuexing; Hou, Xiaoqiang; Huang, Hainan; Yang, Ruimei; Zou, Xiaohuan; Huang, Geng; Xia, Xianzhu

2010-12-01

An immunochromatographic test strip (ICTS) for detecting antibodies to rabies virus was developed, using colloidal gold particles labeled with rabies virus glycoprotein as the tracer. The assay was evaluated using sera from dogs immunized with various commercial rabies vaccines, or from dogs in the clinics and sera from dogs immunized with vaccines against pathogens other than rabies virus, and negative sera from a wide variety of animal sources, including dogs, mice, and cats which had never been vaccinated. The ICTS was found to be highly specific for antibodies against rabies virus, with a detection limit of 0.5IU/ml as measured by the fluorescent antibody virus neutralization (FAVN) test. Compared with the FAVN test, the specificity and sensitivity of ICTS were 98.2% and 90.4%, respectively. There was an excellent agreement between results obtained by the ICTS and FAVN tests (kappa=0.888). Strips stored at 4°C in a plastic bag with a desiccant retained their specificity and sensitivity for at least 15 months, and strips stored at ambient temperature remained stable for 12 months. The immunochromatographic test strip may therefore be useful for clinical laboratories lacking specialized equipment and for diagnosis in the field for rapid detection of rabies virus-specific antibodies. Copyright © 2010 Elsevier B.V. All rights reserved.

Rapid and specific detection of Salmonella in water samples using real-time PCR and High Resolution Melt (HRM) curve analysis.

PubMed

van Blerk, G N; Leibach, L; Mabunda, A; Chapman, A; Louw, D

2011-01-01

A real-time PCR assay combined with a pre-enrichment step for the specific and rapid detection of Salmonella in water samples is described. Following amplification of the invA gene target, High Resolution Melt (HRM) curve analysis was used to discriminate between products formed and to positively identify invA amplification. The real-time PCR assay was evaluated for specificity and sensitivity. The assay displayed 100% specificity for Salmonella and combined with a 16-18 h non-selective pre-enrichment step, the assay proved to be highly sensitive with a detection limit of 1.0 CFU/ml for surface water samples. The detection assay also demonstrated a high intra-run and inter-run repeatability with very little variation in invA amplicon melting temperature. When applied to water samples received routinely by the laboratory, the assay showed the presence of Salmonella in particularly surface water and treated effluent samples. Using the HRM based assay, the time required for Salmonella detection was drastically shortened to less than 24 h compared to several days when using standard culturing methods. This assay provides a useful tool for routine water quality monitoring as well as for quick screening during disease outbreaks.

Improved PCR assay for the specific detection and quantitation of Escherichia coli serotype O157 in water.

PubMed

Cho, Min Seok; Joh, Kiseong; Ahn, Tae-Young; Park, Dong Suk

2014-09-01

Escherichia coli serotype O157 is still a major global healthcare problem. However, only limited information is now available on the molecular and serological detection of pathogenic bacteria. Therefore, the development of appropriate strategies for their rapid identification and monitoring is still needed. In general, the sequence analysis based on stx, slt, eae, hlyA, rfb, and fliCh7 genes is widely employed for the identification of E. coli serotype O157; but there have been critical defects in the diagnosis and identification of E. coli serotype O157, in that they are also present in other E. coli serogroups. In this study, NCBI-BLAST searches using the nucleotide sequences of the putative regulatory protein gene from E. coli O157:H7 str. Sakai found sequence difference at the serotype level. The specific primers from the putative regulatory protein gene were designed and investigated for their sensitivity and specificity for detecting the pathogen in environment water samples. The specificity of the primer set was evaluated using genomic DNA from 8 isolates of E. coli serotype O157 and 32 other reference strains. In addition, the sensitivity and specificity of this assay were confirmed by successful identification of E. coli serotype O157 in environmental water samples. In conclusion, this study showed that the newly developed quantitative serotype-specific PCR method is a highly specific and efficient tool for the surveillance and rapid detection of high-risk E. coli serotype O157.

Rapid detection of methicillin resistance in coagulase-negative staphylococci by a penicillin-binding protein 2a-specific latex agglutination test.

PubMed

Horstkotte, M A; Knobloch, J K; Rohde, H; Mack, D

2001-10-01

The detection of PBP 2a by the MRSA-Screen latex agglutination test with 201 clinical coagulase-negative staphylococci had an initial sensitivity of 98% and a high degree of specificity for Staphylococcus epidermidis strains compared to PCR for mecA. Determination of oxacillin MICs evaluated according to the new breakpoint (0.5 microg/ml) of the National Committee for Clinical Laboratory Standards exhibited an extremely low specificity for this population.

A simple, rapid, cost-effective and sensitive method for detection of Salmonella in environmental and pecan samples.

PubMed

Dobhal, S; Zhang, G; Rohla, C; Smith, M W; Ma, L M

2014-10-01

PCR is widely used in the routine detection of foodborne human pathogens; however, challenges remain in overcoming PCR inhibitors present in some sample matrices. The objective of this study was to develop a simple, sensitive, cost-effective and rapid method for processing large numbers of environmental and pecan samples for Salmonella detection. This study was also aimed at validation of a new protocol for the detection of Salmonella from in-shell pecans. Different DNA template preparation methods, including direct boiling, prespin, multiple washing and commercial DNA extraction kits, were evaluated with pure cultures of Salmonella Typhimurium and with enriched soil, cattle feces and in-shell pecan each spiked individually with Salmonella Typhimurium. PCR detection of Salmonella was conducted using invA and 16S rRNA gene (internal amplification control) specific primers. The effect of amplification facilitators, including bovine serum albumin (BSA), polyvinylpyrrolidone (PVP), polyethylene glycol (PEG) and gelatin on PCR sensitivity, was also evaluated. Conducting a prespin of sample matrices in combination with the addition of 0·4% (w/v) BSA and 1% (w/v) PVP in PCR mix was the simplest, most rapid, cost-effective and sensitive method for PCR detection of Salmonella, with up to 40 CFU Salmonella per reaction detectable in the presence of over 10(9 ) CFU ml(-1) of background micro-organisms from enriched feces soil or pecan samples. The developed method is rapid, cost-effective and sensitive for detection of Salmonella from different matrices. This study provides a method with broad applicability for PCR detection of Salmonella in complex sample matrices. This method has a potential for its application in different research arenas and diagnostic laboratories. © 2014 The Society for Applied Microbiology.

Detection of Streptococcus pyogenes using rapid visual molecular assay.

PubMed

Zhao, Xiangna; He, Xiaoming; Li, Huan; Zhao, Jiangtao; Huang, Simo; Liu, Wei; Wei, Xiao; Ding, Yiwei; Wang, Zhaoyan; Zou, Dayang; Wang, Xuesong; Dong, Derong; Yang, Zhan; Yan, Xiabei; Huang, Liuyu; Du, Shuangkui; Yuan, Jing

2015-09-01

Streptococcus pyogenes is an increasingly important pathogen in many parts of the world. Rapid and accurate detection of S. pyogenes aids in the control of the infection. In this study, a loop-mediated isothermal amplification (LAMP) assay was developed and validated for the specific detection of S. pyogenes. The assay incorporates two methods: a chromogenic analysis using a calcein/Mn(2+) complex and real-time turbidity monitoring to assess the reaction. Both methods detected the target DNA within 60 min under 64°C isothermal conditions. The assay used specifically designed primers to target spy1258, and correctly identified 111 strains of S. pyogenes and 32 non-S. pyogenes strains, including other species of the genus Streptococcus. Tests using reference strains showed that the LAMP assay was highly specific. The sensitivity of the assay, with a detection limit of 1.49 pg DNA, was 10-fold greater than that of PCR. The LAMP assay established in this study is simple, fast and sensitive, and does not rely upon any special equipment; thus, it could be employed in clinical diagnosis. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Rapid and specific detection of porcine parvovirus by isothermal recombinase polymerase amplification assays.

PubMed

Yang, Yang; Qin, Xiaodong; Zhang, Wei; Li, Yanmin; Zhang, Zhidong

2016-10-01

Porcine parvovirus (PPV) is a major cause of swine reproductive failure and reported in many countries worldwide. Recombinase polymerase amplification (RPA) assays using a real-time fluorescent detection (PPV real-time RPA assay) and a lateral flow dipstick (PPV RPA LFD assay) were developed targeting PPV NS1 gene. The detection limit of PPV real-time RPA assay was 300 copies per reaction within 9 min at 38 °C, while the RPA LFD assay has a detection limit of 400 copies per reaction in less than 20 min at 38 °C. In both assays, there were no cross-reactions with porcine circovirus type 2, pseudorabies virus, porcine reproductive and respiratory syndrome virus, classical swine fever virus, and foot-and-mouth disease virus. Based on a total of 128 clinical samples examined, the sensitivity and the specificity of the developed RPA assays for identification of PPV was 94.4% and 100%, respectively, when compared to real-time (qPCR) assay. Therefore, the RPA assay provides a rapid, sensitive and specific alternative for PPV detection. Copyright © 2016 Elsevier Ltd. All rights reserved.

Blinded study determination of high sensitivity and specificity microchip electrophoresis-SSCP/HA to detect mutations in the p53 gene.

PubMed

Hestekin, Christa N; Lin, Jennifer S; Senderowicz, Lionel; Jakupciak, John P; O'Connell, Catherine; Rademaker, Alfred; Barron, Annelise E

2011-11-01

Knowledge of the genetic changes that lead to disease has grown and continues to grow at a rapid pace. However, there is a need for clinical devices that can be used routinely to translate this knowledge into the treatment of patients. Use in a clinical setting requires high sensitivity and specificity (>97%) in order to prevent misdiagnoses. Single-strand conformational polymorphism (SSCP) and heteroduplex analysis (HA) are two DNA-based, complementary methods for mutation detection that are inexpensive and relatively easy to implement. However, both methods are most commonly detected by slab gel electrophoresis, which can be labor-intensive, time-consuming, and often the methods are unable to produce high sensitivity and specificity without the use of multiple analysis conditions. Here, we demonstrate the first blinded study using microchip electrophoresis (ME)-SSCP/HA. We demonstrate the ability of ME-SSCP/HA to detect with 98% sensitivity and specificity >100 samples from the p53 gene exons 5-9 in a blinded study in an analysis time of <10 min. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

[Flicker comparison of optic disc photographs: sensitivity and specificity].

PubMed

Funk, Jens; Lagrèze, Wolf; Zeyen, Thierry

2002-12-01

Examination and documentation of the optic nerve head are essential in monitoring glaucoma patients. Even minor changes in optic nerve head morphology can be visualised using the so-called flicker test: Two optic nerve head photographs, taken at consecutive examinations, are superimposed by projection. When occluding the pictures in a rapid alternating fashion, changes in optic nerve head morphology appear as motion. In this study, we evaluated sensitivity and specificity of the flicker test. A set of 33 pairs of serial optic disc slides was used as gold standard. These 33 pairs had been classified earlier by 3 independent groups of experts. 23 had been classified as "no change over time", 10 had been classified as "change". All 33 pairs were now evaluated by flicker comparison in a masked fashion. Flicker comparison usually took 1 minute per pair of slides. Sensitivity was 90 %, specificity was 65 %. The sensitivity was reasonably high. The moderate specificity was due to some cases showing "change" with the flicker comparison which might have been overlooked by the expert groups. Flicker comparison is an easy, fast and reliable technique to evaluate pairs of consecutive optic disc photographs.

Improving the Specificity of Plasmodium falciparum Malaria Diagnosis in High-Transmission Settings with a Two-Step Rapid Diagnostic Test and Microscopy Algorithm.

PubMed

Murungi, Moses; Fulton, Travis; Reyes, Raquel; Matte, Michael; Ntaro, Moses; Mulogo, Edgar; Nyehangane, Dan; Juliano, Jonathan J; Siedner, Mark J; Boum, Yap; Boyce, Ross M

2017-05-01

Poor specificity may negatively impact rapid diagnostic test (RDT)-based diagnostic strategies for malaria. We performed real-time PCR on a subset of subjects who had undergone diagnostic testing with a multiple-antigen (histidine-rich protein 2 and pan -lactate dehydrogenase pLDH [HRP2/pLDH]) RDT and microscopy. We determined the sensitivity and specificity of the RDT in comparison to results of PCR for the detection of Plasmodium falciparum malaria. We developed and evaluated a two-step algorithm utilizing the multiple-antigen RDT to screen patients, followed by confirmatory microscopy for those individuals with HRP2-positive (HRP2 + )/pLDH-negative (pLDH - ) results. In total, dried blood spots (DBS) were collected from 276 individuals. There were 124 (44.9%) individuals with an HRP2 + /pLDH + result, 94 (34.1%) with an HRP2 + /pLDH - result, and 58 (21%) with a negative RDT result. The sensitivity and specificity of the RDT compared to results with real-time PCR were 99.4% (95% confidence interval [CI], 95.9 to 100.0%) and 46.7% (95% CI, 37.7 to 55.9%), respectively. Of the 94 HRP2 + /pLDH - results, only 32 (34.0%) and 35 (37.2%) were positive by microscopy and PCR, respectively. The sensitivity and specificity of the two-step algorithm compared to results with real-time PCR were 95.5% (95% CI, 90.5 to 98.0%) and 91.0% (95% CI, 84.1 to 95.2), respectively. HRP2 antigen bands demonstrated poor specificity for the diagnosis of malaria compared to that of real-time PCR in a high-transmission setting. The most likely explanation for this finding is the persistence of HRP2 antigenemia following treatment of an acute infection. The two-step diagnostic algorithm utilizing microscopy as a confirmatory test for indeterminate HRP2 + /pLDH - results showed significantly improved specificity with little loss of sensitivity in a high-transmission setting. Copyright © 2017 American Society for Microbiology.

[The establishment of a novel method of nano-immunomagnetic separation and Real-time PCR for detecting Vibrio cholerae from seafood].

PubMed

Cheng, Jinxia; Zeng, Jing; Liu, Li; Wei, Haiyan; Zhao, Xiaojuan; Zhang, Ximeng; Zhang, Lei; Zhang, Haiyu

2014-02-01

A novel method of Nano-Immunomagnetic Separation (Nano-IMS) plus Real-time PCR was established for detecting Vibrio cholerae. The Nano-Immunomagnetic Beads were created by using the monoclonal antibody of Vibrio cholerae, which was named Nano-IMB-Vc. Nano-IMB-Vc has specific adsorption of Vibrio cholerae, combined with Real-time PCR technology, a method for rapid detection of Vibrio cholerae was established. The capture specificity of Nano-IMB-Vc was tested by using 15 bacteria strains. The specificity of Real-time PCR method was tested by using 102 targets and 101 non-targets bacteria strains. The sensitivity of Nano-IMS plus Real-time PCR were tested in pure culture and in artificial samples and compared with NMKL No.156. The capture ratio of Nano-IMB-Vc was reached 70.2% at the level of 10(3) CFU/ml. In pure culture, the sensitivity of Nano-IMS plus Real-time PCR was reached at 5.4×10(2) CFU/ml. The specific of Real-time PCR method was tested by using 102 targets and 101 non-targets bacteria. The results showed that 102 strains of Vibrio cholerae test results were all positive, and the rest of the 101 strains of non-target bacteria test results were negative. No cross-reaction was founded. Add 1 CFU vibrio cholerae per 25 g sample, it could be detect with Nano-IMS plus Real-time PCR method after 8 hours enrichment. The Nano-IMS plus Real-time PCR method of Vibrio cholerae established in this study has good specificity and sensitivity, which could be applied to the rapid detection of Vibrio cholerae.

Evaluation of an Immunochromatographic Test for Rapid and Reliable Serodiagnosis of Human Tularemia and Detection of Francisella tularensis-Specific Antibodies in Sera from Different Mammalian Species ▿

PubMed Central

Splettstoesser, W.; Guglielmo-Viret, V.; Seibold, E.; Thullier, P.

2010-01-01

Tularemia is a highly contagious infectious zoonosis caused by the bacterial agent Francisella tularensis. Serology is still considered to be a cornerstone in tularemia diagnosis due to the low sensitivity of bacterial culture and the lack of standardization in PCR methodology for the direct identification of the pathogen. We developed a novel immunochromatographic test (ICT) to efficiently detect F. tularensis-specific antibodies in sera from humans and other mammalian species (nonhuman primate, pig, and rabbit). This new tool requires none or minimal laboratory equipment, and the results are obtained within 15 min. When compared to the method of microagglutination, which was shown to be more specific than the enzyme-linked immunosorbent assay, the ICT had a sensitivity of 98.3% (58 positive sera were tested) and a specificity of 96.5% (58 negative sera were tested) on human sera. On animal sera, the overall sensitivity was 100% (22 positive sera were tested) and specificity was also 100% (70 negative sera were tested). This rapid test preferentially detects IgG antibodies that may occur early in the course of human tularemia, but further evaluation with human sera is important to prove that the ICT can be a valuable field test to support a presumptive diagnosis of tularemia. The ICT can also be a useful tool to monitor successful vaccination with subunit vaccines or live vaccine strains containing lipopolysaccharide (e.g., LVS) and to detect seropositive individuals or animals in outbreak situations or in the context of epidemiologic surveillance programs in areas of endemicity as recently recommended by the World Health Organization. PMID:20220165

Promoting Indian Library Use. Guide Number 7.

ERIC Educational Resources Information Center

Townley, Charles T.

Individuals, organizations, and American Indian tribes are rapidly recognizing the value of libraries. They are recognizing that libraries and the information services which they offer are necessary to meet Indian goals. Specific sensitivity to Indian ways and alternatives is just developing as library and information services develop in Indian…

How sensitive and specific is urinalysis 'dipstick' testing for detection of hyperglycaemia and ketosis? An audit of findings from coronial autopsies.

PubMed

Mitchell, Rebecca; Thomas, Sunethra Devika; Langlois, Neil E I

2013-10-01

Biochemical analysis of glucose and ketones in the vitreous humour obtained at post-mortem examination is representative of the levels in the blood prior to death. Elevated levels can be indicative of conditions including diabetic ketoacidosis, which can be a cause for unexpected death. A rapid screening test for such conditions can be performed during the autopsy through urinalysis using test strips (urine 'dipstick' testing). The aim of this study was to assess the utility of urinalysis testing for post-mortem detection of derangements of glucose and ketone levels. The results of vitreous biochemical analysis and urinalysis were collated from 188 forensic autopsy cases. A vitreous glucose result of above 10 mmol/L was regarded as high. When this was compared to urinalysis results it was found that any urinalysis result above negative had a sensitivity of 0.83 and a specificity of 0.93. A vitreous ketone level of above 5 mmol/L was regarded as significantly elevated; a urinalysis result above negative had a sensitivity of 1, but a specificity of 0.12. Urinalysis ('dipstick' testing) for glucose has a good sensitivity and specificity for high vitreous glucose levels, which are regarded as indicative of pathological hyperglycaemia during life. It was found that urine testing for ketones either has an excellent sensitivity with low specificity or a poor sensitivity with a good specificity; however, this finding has to be viewed in the context of uncertainty of the biochemical level of significant ketosis.

Dual Electrophoresis Detection System for Rapid and Sensitive Immunoassays with Nanoparticle Signal Amplification

NASA Astrophysics Data System (ADS)

Zhang, Fangfang; Ma, Junjie; Watanabe, Junji; Tang, Jinlong; Liu, Huiyu; Shen, Heyun

2017-02-01

An electrophoretic technique was combined with an enzyme-linked immunosorbent assay (ELISA) system to achieve a rapid and sensitive immunoassay. A cellulose acetate filter modified with polyelectrolyte multilayer (PEM) was used as a solid substrate for three-dimensional antigen-antibody reactions. A dual electrophoresis process was used to induce directional migration and local condensation of antigens and antibodies at the solid substrate, avoiding the long diffusion times associated with antigen-antibody reactions in conventional ELISAs. The electrophoretic forces drove two steps in the ELISA process, namely the adsorption of antigen, and secondary antibody-labelled polystyrene nanoparticles (NP-Ab). The total time needed for dual electrophoresis-driven detection was just 4 min, nearly 2 h faster than a conventional ELISA system. Moreover, the rapid NP-Ab electrophoresis system simultaneously achieved amplification of the specific signal and a reduction in noise, leading to a more sensitive NP-Ab immunoassay with a limit of detection (LOD) of 130 fM, and wide range of detectable concentrations from 0.13 to 130 pM. These results suggest that the combination of dual electrophoresis detection and NP-Ab signal amplification has great potential for future immunoassay systems.

Rapid Clinical Score for the Diagnosis of Tuberculous Meningitis: A Retrospective Cohort Study

PubMed Central

Jipa, Raluca; Olaru, Ioana D.; Manea, Eliza; Merisor, Simona; Hristea, Adriana

2017-01-01

Objective: The aim of our study was to retrospectively validate a previously described rapid clinical score (RCS) in distinguishing tuberculous meningitis (TBM) from viral meningitis (VM) in people who are at increased risk of tuberculosis, as well as from cryptococcal meningitis (CM) in HIV-infected patients. Methods: We performed a retrospective study of patients admitted with a diagnosis of aseptic meningitis between January 2012 and December 2015, to a referral hospital for infectious diseases. The variables included in RCS were duration of symptoms before admission, neurological stage, cerebrospinal fluid (CSF) to blood glucose ratio, and CSF protein. We included in this retrospective study 31 patients with definite or probable TBM including 14 HIV-infected patients, 62 HIV-noninfected patients with VM, and 18 HIV-infected patients with CM. Results: The sensitivity of RCS to distinguish TBM from VM was 96.7%, with a specificity of 81.1% and the area under the receiver operating characteristic (ROC) curve was 0.949 (0.90–0.99). When all four criteria from the RCS were present, the specificity increased at 100%. In HIV-infected patients, the sensitivity and specificity of RCS in differentiating TBM from CM were 86.6% and 27.7%, respectively, and the area under the ROC curve was 0.669 (0.48–0.85). Conclusion: This easy-to-use RCS was found to be helpful in differentiating TBM from VM, with a better sensitivity than molecular amplification techniques and a relatively good specificity. However, the RCS was not useful to differentiate between TBM and CM in HIV-infected patients. PMID:29184338

Inter-laboratory evaluation of the performance parameters of a Lateral Flow Test device for the detection of Bluetongue virus-specific antibodies.

PubMed

Hanon, Jean-Baptiste; Vandenberge, Valerie; Deruelle, Matthias; De Leeuw, Ilse; De Clercq, Kris; Van Borm, Steven; Koenen, Frank; Liu, Lihong; Hoffmann, Bernd; Batten, Carrie Anne; Zientara, Stéphan; Breard, Emmanuel; Van der Stede, Yves

2016-02-01

Bluetongue (BT) is a viral vector-borne disease affecting domestic and wild ruminants worldwide. In this study, a commercial rapid immuno-chromatographic method or Lateral Flow Test (LFT) device, for the detection of BT virus-specific antibodies in animal serum, was evaluated in an international inter-laboratory proficiency test. The evaluation was done with sera samples of variable background (ruminant species, serotype, field samples, experimental infections, vaccinated animals). The diagnostic sensitivity was 100% (95% C.I. [90.5-100]) and the diagnostic specificity was 95.2% (95% C.I. [76.2-99.9]). The repeatability (accordance) and reproducibility (concordance) were 100% for seropositive samples but were lower for two of the seronegative samples (45% and 89% respectively). The analytical sensitivity, evaluated by testing positive sera at increasing dilutions was better for the BT LFT compared to some commercial ELISAs. Seroconversion of an infected sheep was detected at 4 days post infection. Analytical specificity was impaired by cross-reactions observed with some of the samples seropositive for Epizootic Haemorrhagic Disease Virus (EHDV). The agreement (Cohen's kappa) between the LFT and a commercial BT competitive ELISA was 0.79 (95% CI [0.62-0.95]). Based on these results, it can be concluded that the BT LFT device is a rapid and sensitive first-line serological test that can be used in the field, especially in areas endemic for the disease where there is a lack of diagnostic facilities. Copyright © 2015 Elsevier B.V. All rights reserved.

Rapid Characterization of Microorganisms by Mass Spectrometry—What Can Be Learned and How?

NASA Astrophysics Data System (ADS)

Fenselau, Catherine C.

2013-08-01

Strategies for the rapid and reliable analysis of microorganisms have been sought to meet national needs in defense, homeland security, space exploration, food and water safety, and clinical diagnosis. Mass spectrometry has long been a candidate technique because it is extremely rapid and can provide highly specific information. It has excellent sensitivity. Molecular and fragment ion masses provide detailed fingerprints, which can also be interpreted. Mass spectrometry is also a broad band method—everything has a mass—and it is automatable. Mass spectrometry is a physiochemical method that is orthogonal and complementary to biochemical and morphological methods used to characterize microorganisms.

Rapid characterization of microorganisms by mass spectrometry--what can be learned and how?

PubMed

Fenselau, Catherine C

2013-08-01

Strategies for the rapid and reliable analysis of microorganisms have been sought to meet national needs in defense, homeland security, space exploration, food and water safety, and clinical diagnosis. Mass spectrometry has long been a candidate technique because it is extremely rapid and can provide highly specific information. It has excellent sensitivity. Molecular and fragment ion masses provide detailed fingerprints, which can also be interpreted. Mass spectrometry is also a broad band method--everything has a mass--and it is automatable. Mass spectrometry is a physiochemical method that is orthogonal and complementary to biochemical and morphological methods used to characterize microorganisms.

Label-Free Electrical Immunosensor for Highly Sensitive and Specific Detection of Microcystin-LR in Water Samples.

PubMed

Tan, Feng; Saucedo, Nuvia Maria; Ramnani, Pankaj; Mulchandani, Ashok

2015-08-04

Microcystin-LR (MCLR) is one of the most commonly detected and toxic cyclic heptapeptide cyanotoxins released by cyanobacterial blooms in surface waters, for which sensitive and specific detection methods are necessary to carry out its recognition and quantification. Here, we present a single-walled carbon nanotube (SWCNTs)-based label-free chemiresistive immunosensor for highly sensitive and specific detection of MCLR in different source waters. MCLR was initially immobilized on SWCNTs modified interdigitated electrode, followed by incubation with monoclonal anti-MCLR antibody. The competitive binding of MCLR in sample solutions induced departure of the antibody from the antibody-antigen complexes formed on SWCNTs, resulting in change in the conductivity between source and drain of the sensor. The displacement assay greatly improved the sensitivity of the sensor compared with direct immunoassay on the same device. The immunosensor exhibited a wide linear response to log value of MCLR concentration ranging from 1 to 1000 ng/L, with a detection limit of 0.6 ng/L. This method showed good reproducibility, stability and recovery. The proposed method provides a powerful tool for rapid and sensitive monitoring of MCLR in environmental samples.

Diagnostic Value of Cerebrospinal Fluid T-SPOT.TB for Tuberculousis Meningitis in China.

PubMed

Li, Xue Lian; Xie, Na; Wang, Song Wang; Wu, Qian Hong; Ma, Yan; Shu, Wei; Chen, Hong Mei; Zhang, Li Qun; Wu, Xiao Guang; Ma, Li Ping; Che, Nan Ying; Gao, Meng Qiu

2017-09-01

The aim of this study was to evaluate the diagnostic value of the cerebrospinal fluid (CSF) T-SPOT.TB test for the diagnosis of TB meningitis (TBM). A retrospective analysis of 96 patients with manifested meningitis was conducted; T-SPOT.TB test was performed for diagnosing TBM to determine the diagnostic sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). A receiver operating characteristic (ROC) curve was also drawn to assess the diagnostic accuracy. The sensitivity, specificity, PPV, and NPV of CSF T-SPOT.TB test were 97.8%, 78.0%, 80.3%, and 97.5%, respectively, for 52 patients (54.2%) of the 96 enrolled patients. The area under the curve (AUC) was 0.910, and the sensitivities of CSF T-SPOT.TB for patients with stages I, II, and III of TBM were 96.7%, 97.2%, and 98.9%, respectively. CSF T-SPOT.TB test is a rapid and accurate diagnostic method with higher sensitivity and specificity for diagnosing TBM. Copyright © 2017 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

Rapid Detection of Enterobacter Sakazakii in milk Powder using amino modified chitosan immunomagnetic beads.

PubMed

Zhu, Yinglian; Wang, Dongfeng

2016-12-01

Chitosan immunomagnetic beads (CIBs) were first prepared through converting hydroxyl groups of natural polymer material-chitosan into amino groups using epichlorohydrin and ethylenediamine as modification agent and then coupling with polyclonal antibodies of Enterobacter sakazakii using glutaraldehyde as cross-linking agent. The beads before coupling with antibodies were characterized by magnetic property measurement, FTIR, SEM and XRD technologies. In the assay a natural polysaccharide-chitosan, which has good biological and chemical properties such as non-toxicity, biocompatibility and high chemical reactivity was first used for synthesis of immunomagnetic beads. The detection method first established in this paper that combined the beads with chromogenic medium together to rapid detect E. sakazakii in milk powder could greatly improve the detection specificity and working efficiency. The beads exhibited a maximum capturing capacity of 1×10 6 cfu/g with the detection sensitivity of 4cfu/g. The results demonstrate that the assay is a straightforward, specific and sensitive alternative for rapid detection of E.sakazakii in food matrix. The total analysis time was as little as about 25h, which greatly shorten the detection time. The method can provides new ideas not only to preparation technique of immunomagnetic beads but to imunne detection technique in food safety. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparison of Real-Time PCR and Conventional Biochemical Methods for Identification of Staphylococcus lugdunensis▿

PubMed Central

Pinsky, Benjamin A.; Samson, Divinia; Ghafghaichi, Laleh; Baron, Ellen J.; Banaei, Niaz

2009-01-01

Staphylococcus lugdunensis is an aggressive, virulent member of the coagulase-negative staphylococci (CoNS) that is responsible for severe, rapidly progressive skin and soft tissue infections and native valve endocarditis. To facilitate prompt identification and appropriate therapy, we describe here a rapid and robust multiplex real-time PCR assay that is able to definitively distinguish S. lugdunensis from other staphylococci. Using melting curve analysis, the assay also identifies Staphylococcus aureus and CoNS other than S. lugdunensis and determines MecA-dependent resistance to methicillin (meticillin). When applied to a panel of well-characterized staphylococcal reference strains, as well as 165 clinical isolates previously identified by conventional methods, the assay was both sensitive and specific for S. lugdunensis, correctly identifying the reference strain and all 47 S. lugdunensis isolates without inappropriate amplification of other staphylococci. Furthermore, rapid biochemical identification using the WEE-TAB system to detect ornithine decarboxylase activity was found to be unsuitable as an alternative to PCR identification, displaying just 31% sensitivity and 77% specificity when tested on a subset (90 isolates) of the clinical strains. We therefore propose that this simple, accurate PCR approach will allow for the routine and timely identification of S. lugdunensis in the clinical microbiology laboratory. PMID:19741081

Development of simple and rapid assay to detect viral RNA of tick-borne encephalitis virus by reverse transcription-loop-mediated isothermal amplification.

PubMed

Hayasaka, Daisuke; Aoki, Kotaro; Morita, Kouichi

2013-03-04

Tick-borne encephalitis virus (TBEV) is a causative agent of acute central nervous system disease in humans. It has three subtypes, far eastern (FE), Siberian (Sib) and European (Eu) subtypes, which are distributed over a wide area of Europe and Asia. The objective of this study was to develop a simple and rapid assay for the detection of TBEV RNA by using reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) method that can differentiate the three subtypes of TBEV and can be used for clinical diagnosis and epidemiological study. Primers for TBEV-specific and subtype-specific RT-LAMP assay were designed to target the consensus sequence in NS1 of all subtypes and the consensus sequence in the E gene of each subtype, respectiveluy. In vitro transcribed RNA of Oshima strain that belongs to FE subtype was serially diluted and used to examine the sensitivity of the assay. Cross-reactivity of subtype-specific RT-LAMP assay was tested by using the RNA of Oshima and Sofjin (FE), IR-99 (Sib) and Hochosterwitz (Eu) strains. RNA extracted from the mixtures of TBEV and ticks, and of TBEV and human blood, and the mouse tissues infected with TBEV, were evaluated in the assay. Positive amplification was observed by real-time monitoring of turbidity and by visual detection of color change. The sensitivity of TBEV-specific RT-LAMP assay was 102 copies of target RNA per reaction volume. FE-specific RT-LAMP assay amplified viral genes of Oshima and Sofjin strains but not of IR-99 and Hochosterwitz strains, and of Japanese encephalitis virus. RT-LAMP assay for Sib and for Eu specifically amplified viral genes of IR-99 and Hochosterwitz strains, respectively. We also showed that tick or human blood extract did not inhibit the amplification of viral gene during the assay. Furthermore, we confirmed that the TBEV RT-LAMP could detect virus RNA from peripheral and central nervous system tissues of laboratory mice infected with TBEV. TBEV RT-LAMP assay offers a sensitive, specific, rapid and easy-to-handle method for the detection of TBEV RNA in tick samples and this may be applied in the clinical samples collected from TBE-suspected patients.

Rapid detection of Streptococcus pneumoniae by real-time fluorescence loop-mediated isothermal amplification

PubMed Central

Guo, Xu-Guang; Zhou, Shan

2014-01-01

Background and aim of study A significant human pathogenic bacterium, Streptococcus pneumoniae was recognized as a major cause of pneumonia, and is the subject of many humoral immunity studies. Diagnosis is generally made based on clinical suspicion along with a positive culture from a sample from virtually any place in the body. But the testing time is too long. This study is to establish a rapid diagnostic method to identification of Streptococcus pneumoniae. Methods Our laboratory has recently developed a new platform called real-amp, which combines loop-mediated isothermal amplification (LAMP) with a portable tube scanner real-time isothermal instrument for the rapid detection of Streptococcus pneumonia. Two pairs of amplification primers required for this method were derived from a conserved DNA sequence unique to the Streptococcus pneumoniae. The amplification was carried out at 63 degree Celsius using SYBR Green for 60 minutes with the tube scanner set to collect fluorescence signals. Clinical samples of Streptococcus pneumoniae and other bacteria were used to determine the sensitivity and specificity of the primers by comparing with traditional culture method. Results The new set of primers consistently detected in laboratory-maintained isolates of Streptococcus pneumoniae from our hospital. The new primers also proved to be more sensitive than the published species-specific primers specifically developed for the LAMP method in detecting Streptococcus pneumoniae. Conclusions This study demonstrates that the Streptococcus pneumoniae LAMP primers developed here have the ability to accurately detect Streptococcus pneumoniae infections by real-time fluorescence LAMP. PMID:25276360

A novel DANP-coupled hairpin RT-PCR for rapid detection of Chikungunya virus.

PubMed

Chen, Huixin; Takei, Fumie; Koay, Evelyn Siew-Chuan; Nakatani, Kazuhiko; Chu, Justin Jang Hann

2013-03-01

Chikungunya has re-emerged as an important arboviral infection of global health significance. Because of lack of a vaccine and effective treatment, rapid diagnosis plays an important role in early clinical management of patients. In this study, we have developed a novel molecular diagnostic platform that ensures a rapid and cost-effective one-step RT-PCR assay, with high sensitivity and specificity, for the early detection of the Chikungunya virus (CHIKV). It uses 2,7-diamino-1,8-naphthyridine derivative (DANP)-labeled cytosine-bulge hairpin primers to amplify the nsP2 region of the CHIKV genome, followed by measurement of the fluorescence emitted from DANP-primer complexes after PCRs. The detection limit of our assay was 0.01 plaque-forming units per reaction of CHIKV. Furthermore, the HP-nsP2 primers were highly specific in detecting CHIKV, without any cross-reactivity with the panel of RNA viruses validated in this study. The feasibility of the DANP-coupled hairpin RT-PCR for clinical diagnosis was evaluated using clinical serum samples from CHIKV-infected patients, and the specificity and sensitivity were 100% (95% CI, 80.0% to 100%) and 95.5% (95% CI, 75.1% to 99.8%), respectively. These findings confirmed its potential as a point-of-care clinical molecular diagnostic assay for CHIKV in acute-phase patient serum samples. Copyright © 2013 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

Comparison of a new gold immunochromatographic assay for the rapid diagnosis of the novel influenza A (H7N9) virus with cell culture and a real-time reverse-transcription PCR assay.

PubMed

Jin, Changzhong; Wu, Nanping; Peng, Xiaorong; Yao, Hangping; Lu, Xiangyun; Chen, Yu; Wu, Haibo; Xie, Tiansheng; Cheng, Linfang; Liu, Fumin; Kang, Keren; Tang, Shixing; Li, Lanjuan

2014-01-01

We assessed a colloidal gold immunochromatographic assay (GICA) for rapid detection of influenza A (H7N9) and compared it with reverse-transcription-polymerase chain reaction (RT-PCR) and viral culture. Samples from 35 H7N9 infected patients were collected, including 45 throat swab samples, 56 sputum samples, and 39 feces samples. All samples were tested by GICA, viral culture, and RT-PCR. GICA specifically reacted with recombinant HA proteins, virus lysates, and clinical samples from H7 subtype viruses. Compared with RT-PCR, GICA demonstrated low sensitivity (33.33%) but high specificity (97.56%). The positive rate of GICA tests for samples collected in the period from 8 to 21 days after contact with poultry was much higher than those for samples collected before or after this period. Compared with viral culture, GICA showed sensitivity of 91.67% and specificity of 82.03%. Sputum specimens were more likely to test positive for H7N9 virus than samples from throat swabs and feces. The GICA-based H7 test is a reliable, rapid, and convenient method for the screening and diagnosis of influenza A (H7N9) disease, especially for the sputum specimens with high viral load. It may be helpful in managing H7N9 epidemics and preliminary diagnosis in early stages in resource-limited settings.

Comparison of a New Gold Immunochromatographic Assay for the Rapid Diagnosis of the Novel Influenza A (H7N9) Virus with Cell Culture and a Real-Time Reverse-Transcription PCR Assay

PubMed Central

Wu, Nanping; Peng, Xiaorong; Yao, Hangping; Lu, Xiangyun; Chen, Yu; Wu, Haibo; Xie, Tiansheng; Cheng, Linfang; Liu, Fumin; Kang, Keren; Tang, Shixing; Li, Lanjuan

2014-01-01

We assessed a colloidal gold immunochromatographic assay (GICA) for rapid detection of influenza A (H7N9) and compared it with reverse-transcription-polymerase chain reaction (RT-PCR) and viral culture. Samples from 35 H7N9 infected patients were collected, including 45 throat swab samples, 56 sputum samples, and 39 feces samples. All samples were tested by GICA, viral culture, and RT-PCR. GICA specifically reacted with recombinant HA proteins, virus lysates, and clinical samples from H7 subtype viruses. Compared with RT-PCR, GICA demonstrated low sensitivity (33.33%) but high specificity (97.56%). The positive rate of GICA tests for samples collected in the period from 8 to 21 days after contact with poultry was much higher than those for samples collected before or after this period. Compared with viral culture, GICA showed sensitivity of 91.67% and specificity of 82.03%. Sputum specimens were more likely to test positive for H7N9 virus than samples from throat swabs and feces. The GICA-based H7 test is a reliable, rapid, and convenient method for the screening and diagnosis of influenza A (H7N9) disease, especially for the sputum specimens with high viral load. It may be helpful in managing H7N9 epidemics and preliminary diagnosis in early stages in resource-limited settings. PMID:24822207

Changes in plasma chemistry after drug-induced liver disease or muscle necrosis in racing pigeons (Columba livia domestica).

PubMed

Lumeij, J T; Meidam, M; Wolfswinkel, J; Van der Hage, M H; Dorrestein, G M

1988-01-01

Changes in plasma variables as a result of liver damage induced by ethylene glycol (group A) or D-galactosamine (group B) and of muscle damage induced by doxycycline were compared. Plasma bile acid concentration was both a specific and a sensitive indicator of liver disease. Another specific, but less sensitive indicator of liver disease was 7-GT. Plasma AS AT activity was the most sensitive indicator of disease of the liver, but was not specific, since increased ASAT activities were also seen during muscle disease. ALAT activity was slightly more sensitive to liver damage than 7-GT, but was also not specific, being increased also after muscle damage. Plasma GLDH activity was increased only as a result of extensive liver necrosis. AP activity was of no value for detecting liver disease in the pigeon. CK activity was specific for muscle injury, though the activities of ALAT, ASAT and LD were also increased. Because of its long elimination half-life, increased ALAT activity persisted for 9 days after muscle damage, whereas CK activity returned to reference values within 3 days. LDH was a poor indicator of damage to liver and muscle, despite its relatively high tissue concentrations in both tissues. The rapid disappearance rate of LDH from plasma probably explains this observation.

Rapid diagnosis of pulmonary tuberculosis and detection of drug resistance by combined simultaneous amplification testing and reverse dot blot.

PubMed

Chen, Yiwen; Zhang, Lahong; Hong, Liquan; Luo, Xian; Chen, Juping; Tang, Leiming; Chen, Jiahuan; Liu, Xia; Chen, Zhaojun

2018-06-01

Making a correct and rapid diagnosis is essential for managing pulmonary tuberculosis (PTB), particularly multidrug-resistant tuberculosis. We aimed to evaluate the efficacy of the combination of simultaneous amplification testing (SAT) and reverse dot blot (RDB) for the rapid detection of Mycobacterium tuberculosis (MTB) and drug-resistant mutants in respiratory samples. 225 suspected PTB and 32 non-TB pulmonary disease samples were collected. All sputum samples were sent for acid-fast bacilli smear, SAT, culture and drug susceptibility testing (DST) by the BACTEC TM MGIT TM 960 system. 53 PTB samples were tested by both RDB and DNA sequencing to identify drug resistance genes and mutated sites. The SAT positive rate (64.9%) was higher than the culture positive rate (55.1%), with a coincidence rate of 83.7%. The sensitivity and specificity of SAT for diagnosing PTB were 66.7% and 100%, respectively, while those for culture were 53.9% and 84.2%, respectively. RDB has high sensitivity and specificity in identifying drug resistance genes and mutated sites. The results of RDB correlated well with those of DST and DNA sequencing, with coincidence rates of 92.5% and 98.1%, respectively. The combination of SAT and RDB is promising for rapidly detecting PTB and monitoring drug resistance in clinical laboratories. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

A rapid and ultrasensitive SERRS assay for histidine and tyrosine based on azo coupling.

PubMed

Sui, Huimin; Wang, Yue; Yu, Zhi; Cong, Qian; Han, Xiao Xia; Zhao, Bing

2016-10-01

A simple and highly sensitive surface-enhanced resonance Raman scattering (SERRS)-based approach coupled with azo coupling reaction has been put forward for quantitative analysis of histidine and tyrosine. The SERRS-based assay is simple and rapid by mixing the azo reaction products with silver nanoparticles (AgNPs) for measurements within 2min. The limits of detection (LODs) of the method are as low as 4.33×10(-11) and 8.80×10(-11)M for histidine and tyrosine, respectively. Moreover, the SERRS fingerprint information specific to corresponding amino acids guarantees the selective detection for the target histidine and tyrosine. The results from serum indicated the potential application of the proposed approach into biological samples. Compared with the methods ever reported, the main advantages of this methodology are simpleness, rapidity without time-consuming separation or pretreatment steps, high sensitivity, selectivity and the potential for determination of other molecules containing imidazole or phenol groups. Copyright © 2016 Elsevier B.V. All rights reserved.

Sensitivity to image recurrence across eye-movement-like image transitions through local serial inhibition in the retina

PubMed Central

Krishnamoorthy, Vidhyasankar; Weick, Michael; Gollisch, Tim

2017-01-01

Standard models of stimulus encoding in the retina postulate that image presentations activate neurons according to the increase of preferred contrast inside the receptive field. During natural vision, however, images do not arrive in isolation, but follow each other rapidly, separated by sudden gaze shifts. We here report that, contrary to standard models, specific ganglion cells in mouse retina are suppressed after a rapid image transition by changes in visual patterns across the transition, but respond with a distinct spike burst when the same pattern reappears. This sensitivity to image recurrence depends on opposing effects of glycinergic and GABAergic inhibition and can be explained by a circuit of local serial inhibition. Rapid image transitions thus trigger a mode of operation that differs from the processing of simpler stimuli and allows the retina to tag particular image parts or to detect transition types that lead to recurring stimulus patterns. DOI: http://dx.doi.org/10.7554/eLife.22431.001 PMID:28230526

Development of Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid and Sensitive Identification of Ostrich Meat

PubMed Central

Abdulmawjood, Amir; Grabowski, Nils; Fohler, Svenja; Kittler, Sophie; Nagengast, Helga; Klein, Guenter

2014-01-01

Animal species identification is one of the primary duties of official food control. Since ostrich meat is difficult to be differentiated macroscopically from beef, therefore new analytical methods are needed. To enforce labeling regulations for the authentication of ostrich meat, it might be of importance to develop and evaluate a rapid and reliable assay. In the present study, a loop-mediated isothermal amplification (LAMP) assay based on the cytochrome b gene of the mitochondrial DNA of the species Struthio camelus was developed. The LAMP assay was used in combination with a real-time fluorometer. The developed system allowed the detection of 0.01% ostrich meat products. In parallel, a direct swab method without nucleic acid extraction using the HYPLEX LPTV buffer was also evaluated. This rapid processing method allowed detection of ostrich meat without major incubation steps. In summary, the LAMP assay had excellent sensitivity and specificity for detecting ostrich meat and could provide a sampling-to-result identification-time of 15 to 20 minutes. PMID:24963709

Serum-Stable, Long-Circulating, pH-Sensitive PEGylated Liposomes.

PubMed

Bertrand, Nicolas; Simard, Pierre; Leroux, Jean-Christophe

2017-01-01

pH-sensitive liposomes have been designed to deliver active compounds, specifically to acidic intracellular organelles, and to augment their cytoplasmic concentrations. These systems combine the protective effects of other liposomal formulations with specific environment-controlled drug release. They are stable at physiological pH, but abruptly discharge their contents when endocytosed into acidic compartments, allowing the drug to be released before it is exposed to the harsh environment of the lysosomes.Serum-stable formulations with minimal leakage at physiological pH and rapid drug release at pH 5.0 to 5.5 can be easily prepared by inserting a hydrophobically modified N-isopropylacrylamide/methacrylic acid copolymer (poly(NIPAM-co-MAA)) in the lipid bilayer of sterically stabilized liposomes. The present chapter describes polymer synthesis, as well as the preparation and characterization of large unilamellar pH-sensitive vesicles.

Development of a rapid loop-mediated isothermal amplification assay for diagnosis and assessment of cure of Leishmania infection.

PubMed

Verma, Sandeep; Singh, Ruchi; Sharma, Vanila; Bumb, Ram Avtar; Negi, Narendra Singh; Ramesh, V; Salotra, Poonam

2017-03-23

Leishmaniasis is a spectrum of diseases with great relevance to public health. Conventional diagnostic methods are time consuming, needing trained personnel. A robust, rapid and cost effective diagnostic test is warranted for on-time diagnosis and field application. We have developed a loop mediated isothermal amplification (LAMP) assay with primers (n = 6) based on Leishmania donovani kDNA for detection of Leishmania infection, using a closed tube to prevent cross-contamination. The assay was used to detect Leishmania infection in biological samples obtained from patients of visceral leishmaniasis (VL), post kala-azar dermal leishmaniasis (PKDL) and cutaneous leishmaniasis (CL). The assay was positive for L. donovani, L. tropica and L. major parasites, with the highest sensitivity towards L. donovani (1 fg DNA). The high sensitivity of the assay for detection of L. donovani was reflected in its ability to detect parasite DNA within 30 min of amplification time with a threshold detection limit of ≥25 copies per reaction. The assay detected parasite in 64 of 66 VL blood samples (sensitivity, 96.9%; 95% CI: 89.6-99.2%), 15 of 15 VL bone marrow aspirate samples (sensitivity, 100%; 95% CI:79.6-100%), 65 of 67 PKDL tissue biopsy samples (sensitivity, 97%; 95% CI:89.7-99.2%). The assay was evaluated in a few cases of CL wherein it was found positive in 8 of 10 tissue biopsies (sensitivity, 80%; 95% CI: 49-94.3%). The assay was negative in all control blood (n = 76) and tissue biopsy (n = 24) samples (specificity, 100%; 95% CI: 96.3-100%). Further, the assay was evaluated for its utility in assessment of cure in treated VL and PKDL patients. The assay detected parasite DNA in 2 of 20VL blood samples and 2 of 21 PKDL tissue samples. Out of 4 cases that were positive for parasite DNA at post treatment stage, 2 patients (1VL and 1 PKDL) returned with relapse. The study demonstrated a Leishmania genus specific closed tube LAMP assay for reliable and rapid molecular diagnosis of VL and PKDL with potential for application in assessment of cure.

A biosensor platform for rapid detection of E. coli in drinking water.

PubMed

Hesari, Nikou; Alum, Absar; Elzein, Mohamad; Abbaszadegan, Morteza

2016-02-01

There remains a need for rapid, specific and sensitive assays for the detection of bacterial indicators for water quality monitoring. In this study, a strategy for rapid detection of Escherichia coli in drinking water has been developed. This strategy is based on the use of the substrate 4-methylumbelliferyl-β-d-glucuronide (MUG), which is hydrolyzed rapidly by the action of E. coli β-d-glucuronidase (GUD) enzyme to yield a fluorogenic 4-methylumbelliferone (4-MU) product that can be quantified and related to the number of E. coli cells present in water samples. In this study, the detection time required for the biosensor response ranged between 20 and 120 min, depending on the number of bacteria in the sample. This approach does not need extensive sample processing with a rapid detection capability. The specificity of the MUG substrate was examined in both, pure cultures of non-target bacterial genera such as Klebsiella, Salmonella, Enterobacter and Bacillus. Non-target substrates that included 4-methylumbelliferyl-β-d-galactopyranoside (MUGal) and l-leucine β-naphthylamide aminopeptidase (LLβ-N) were also investigated to identify nonspecific patterns of enzymatic activities in E. coli. GUD activity was found to be specific for E. coli and no further enzymatic activity was detected by other species. In addition, fluorescence assays were performed for the detection of E. coli to generate standard curves; and the sensitivity of the GUD enzymatic response was measured and repeatedly determined to be less than 10 E. coli cells in a reaction vial. The applicability of the method was tested by performing multiple fluorescence assays under pure and mixed bacterial flora in environmental samples. The results of this study showed that the fluorescence signals generated in samples using specific substrate molecules can be utilized to develop a bio-sensing platform for the detection of E. coli in drinking water. Furthermore, this system can be applied independently or in conjunction with other methods as a part of an array of biochemical assays in order to reliably detect E. coli in water. Copyright © 2015 Elsevier Inc. All rights reserved.

[Evaluation of Mascia Brunelli rapid antigen test in the diagnosis of group A streptococcal pharyngitis].

PubMed

Barış, Ayşe; Anlıaçık, Nur; Bulut, Mehmet Emin; Deniz, Rıdvan; Yücel, Elif; Aktaş, Elif

2017-01-01

Pharyngitis in most cases is due to viral microorganisms however drug therapy without the detection of etiological agent leads to unnecessary use of antibiotics. On the other hand, when the etiologic agent is group A beta-hemolytic streptococci (GAS) it is important to identify the etiologic agent rapidly which will guide the treatment with appropriate antibiotics. The use of highly sensitive rapid tests will contribute significantly to early diagnosis and appropriate therapy. The aim of this study is to evaluate the efficacy of Mascia Brunelli rapid antigen test for the detection of GAS in throat swab samples. A total of 833 throat swab samples submitted to our laboratory with pre-diagnosis of pharyngitis were assessed between June 2016 and August 2016. The samples were simultaneously cultured and tested by rapid Mascia Brunelli Strep-A Card (Mascia Brunelli S.p.a, Italy). For identification, bacitracin sensitivity, PYR test and latex agglutination test in addition to Bruker MALDI-TOF MS (Daltonics, Germany) system were used. The density of GAS growth in the culture was noted. The samples that were false negative with Mascia Brunelli test were re-tested with QuickVue + Strep A Test (Quidel Corporation, San Diego, USA) rapid antigen test. A total of 833 patients, 376 (45.2%) female and 457 (54.8%) male were included in the study. The age range was between 0-94 years with a mean value of 7.86 ± 6.72. 125 (15%) and 94 (11.28%) of the samples were positive with culture and rapid antigen test, respectively. Mascia Brunelli antigen test gave negative results for 31 culture positive samples. Of these 31 samples, 28 were found positive by QuickVue + Strep A antigen test. As a result, the sensitivity of the test was found to be independent of the inoculum effect. The culture positivity rate in patients between 5-15 years was 18.4%. The sensitivity, specificity, positive predictive value, negative predictive value and the accuracy of Mascia Brunelli antigen test, with respect to culture, were 75.2%, 100%, 100%, 95.81% and 96.28%, respectively. In conclusion, the selection of rapid antigen tests with high sensitivity in the diagnosis of GAS pharyngitis will contribute to the prevention of resistance development by appropriate use of antibiotics as well as early diagnosis and appropriate treatment. However, confirmation of negative rapid antigen test results by culture is very important in terms of false diagnosis and prevention of incomplete treatment.

Rapid detection of fungal keratitis with DNA-stabilizing FTA filter paper.

PubMed

Menassa, Nardine; Bosshard, Philipp P; Kaufmann, Claude; Grimm, Christian; Auffarth, Gerd U; Thiel, Michael A

2010-04-01

Purpose. Polymerase chain reaction (PCR) is increasingly important for the rapid detection of fungal keratitis. However, techniques of specimen collection and DNA extraction before PCR may interfere with test sensitivity. The purpose of this study was to investigate the use of DNA-stabilizing FTA filter paper (Indicating FTA filter paper; Whatman International, Ltd., Maidstone, UK) for specimen collection without DNA extraction in a single-step, nonnested PCR for fungal keratitis. Methods. Specimens were collected from ocular surfaces with FTA filter discs, which automatically lyse collected cells and stabilize nucleic acids. Filter discs were directly used in single-step PCR reactions to detect fungal DNA. Test sensitivity was evaluated with serial dilutions of Candida albicans, Fusarium oxysporum, and Aspergillus fumigatus cultures. Test specificity was analyzed by comparing 196 and 155 healthy individuals from Switzerland and Egypt, respectively, with 15 patients with a diagnosis of microbial keratitis. Results. PCR with filter discs detected 3 C. albicans, 25 F. oxysporum, and 125 A. fumigatus organisms. In healthy volunteers, fungal PCR was positive in 1.0% and 8.4% of eyes from Switzerland and Egypt, respectively. Fungal PCR remained negative in 10 cases of culture-proven bacterial keratitis, became positive in 4 cases of fungal keratitis, but missed 1 case of culture-proven A. fumigatus keratitis. Conclusions. FTA filter paper for specimen collection together with direct PCR is a promising method of detecting fungal keratitis. The analytical sensitivity is high without the need for a semi-nested or nested second PCR, the clinical specificity is 91.7% to 99.0%, and the method is rapid and inexpensive.

A rapid, highly accurate method for quantifying CALR mutant allele burden in persons with myeloproliferative neoplasms.

PubMed

Yao, Qiu-Mei; Zhou, Jiao; Gale, Robert Peter; Li, Jin-Lan; Li, Ling-Di; Li, Ning; Chen, Shan-Shan; Ruan, Guo-Rui

2015-10-01

Calreticulin (CALR) mutations were recently identified in a substantial proportion of persons with essential thrombocythemia (ET) and with primary myelofibrosis (PMF) without JAK2(V617F). Consequently rapid, sensitive, and specific methods to detect and quantify these mutations are needed. We studied samples from 1088 persons with myeloproliferative neoplasms (MPNs) including 421 JAK2(V617F) negative subjects with ET, PMF, polycythemia vera (PV), chronic myeloid leukemia (CML) and hyper-eosinophilic syndrome (HES). Detection of CALR exon 9 mutations was done by PCR amplification followed by fragment length analysis and direct sequencing. Dilution assays were used to determine CALR mutant allele burden. We detected CALR mutations in blood and bone marrow samples from 152 subjects with ET and with PMF but not in samples from normal or persons with PV, CML, or HES. CALR mutant peaks were distinct from wild-type peaks and dilution experiments indicated a sensitivity level of 0.5-5% for a CALR mutant allele in a wild-type background. Diverse types of mutations were detected including deletions, insertions, and complex indels. All mutations were confirmed by direct sequencing. We also used dilution experiments to quantify mutant allele burden. We were able to reproducibly detect mutant allele levels as low 5% (0.5-5%) in a wild-type background. PCR amplification followed by fragment length analysis is a rapid, sensitive, and specific method for screening persons with MPNs for CALR mutations, especially those with ET and PMF and for estimating mutant allele burden.

[Rapid Detection of Adenovirus in Fecal Samples by Capillary Electrophoresis-laser Induced Fluorescence and Microchip Capillary Electrophoresis-laser Induced Fluorescence].

PubMed

Ruan, Jia; Ren, Dong-xia; Yang, Dan-ni; Long, Pin-pin; Zhao, Hong-yue; Wang, Yi-qi; Li, Yong-xin

2015-07-01

To establish a rapid and sensitive method based on polymerase chain reaction (PCR) combined with capillary electrophoresis-laser induced fluorescence (CE-LIF) and microchip capillary electrophoresis-laser induced fluorescence (MCE-LIF) for detecting adenoviruses in fecal samples. The DNA of adenovirus in fecal samples were extracted by the commercial kits and the conserved region of hexon gene was selected as the target gene and amplified by PCR reaction. After labeling highly sensitive nucleic acid fluorescent dye SYBR Gold and SYBR Orange respectively, PCR amplification products were separated by CE and MCE under the optimized condition and detected by LIF detector. PCR amplification products could be detected within 9 min by CE-LIF and 6 min by MCE-LIF under the optimized separation condition. The sequenced PCR product showed good specificity in comparison with the prototype sequences from NCBI. The intraday and inter-day relative standard deviation (RSD) of the size (bp) of the target DNA was in the range of 1.14%-1.34% and 1.27%- 2.76%, respectively, for CE-LIF, and 1.18%-1.48% and 2.85%-4.06%, respectively, for MCE-LIF. The detection limits was 2.33 x 10(2) copies/mL for CE-LIF and 2.33 x 10(3) copies/mL for MCE-LIF. The two proposed methods were applied to detect fecal samples, both showing high accuracy. The two proposed methods of PCR-CE-LIF and PCR-MCE-LIF can detect adenovirus in fecal samples rapidly, sensitively and specifically.

Development of recombinase polymerase amplification assays for the rapid detection of peste des petits ruminants virus.

PubMed

Zhang, Yongning; Wang, Jianchang; Zhang, Zhou; Mei, Lin; Wang, Jinfeng; Wu, Shaoqiang; Lin, Xiangmei

2018-04-01

Peste des petits ruminants (PPR) is a severe infectious disease of small ruminants caused by PPR virus (PPRV). Rapid and sensitive detection of PPRV is critical for controlling PPR. This report describes the development and evaluation of a conventional reverse transcription recombinase polymerase amplification (RT-RPA) assay and a real-time RT-RPA assay, targeting the PPRV N gene. Sensitivity analysis revealed that the conventional RT-RPA assay could detect 852 copies of standard PPRV RNA per reaction at 95% probability within 20 min at 41 °C, and the real-time RT-RPA assay could detect 103 copies of RNA molecules per reaction at 95% probability. Specificity analysis showed that both assays have no cross-reactivity with nucleic acid templates prepared from other selected viruses or common pathogens. Clinical evaluation using 162 ovine and hircine serum and nasal swab samples showed that the performance of both the real-time RT-RPA assay and the conventional RT-RPA assay were comparable to that of real-time RT-PCR. The overall agreements between real-time RT-PCR and real-time RT-RPA, and conventional RT-RPA were 99.4% (161/162) and 98.8% (160/162), respectively. The R 2 value of real-time RT-RPA and real-time RT-PCR was 0.900 by linear regression analysis. Our results suggest that both RT-RPA assays have a potential application in the rapid, sensitive and specific detection of PPRV. Copyright © 2018 Elsevier B.V. All rights reserved.

Comparison of Rapid Point-of-Care Tests for Detection of Antibodies to Hepatitis C Virus

PubMed Central

Fisher, Dennis G.; Hess, Kristen L.; Erlyana, Erlyana; Reynolds, Grace L.; Cummins, Catherine A.; Alonzo, Todd A.

2015-01-01

Background. Hepatitis C is one of the most prevalent blood-borne diseases in the United States. Despite the benefits of early screening, among 3.2 million Americans who are infected with hepatitis C virus (HCV), 50%–70% are unaware of their infection status. Methods. Data were collected between 2011 and 2014, from 1048 clients who were in the following groups: (1) injection drug users, (2) women at sexual risk, (3) gay and bisexual men, and (4) transgender individuals. The sensitivity and specificity of point-of-care tests included (1) the MedMira rapid human immunodeficiency virus (HIV)/HCV antibody test, (2) MedMira hepatitis B (HBV)/HIV/HCV antibody test, (3) Chembio HCV Screen Assay used with both whole blood and (4) oral specimens, (5) Chembio HIV-HCV Assay also used with both whole blood and (6) oral specimens, (7) Chembio HIV-HCV-Syphilis Assay, and (8) OraSure HCV Rapid Antibody Test used with whole blood. The gold standard for the HCV tests were HCV enzyme immunoassay (EIA) 2.0. Results. OraSure had the highest sensitivity at 92.7% (95% confidence interval [CI] = 88.8%–96.5%) followed closely by Chembio's 3 blood tests at 92.1% (95% CI = 87.7%–96.4%), 91.5% (95% CI = 87.2%–95.7%), and 92.3% (95% CI = 88.4%–96.2%). The sensitivities of MedMira HIV/HCV and MedMira HIV/HCV/HBV tests were the lowest, at 79.1% (95% CI = 72.6%–85.5%), and 81.5% (95% CI = 75.2%–87.8%), respectively. Specificity for the OraSure was 99.8% (95% CI = 99.4%–100%); specificity for the Chembio blood tests was 99.2% (95% CI = 98.6%–99.9%), 99.4% (95% CI = 98.8%–99.9%), and 99.3% (95% CI = 98.8%–99.9%); and specificity for the MedMira was100% and 100%. False-negative results were associated with HIV and hepatitis B core antibody serostatus. Conclusions. The OraSure and Chembio blood tests (including those multiplexed with HIV and syphilis) appear to good performance characteristics. This study has identified potential limitations of rapid testing in those testing positive for HIV and HBcAb. There should be discussion of updates to the 2013 CDC guidance. PMID:26269795

Highly sensitive label-free dual sensor array for rapid detection of wound bacteria.

PubMed

Sheybani, Roya; Shukla, Anita

2017-06-15

Wound infections are a critical healthcare concern worldwide. Rapid and effective antibiotic treatments that can mitigate infection severity and prevent the spread of antibiotic resistance are contingent upon timely infection detection. In this work, dual electrochemical pH and cell-attachment sensor arrays were developed for the real-time spatial and temporal monitoring of potential wound infections. Biocompatible polymeric device coatings were integrated to stabilize the sensors and promote bacteria attachment while preventing non-specific cell and protein fouling. High sensitivity (bacteria concentration of 10 2 colony forming units (CFU)/mL and -88.1±6.3mV/pH over a pH range of 1-13) and stability over 14 days were achieved without the addition of biological recognition elements. The dual sensor array was demonstrated to successfully monitor the growth of both gram-positive (Staphylococcus aureus and Streptococcus pyogenes) and gram-negative bacteria (Pseudomonas aeruginosa and Escherichia coli) over time through lag and log growth phases and following antibiotic administration and in simulated shallow wounds conditions. The versatile fabrication methods utilized in sensor development, superior sensitivity, prolonged stability, and lack of non-specific sensor fouling may enable long-term in situ sensor array operation in low resource settings. Copyright © 2016 Elsevier B.V. All rights reserved.

Performance comparison of CareStart™ HRP2/pLDH combo rapid malaria test with light microscopy in north-western Tigray, Ethiopia: a cross-sectional study.

PubMed

Feleke, Daniel Getacher; Tarko, Shambel; Hadush, Haftom

2017-06-06

Rapid diagnostic tests (RDTs) are alternative methods for microscopy in the diagnosis of malaria in resource limited settings. Among commercially available RDTs, CareStart™ Malaria test was found to show reliable results. This study evaluated the performance of CareStart™ Malaria Combo test kit in Northwestern Tigray in Ethiopia. Blood samples were collected from 320 malaria-suspected patients at Mayani Hospital in Northwestern Tigray from December 2015 to March 2016. All blood samples were examined using both light microscopy and CareStart™ Malaria HRP2/pLDH Combo Test kit. Statistical analyses were performed using SPSS version 20. The overall parasite positivity using light microscopy and CareStart™ RDT was 41 (12.8%) and 43 (13.4%), respectively. The sensitivity and specificity of CareStart™ RDT, regardless of species, were found to be 95.4 and 99.3%, respectively. Furthermore, the sensitivity of CareStart™ RDT for Plasmodium falciparum or mixed infection and non-falciparum malaria parasites was 94.4 and 85.0%, respectively while the specificity was found to be 98.9 and 99.7%, respectively. The agreement between the two test methods was "excellent" with a kappa value of 0.92. CareStart™ RDT has very good sensitivity and specificity for malaria diagnosis. The test kit also has an excellent agreement with light microscopy. It is therefore useful in resource-limited areas where microscopy is not available.

Evaluation of the Clearview® malaria pLDH malaria rapid diagnostic test in a non-endemic setting

PubMed Central

2011-01-01

Background Malaria Rapid Diagnostic Tests (RDTs) are widely used to diagnose malaria. The present study evaluated a new RDT, the Clearview® Malaria pLDH test targeting the pan-Plasmodium antigen lactate dehydrogenase (pLDH). Methods The Clearview® Malaria pLDH test was evaluated on fresh samples obtained in returned international travellers using microscopy corrected by PCR as the reference method. Included samples were Plasmodium falciparum (139), Plasmodium vivax (22), Plasmodium ovale (20), Plasmodium malariae (7), and 102 negative. Results Overall sensitivity for the detection of Plasmodium spp was 93.2%. For P. falciparum, the sensitivity was 98.6%; for P. vivax, P. ovale and P. malariae, overall sensitivities were 90.9%, 60.0% and 85.7% respectively. For P. falciparum and for P. vivax, the sensitivities increased to 100% at parasite densities above 100/μl. The specificity was 100%. The test was easily to perform and the result was stable for at least 1 hour. Conclusion The Clearview® Malaria pLDH was efficient for the diagnosis of malaria. The test was very sensitive for P. falciparum and P. vivax detection. The sensitivities for P. ovale and P. malariae were better than other RDTs PMID:21951996

Evaluation of the Rapid Polymyxin NP Test for Polymyxin B Resistance Detection Using Enterobacter cloacae and Enterobacter aerogenes Isolates.

PubMed

Simar, Shelby; Sibley, Diane; Ashcraft, Deborah; Pankey, George

2017-10-01

Polymyxin resistance is an increasing problem worldwide. Currently, determining susceptibility to polymyxins is problematic and lengthy. Polymyxins diffuse poorly into agar, potentially giving inaccurate disk diffusion and Etest results. A rapid screening test (2 h) for the detection of polymyxin resistance in Enterobacteriaceae , developed by P. Nordmann and L. Poirel (rapid polymyxin NP test) in 2016, detects glucose metabolization in the presence of polymyxin E (PE) and PB via pH-induced color change. The sensitivity and specificity were 99.3 and 95.4%, respectively, with results obtained in ≤2 h. Our goal was to evaluate this test using PB against larger numbers of Enterobacter A total of 143 nonduplicate Enterobacter isolates (102 E. cloacae complex, 41 E. aerogenes ) were tested, including 136 collected from Ochsner Health System patients from March to May 2016 and 7 previously determined PB-resistant E. cloacae isolates from JMI Laboratories. MICs were determined via broth microdilution. For the rapid polymyxin NP test, a color change from orange to yellow is positive; a weak/no color change is deemed negative after 4 h. Of 143 Enterobacter isolates, 25 were determined to be PB resistant by broth microdilution (MIC > 2 μg/ml), including all 7 JMI isolates. Of these 25, 7 were positive by the rapid polymyxin NP test (included 3/7 JMI isolates). All 118 isolates determined to be PB susceptible by broth microdilution were NP test negative. The sensitivity and specificity for the rapid polymyxin NP test were 25 and 100%, respectively, compared to broth microdilution. Although the rapid polymyxin NP test is a much faster method (2 to 4 h) for polymyxin resistance determination compared to broth microdilution (16 to 20 h), our study indicates that it may be subject to limitations when testing Enterobacter . Copyright © 2017 American Society for Microbiology.

Liver Rapid Reference Set Application (#1): David Lubman - Univ of Michigan (2009) — EDRN Public Portal

Cancer.gov

We are requesting the reference set, which includes 50 HCC cases and 50 cirrhotic controls. In our preliminary study, AFP had a AUROC of 0.66 while the AUROC for the 5 glycoproteins was 0.81. The sensitivity and specificity for the 5 glycoproteins was 79% and 72% at the point that maximizes sensitivity+specificity in the ROC curve, and it was 79% and 35%, respectively, for AFP at the same point in the ROC curve. The reference set will allow us to determine the best performance of the 5 glycoproteins by themselves or whether their combination has a better sensitivity and/or specificity and AUROC. While a direct comparison with AFP will be made, the reference set will not allow a robust comparison due to the low sample size. If the glycoproteins are complementary or have better performance than AFP, then the next step would be to test them in the entire phase 2 hepatocellular carcinoma set.

Liver Rapid Reference Set Application ( #2): Lubman - Univ of Michigan (2010) — EDRN Public Portal

Cancer.gov

We are requesting the reference set, which includes 50 HCC cases and 50 cirrhotic controls. In our preliminary study, AFP had a AUROC of 0.66 while the AUROC for the 5 glycoproteins was 0.81. The sensitivity and specificity for the 5 glycoproteins was 79% and 72% at the point that maximizes sensitivity+specificity in the ROC curve, and it was 79% and 35%, respectively, for AFP at the same point in the ROC curve. The reference set will allow us to determine the best performance of the 5 glycoproteins by themselves or whether their combination has a better sensitivity and/or specificity and AUROC. While a direct comparison with AFP will be made, the reference set will not allow a robust comparison due to the low sample size. If the glycoproteins are complementary or have better performance than AFP, then the next step would be to test them in the entire phase 2 hepatocellular carcinoma set.

Comparison of real-time PCR methods for the detection of Naegleria fowleri in surface water and sediment.

PubMed

Streby, Ashleigh; Mull, Bonnie J; Levy, Karen; Hill, Vincent R

2015-05-01

Naegleria fowleri is a thermophilic free-living ameba found in freshwater environments worldwide. It is the cause of a rare but potentially fatal disease in humans known as primary amebic meningoencephalitis. Established N. fowleri detection methods rely on conventional culture techniques and morphological examination followed by molecular testing. Multiple alternative real-time PCR assays have been published for rapid detection of Naegleria spp. and N. fowleri. Foursuch assays were evaluated for the detection of N. fowleri from surface water and sediment. The assays were compared for thermodynamic stability, analytical sensitivity and specificity, detection limits, humic acid inhibition effects, and performance with seeded environmental matrices. Twenty-one ameba isolates were included in the DNA panel used for analytical sensitivity and specificity analyses. N. fowleri genotypes I and III were used for method performance testing. Two of the real-time PCR assays were determined to yield similar performance data for specificity and sensitivity for detecting N. fowleri in environmental matrices.

Comparison of real-time PCR methods for the detection of Naegleria fowleri in surface water and sediment

PubMed Central

Streby, Ashleigh; Mull, Bonnie J.; Levy, Karen

2015-01-01

Naegleria fowleri is a thermophilic free-living ameba found in freshwater environments worldwide. It is the cause of a rare but potentially fatal disease in humans known as primary amebic meningoencephalitis. Established N. fowleri detection methods rely on conventional culture techniques and morphological examination followed by molecular testing. Multiple alternative real-time PCR assays have been published for rapid detection of Naegleria spp. and N. fowleri. Four such assays were evaluated for the detection of N. fowleri from surface water and sediment. The assays were compared for thermodynamic stability, analytical sensitivity and specificity, detection limits, humic acid inhibition effects, and performance with seeded environmental matrices. Twenty-one ameba isolates were included in the DNA panel used for analytical sensitivity and specificity analyses. N. fowleri genotypes I and III were used for method performance testing. Two of the real-time PCR assays were determined to yield similar performance data for specificity and sensitivity for detecting N. fowleri in environmental matrices. PMID:25855343

Bacteriophage-based assays for the rapid detection of rifampicin resistance in Mycobacterium tuberculosis: a meta-analysis.

PubMed

Pai, Madhukar; Kalantri, Shriprakash; Pascopella, Lisa; Riley, Lee W; Reingold, Arthur L

2005-10-01

To summarize, using meta-analysis, the accuracy of bacteriophage-based assays for the detection of rifampicin resistance in Mycobacterium tuberculosis. By searching multiple databases and sources we identified a total of 21 studies eligible for meta-analysis. Of these, 14 studies used phage amplification assays (including eight studies on the commercial FASTPlaque-TB kits), and seven used luciferase reporter phage (LRP) assays. Sensitivity, specificity, and agreement between phage assay and reference standard (e.g. agar proportion method or BACTEC 460) results were the main outcomes of interest. When performed on culture isolates (N=19 studies), phage assays appear to have relatively high sensitivity and specificity. Eleven of 19 (58%) studies reported sensitivity and specificity estimates > or =95%, and 13 of 19 (68%) studies reported > or =95% agreement with reference standard results. Specificity estimates were slightly lower and more variable than sensitivity; 5 of 19 (26%) studies reported specificity <90%. Only two studies performed phage assays directly on sputum specimens; although one study reported sensitivity and specificity of 100 and 99%, respectively, another reported sensitivity of 86% and specificity of 73%. Current evidence is largely restricted to the use of phage assays for the detection of rifampicin resistance in culture isolates. When used on culture isolates, these assays appear to have high sensitivity, but variable and slightly lower specificity. In contrast, evidence is lacking on the accuracy of these assays when they are directly applied to sputum specimens. If phage-based assays can be directly used on clinical specimens and if they are shown to have high accuracy, they have the potential to improve the diagnosis of MDR-TB. However, before phage assays can be successfully used in routine practice, several concerns have to be addressed, including unexplained false positives in some studies, potential for contamination and indeterminate results.

Detection of prostate-specific antigen with biomolecule-gated AlGaN/GaN high electron mobility transistors

NASA Astrophysics Data System (ADS)

Li, Jia-dong; Cheng, Jun-jie; Miao, Bin; Wei, Xiao-wei; Xie, Jie; Zhang, Jin-cheng; Zhang, Zhi-qiang; Wu, Dong-min

2014-07-01

In order to improve the sensitivity of AlGaN/GaN high electron mobility transistor (HEMT) biosensors, a simple biomolecule-gated AlGaN/GaN HEMT structure was designed and successfully fabricated for prostate specific antigen (PSA) detection. UV/ozone was used to oxidize the GaN surface and then a 3-aminopropyl trimethoxysilane (APTES) self-assembled monolayer was bound to the sensing region. This monolayer serves as a binding layer for attachment of the prostate specific antibody (anti-PSA). The biomolecule-gated AlGaN/GaN HEMT sensor shows a rapid and sensitive response when the target prostate-specific antigen in buffer solution was added to the antibody-immobilized sensing area. The current change showed a logarithm relationship against the PSA concentration from 0.1 pg/ml to 0.993 ng/ml. The sensitivity of 0.215% is determined for 0.1 pg/ml PSA solution. The above experimental result of the biomolecule-gated AlGaN/GaN HEMT biosensor suggested that this biosensor might be a useful tool for prostate cancer screening.

Bacteriophage Amplification-Coupled Detection and Identification of Bacterial Pathogens

NASA Astrophysics Data System (ADS)

Cox, Christopher R.; Voorhees, Kent J.

Current methods of species-specific bacterial detection and identification are complex, time-consuming, and often require expensive specialized equipment and highly trained personnel. Numerous biochemical and genotypic identification methods have been applied to bacterial characterization, but all rely on tedious microbiological culturing practices and/or costly sequencing protocols which render them impractical for deployment as rapid, cost-effective point-of-care or field detection and identification methods. With a view towards addressing these shortcomings, we have exploited the evolutionarily conserved interactions between a bacteriophage (phage) and its bacterial host to develop species-specific detection methods. Phage amplification-coupled matrix assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF-MS) was utilized to rapidly detect phage propagation resulting from species-specific in vitro bacterial infection. This novel signal amplification method allowed for bacterial detection and identification in as little as 2 h, and when combined with disulfide bond reduction methods developed in our laboratory to enhance MALDI-TOF-MS resolution, was observed to lower the limit of detection by several orders of magnitude over conventional spectroscopy and phage typing methods. Phage amplification has been combined with lateral flow immunochromatography (LFI) to develop rapid, easy-to-operate, portable, species-specific point-of-care (POC) detection devices. Prototype LFI detectors have been developed and characterized for Yersinia pestis and Bacillus anthracis, the etiologic agents of plague and anthrax, respectively. Comparable sensitivity and rapidity was observed when phage amplification was adapted to a species-specific handheld LFI detector, thus allowing for rapid, simple, POC bacterial detection and identification while eliminating the need for bacterial culturing or DNA isolation and amplification techniques.

Utility of the Determine Syphilis TP rapid test in commercial sex venues in Peru.

PubMed

Campos, P E; Buffardi, A L; Chiappe, M; Buendía, C; Garcia, P J; Carcamo, C P; Garnett, G; White, P; Holmes, K K

2006-12-01

This study sought to evaluate the utility of the Determine Syphilis TP test performed in Peruvian commercial sex venues for the detection of active syphilis; and determine the feasibility of integrating rapid syphilis testing for female sex workers (FSW) into existing health outreach services. We tested 3586 female sex workers for syphilis by Determine in the field using whole blood fingerstick, and by rapid plasma reagin (RPR) and Treponema pallidum haemagglutination assay (TPHA) in a central laboratory in Lima using sera. 97.4% of the FSW offered rapid syphilis testing participated; and among those who tested positive, 87% visited the local health centre for treatment. More than twice as many specimens were RPR reactive using serum in Lima (5.7%) than tested positive by whole blood Determine in the field (2.8%), and although most were confirmed by TPHA, only a small proportion (0.7%) were RPR reactive at >or=1:8 dilutions, and likely indicating active syphilis. Sensitivity, specificity and positive predictive value of the Determine Syphilis TP test in whole blood when compared to serum RPR reactivity at any dilution confirmed by TPHA as the gold standard were 39.3%, 99.2% and 71.4%, respectively. Sensitivity improved to 64.0% when using serum RPR >or=1:8 confirmed by TPHA. Invalid tests were rare (0.3%). Rapid syphilis testing in sex work venues proved feasible, but Determine using whole blood obtained by fingerstick was substantially less sensitive than reported in previous laboratory-based studies using serum. Although easy to perform in outreach venues, the utility of this rapid syphilis test was relatively low in settings where a large proportion of the targeted population has been previously tested and treated.

Utility of the Determine Syphilis TP rapid test in commercial sex venues in Peru

PubMed Central

Campos, P E; Buffardi, A L; Chiappe, M; Buendía, C; Garcia, P J; Carcamo, C P; Garnett, G; White, P

2006-01-01

Objectives This study sought to evaluate the utility of the Determine Syphilis TP test performed in Peruvian commercial sex venues for the detection of active syphilis; and determine the feasibility of integrating rapid syphilis testing for female sex workers (FSW) into existing health outreach services. Methods We tested 3586 female sex workers for syphilis by Determine in the field using whole blood fingerstick, and by rapid plasma reagin (RPR) and Treponema pallidum haemagglutination assay (TPHA) in a central laboratory in Lima using sera. Results 97.4% of the FSW offered rapid syphilis testing participated; and among those who tested positive, 87% visited the local health centre for treatment. More than twice as many specimens were RPR reactive using serum in Lima (5.7%) than tested positive by whole blood Determine in the field (2.8%), and although most were confirmed by TPHA, only a small proportion (0.7%) were RPR reactive at ⩾1:8 dilutions, and likely indicating active syphilis. Sensitivity, specificity and positive predictive value of the Determine Syphilis TP test in whole blood when compared to serum RPR reactivity at any dilution confirmed by TPHA as the gold standard were 39.3%, 99.2% and 71.4%, respectively. Sensitivity improved to 64.0% when using serum RPR ⩾1:8 confirmed by TPHA. Invalid tests were rare (0.3%). Conclusions Rapid syphilis testing in sex work venues proved feasible, but Determine using whole blood obtained by fingerstick was substantially less sensitive than reported in previous laboratory‐based studies using serum. Although easy to perform in outreach venues, the utility of this rapid syphilis test was relatively low in settings where a large proportion of the targeted population has been previously tested and treated. PMID:17116642

Rapid and visual detection of Leptospira in urine by LigB-LAMP assay with pre-addition of dye.

PubMed

Ali, Syed Atif; Kaur, Gurpreet; Boby, Nongthombam; Sabarinath, T; Solanki, Khushal; Pal, Dheeraj; Chaudhuri, Pallab

2017-12-01

Leptospirosis is considered to be the most widespread zoonotic disease caused by pathogenic species of Leptospira. The present study reports a novel set of primers targeting LigB gene for visual detection of pathogenic Leptospira in urine samples through Loop-mediated isothermal amplification (LAMP). The results were recorded by using Hydroxyl napthol blue (HNB), SYBR GREEN I and calcein. Analytical sensitivity of LAMP was as few as 10 leptospiral organisms in spiked urine samples from cattle and dog. LigB gene based LAMP, termed as LigB-LAMP, was found 10 times more sensitive than conventional PCR. The diagnostic specificity of LAMP was 100% when compared to SYBR green qPCR for detection of Leptospira in urine samples. Though qPCR was found more sensitive, the rapidity and simplicity in setting LAMP test followed by visual detection of Leptospira infection in clinical samples makes LigB-LAMP an alternative and favourable diagnostic tool in resource poor setting. Copyright © 2017 Elsevier Ltd. All rights reserved.

A rapid, ratiometric, enzyme-free, and sensitive single-step miRNA detection using three-way junction based FRET probes

NASA Astrophysics Data System (ADS)

Luo, Qingying; Liu, Lin; Yang, Cai; Yuan, Jing; Feng, Hongtao; Chen, Yan; Zhao, Peng; Yu, Zhiqiang; Jin, Zongwen

2018-03-01

MicroRNAs (miRNAs) are single stranded endogenous molecules composed of only 18-24 nucleotides which are critical for gene expression regulating the translation of messenger RNAs. Conventional methods based on enzyme-assisted nucleic acid amplification techniques have many problems, such as easy contamination, high cost, susceptibility to false amplification, and tendency to have sequence mismatches. Here we report a rapid, ratiometric, enzyme-free, sensitive, and highly selective single-step miRNA detection using three-way junction assembled (or self-assembled) FRET probes. The developed strategy can be operated within the linear range from subnanomolar to hundred nanomolar concentrations of miRNAs. In comparison with the traditional approaches, our method showed high sensitivity for the miRNA detection and extreme selectivity for the efficient discrimination of single-base mismatches. The results reveal that the strategy paved a new avenue for the design of novel highly specific probes applicable in diagnostics and potentially in microscopic imaging of miRNAs in real biological environments.

Detection of Echinoderm Microtubule Associated Protein Like 4-Anaplastic Lymphoma Kinase Fusion Genes in Non-small Cell Lung Cancer Clinical Samples by a Real-time Quantitative Reverse Transcription Polymerase Chain Reaction Method.

PubMed

Zhao, Jing; Zhao, Jin-Yin; Chen, Zhi-Xia; Zhong, Wei; Li, Long-Yun; Liu, Li-Cheng; Hu, Xiao-Xu; Chen, Wei-Jun; Wang, Meng-Zhao

2016-12-20

Objective To establish a real-time quantitative reverse transcription polymerase chain reaction assay (qRT-PCR) for the rapid, sensitive, and specific detection of echinoderm microtubule associated protein like 4-anaplastic lymphoma kinase (EML4-ALK) fusion genes in non-small cell lung cancer. Methods The specific primers for the four variants of EML4-ALK fusion genes (V1, V2, V3a, and V3b) and Taqman fluorescence probes for the detection of the target sequences were carefully designed by the Primer Premier 5.0 software. Then, using pseudovirus containing EML4-ALK fusion genes variants (V1, V2, V3a, and V3b) as the study objects, we further analyzed the lower limit, sensitivity, and specificity of this method. Finally, 50 clinical samples, including 3 ALK-fluorescence in situ hybridization (FISH) positive specimens, were collected and used to detect EML4-ALK fusion genes using this method. Results The lower limit of this method for the detection of EML4-ALK fusion genes was 10 copies/μl if no interference of background RNA existed. Regarding the method's sensitivity, the detection resolution was as high as 1% and 0.5% in the background of 500 and 5000 copies/μl wild-type ALK gene, respectively. Regarding the method's specificity, no non-specific amplification was found when it was used to detect EML4-ALK fusion genes in leukocyte and plasma RNA samples from healthy volunteers. Among the 50 clinical samples, 47 ALK-FISH negative samples were also negative. Among 3 ALK-FISH positive samples, 2 cases were detected positive using this method, but another was not detected because of the failure of RNA extraction. Conclusion The proposed qRT-PCR assay for the detection of EML4-ALK fusion genes is rapid, simple, sensitive, and specific, which is deserved to be validated and widely used in clinical settings.

Loop-mediated isothermal amplification for detection of the tomato and potato late blight pathogen, Phytophthora infestans

USDA-ARS?s Scientific Manuscript database

Aims: To design and validate a colorimetric loop-mediated isothermal amplification assay for rapid detection of P. infestans DNA. Methods and Results: Two sets of LAMP primers were designed and evaluated for their sensitivity and specificity for P. infestans. ITSII primers targeted a portion of the ...

A rapid spectrophotometric blood cyanide determination applicable to emergency toxicology.

PubMed

Laforge, M; Buneaux, F; Houeto, P; Bourgeois, F; Bourdon, R; Levillain, P

1994-01-01

Cyanide determination in whole blood can be performed by spectrophotometry after using diffusion coupled with coloration by hydroxocobalamin in a Conway dish. The technique may be accelerated by the use of a heating sheet at 45 degrees C. The method proved to be specific, sensitive, and fast, thus permitting measurements in emergency situations.

Development of one-step Loop-Mediated Isothermal Amplification (LAMP) for the detection of norovirus in oysters

USDA-ARS?s Scientific Manuscript database

The aim of this study was to develop a simple and rapid technique for detecting human norovirus (NoV). The loop-mediated isothermal amplification (LAMP) technique was evaluated and found to be sensitive, highly specific, and useful for routine oyster testing. Reverse transcription-LAMP (RT-LAMP) pri...

Avian influenza virus detection and quantitation by real-time RT-PCR

USDA-ARS?s Scientific Manuscript database

Real-time RT-PCR (rRT-PCR) has been used for avian influenza virus (AIV) detection since the early 2000’s for routine surveillance, during outbreaks and for research. Some of the advantages of rRT-PCR are: high sensitivity, high specificity, rapid time-to-result, scalability, cost, and its inherentl...

Fabrication of a novel transparent SERS substrate comprised of Ag-nanoparticle arrays and its application in rapid detection of ractopamine on meat

USDA-ARS?s Scientific Manuscript database

Surface-enhanced Raman spectroscopy (SERS) is an emerging analytical tool that boasts the feature of high detection sensitivity and molecular fingerprint specificity attracting increased attention and showing promise in applications including detecting residues of veterinary drugs. In practice, spec...

Comparison of Directigen Group A Strep Test with a traditional culture technique for detection of group A beta-hemolytic streptococci.

PubMed Central

McCusker, J J; McCoy, E L; Young, C L; Alamares, R; Hirsch, L S

1984-01-01

The Directigen Group A Strep Test (DGAST), a new rapid method of detecting group A beta-hemolytic streptococci directly from throat swabs, was compared with a traditional culture technique for the detection of group A beta-hemolytic streptococci. Five hundred oropharyngeal swabs from pediatric and adult patients were cultured and then processed by using the DGAST. Of the 144 specimens positive by culture, 131 were DGAST positive (sensitivity, 90.9%). Of the 356 specimens negative by culture, 353 were DGAST negative (specificity, 99.2%). Twelve of the 13 false-negative DGAST results were from pediatric patients. One hundred isolates of non-group A beta-hemolytic streptococci were recovered, primarily groups C, F, and G. The DGAST is easy to perform, rapid, sensitive, and very specific for detection of group A beta-hemolytic streptococci directly from swabs. Supplementing the DGAST with a culture on a 5% sheep blood agar plate would enhance detection of group A beta-hemolytic streptococci, especially in pediatric patients. PMID:6386884

Performance of an HRP-2 Rapid Diagnostic Test in Nigerian Children Less Than 5 Years of Age

PubMed Central

Ajumobi, Olufemi; Sabitu, Kabir; Nguku, Patrick; Kwaga, Jacob; Ntadom, Godwin; Gitta, Sheba; Elizeus, Rutebemberwa; Oyibo, Wellington; Nsubuga, Peter; Maire, Mark; Poggensee, Gabriele

2015-01-01

The diagnostic performance of histidine-rich protein 2 (HRP-2)–based malaria rapid diagnostic test (RDT) was evaluated in a mesoendemic area for malaria, Kaduna, Nigeria. We compared RDT results with expert microscopy results of blood samples from 295 febrile children under 5 years. Overall, 11.9% (35/295) tested positive with RDT compared with 10.5% (31/295) by microscopy: sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were 100%, 98.5%, 88.6%, and 100%, respectively. The RDT sensitivity was not affected by transmission season, parasite density, and age. Specificity and positive PV decreased slightly during the high-transmission season (97.5% and 83.3%). The RDT test positivity rates in the low- and high-transmission seasons were 9.4% and 13.5%, respectively. Overall, the test performance of this RDT was satisfactory. The findings of a low proportion of RDT false positives, no invalid and no false-negative results should validate the performance of RDTs in this context. PMID:25711608

Evaluation of 9 rapid diagnostic tests for screening HIV infection, in Lomé, Togo.

PubMed

Dagnra, A Y; Dossim, S; Salou, M; Nyasenu, T; Ali-Edje, K; Ouro-Médeli, A; Doufan, M; Ehlan, A; Prince-David, M

2014-12-01

HIV rapid diagnostic tests (RDT) could be greatly contributive for a universal access to HIV diagnosis. However, according to the WHO, these tests need to be assessed before they can be used in routine. We assessed 9 RDT in routine clinical use between 2009 and 2013. The sensitivity and specificity observed for 7 tests were≥99% and≥98%, respectively: FIRST RESPONSE HIV1-2-O PMC Medical, India, GENIE Fast HIV 1-2 and GENIE™ III HIV(1/2) Bio-Rad, France, HIV TRI-DOT+Ag;J. Mitra, INDIA; SD BIOLINE HIV(1/2) 3.0 and SD BIOLINE HIV/SYPHILIS DUO Standard Diagnostic, Korea; and VIKIA HIV(1/2); BioMérieux, France. Two tests had performances inferior to WHO recommendations: INSTI HIV1/2 Biolytical Canada; sensitivity=97.8% and HEXAGON HIV HUMAN GmbH Germany; specificity=94.8%. Seven of 9 RDT had excellent performances. Nevertheless, they can be used only after training staff, and taking into account national algorithm for their safe use. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Development of a polymerase chain reaction assay for the detection of pseudorabies virus in clinical samples

PubMed Central

Pérez, Lester J.; de Arce, Heidy Díaz

2009-01-01

Aujeszky’s disease, also known as pseudorabies causes severe economic losses in swine industry and affects the pig husbandry all over the world. The conventional diagnostic procedure is time-consuming and false-negative results may occur in submissions from latently infected animals. The development, optimization and evaluation of a polymerase chain reaction (PCR) assay are presented for the diagnosis of pseudorabies infection. This assay was based on the amplification of a highly conserved viral gD gene fragment. PCR products of the expected size were obtained from PRV strains. Non-specific reactions were not observed when a related herpesvirus, other porcine DNA genome viruses and uninfected cells were used to assess PCR. The analytical sensitivity of the test was estimated to be 1.34 TCID50/ 50 uL. The analysis of tissue homogenate samples from naturally infected animals proved the potential usefulness of the method for a rapid disease diagnosis from field cases. A rapid, sensitive and specific PCR-based diagnostic assay to detect pseudorabies virus in clinical samples is provided. PMID:24031383

A colorimetric and fluorescent probe for detecting intracellular biothiols.

PubMed

Chen, Chunyang; Liu, Wei; Xu, Cong; Liu, Weisheng

2016-11-15

A new rapid and highly sensitive coumarin-based probe (probe 1) has been designed and synthesized for detecting intracellular thiols. Probe 1 was prepared by a 4-step procedure as a latent fluorescence probe to achieve high sensitivity and fluorescence turn-on response toward cysteine and homocysteine over GSH and other various natural amino acids under physiological conditions. Owing to specific cyclization between thiols and aldehyde group, probe 1 displayed a highly selectivity toward cysteine and homocysteine. Above all, probe 1 was successfully used for fluorescence imaging of biothiols in Hela cells, and quantitative determination had been achieved within a certain range. Then specific fluorescence imaging of mice organ tissues was obtained for proving the permeability of probe 1. Simultaneously, the viability was measured to be more than 80%, which shows probe 1 can be a rapid and biocompatible probe for biothiols in cells. Furthermore, the measurement of thiols detection in 5 kinds of animal serum showed that probe 1 can be used in determination of biothiols in blood. Copyright © 2016 Elsevier B.V. All rights reserved.

Multiplex-PCR As a Rapid and Sensitive Method for Identification of Meat Species in Halal-Meat Products.

PubMed

Alikord, Mahsa; Keramat, Javad; Kadivar, Mahdi; Momtaz, Hassan; Eshtiaghi, Mohammad N; Homayouni-Rad, Aziz

2017-01-01

Species identification and authentication in meat products are important subjects for ensuring the health of consumers. The multiplex-PCR amplification and species- specific primer set were used for the identification of horse, donkey, pig and other ruminants in raw and processed meat products. Oligonucleotid primers were designed and patented for amplification of species-specific mitochondrial DNA sequences of each species and samples were prepared from binary meat mixtures. The results showed that meat species were accurately determined in all combinations by multiplex-PCR, and the sensitivity of this method was 0.001 ng, rendering this technique open to and suitable for use in industrial meat products. It is concluded that more fraud is seen in lower percentage industrial meat products than in higher percentage ones. There was also more fraud found in processed products than in raw ones. This rapid and useful test is recommended for quality control firms for applying more rigorous controls over industrial meat products, for the benefit of target consumers. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Detection of Shigella in Milk and Clinical Samples by Magnetic Immunocaptured-Loop-Mediated Isothermal Amplification Assay

PubMed Central

Zhang, Liding; Wei, Qiujiang; Han, Qinqin; Chen, Qiang; Tai, Wenlin; Zhang, Jinyang; Song, Yuzhu; Xia, Xueshan

2018-01-01

Shigella is an important human food-borne zoonosis bacterial pathogen, and can cause clinically severe diarrhea. There is an urgent need to develop a specific, sensitive, and rapid methodology for detection of this pathogen. In this study, loop-mediated isothermal amplification (LAMP) combined with magnetic immunocapture assay (IC-LAMP) was first developed for the detection of Shigella in pure culture, artificial milk, and clinical stool samples. This method exhibited a detection limit of 8.7 CFU/mL. Compared with polymerase chain reaction, IC-LAMP is sensitive, specific, and reliable for monitoring Shigella. Additionally, IC-LAMP is more convenient, efficient, and rapid than ordinary LAMP, as it is more efficiently enriches pathogen cells without extraction of genomic DNA. Under isothermal conditions, the amplification curves and the green fluorescence were detected within 30 min in the presence of genomic DNA template. The overall analysis time was approximately 1 h, including the enrichment and lysis of the bacterial cells, a significantly short detection time. Therefore, the IC-LAMP methodology described here is potentially useful for the efficient detection of Shigella in various samples. PMID:29467730

Reverse transcription strand invasion based amplification (RT-SIBA): a method for rapid detection of influenza A and B.

PubMed

Eboigbodin, Kevin; Filén, Sanna; Ojalehto, Tuomas; Brummer, Mirko; Elf, Sonja; Pousi, Kirsi; Hoser, Mark

2016-06-01

Rapid and accurate diagnosis of influenza viruses plays an important role in infection control, as well as in preventing the misuse of antibiotics. Isothermal nucleic acid amplification methods offer significant advantages over the polymerase chain reaction (PCR), since they are more rapid and do not require the sophisticated instruments needed for thermal cycling. We previously described a novel isothermal nucleic acid amplification method, 'Strand Invasion Based Amplification' (SIBA®), with high analytical sensitivity and specificity, for the detection of DNA. In this study, we describe the development of a variant of the SIBA method, namely, reverse transcription SIBA (RT-SIBA), for the rapid detection of viral RNA targets. The RT-SIBA method includes a reverse transcriptase enzyme that allows one-step reverse transcription of RNA to complementary DNA (cDNA) and simultaneous amplification and detection of the cDNA by SIBA under isothermal reaction conditions. The RT-SIBA method was found to be more sensitive than PCR for the detection of influenza A and B and could detect 100 copies of influenza RNA within 15 min. The development of RT-SIBA will enable rapid and accurate diagnosis of viral RNA targets within point-of-care or central laboratory settings.

Recent developments in detection and enumeration of waterborne bacteria: a retrospective minireview.

PubMed

Deshmukh, Rehan A; Joshi, Kopal; Bhand, Sunil; Roy, Utpal

2016-12-01

Waterborne diseases have emerged as global health problems and their rapid and sensitive detection in environmental water samples is of great importance. Bacterial identification and enumeration in water samples is significant as it helps to maintain safe drinking water for public consumption. Culture-based methods are laborious, time-consuming, and yield false-positive results, whereas viable but nonculturable (VBNCs) microorganisms cannot be recovered. Hence, numerous methods have been developed for rapid detection and quantification of waterborne pathogenic bacteria in water. These rapid methods can be classified into nucleic acid-based, immunology-based, and biosensor-based detection methods. This review summarizes the principle and current state of rapid methods for the monitoring and detection of waterborne bacterial pathogens. Rapid methods outlined are polymerase chain reaction (PCR), digital droplet PCR, real-time PCR, multiplex PCR, DNA microarray, Next-generation sequencing (pyrosequencing, Illumina technology and genomics), and fluorescence in situ hybridization that are categorized as nucleic acid-based methods. Enzyme-linked immunosorbent assay (ELISA) and immunofluorescence are classified into immunology-based methods. Optical, electrochemical, and mass-based biosensors are grouped into biosensor-based methods. Overall, these methods are sensitive, specific, time-effective, and important in prevention and diagnosis of waterborne bacterial diseases. © 2016 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Immuno capture PCR for rapid and sensitive identification of pathogenic Bacillus anthracis.

PubMed

Makam, Shivakiran S; Majumder, Saugata; Kingston, Joseph J; Urs, Radhika M; Tuteja, Urmil; Sripathi, Murali H; Batra, Harsh V

2013-12-01

Immuno capture PCR (IPCR) is a technique capable of detecting the pathogens with high specificity and sensitivity. Rapid and accurate detection of Bacillus anthracis was achieved using anti-EA1 antibodies to capture the cells and two primer sets targeting the virulence factors of the pathogen i.e., protective antigen (pag) and capsule (cap) in an IPCR format. Monoclonal antibodies specific to B. anthracis were generated against extractable antigen 1 protein and used as capture antibody onto 96 well polystyrene plates. Following the binding of the pathogen, the DNA extraction was carried out in the well itself and further processed for PCR assay. We compared IPCR described here with conventional duplex PCR using the same primers and sandwich ELISA using the monoclonal antibodies developed in the present study. IPCR was capable of detecting as few as 10 and 100 cfu ml⁻¹ of bacterial cells and spores, respectively. IPCR was found to be 2-3 logs more sensitive than conventional duplex PCR and the sandwich ELISA. The effect of other bacteria and any organic materials on IPCR was also analyzed and found that this method was robust with little change in the sensitivity in the presence of interfering agents. Moreover, we could demonstrate a simple process of microwave treatment for spore disruption which otherwise are resistant to chemical treatments. Also, the IPCR could clearly distinguish the pathogenic and nonpathogenic strains of B. anthracis in the same assay. This can help in saving resources on unnecessary decontamination procedures during false alarms.

Development and clinical performance of high throughput loop-mediated isothermal amplification for detection of malaria

PubMed Central

Perera, Rushini S.; Ding, Xavier C.; Tully, Frank; Oliver, James; Bright, Nigel; Bell, David; Chiodini, Peter L.; Gonzalez, Iveth J.; Polley, Spencer D.

2017-01-01

Background Accurate and efficient detection of sub-microscopic malaria infections is crucial for enabling rapid treatment and interruption of transmission. Commercially available malaria LAMP kits have excellent diagnostic performance, though throughput is limited by the need to prepare samples individually. Here, we evaluate the clinical performance of a newly developed high throughput (HTP) sample processing system for use in conjunction with the Eiken malaria LAMP kit. Methods The HTP system utilised dried blood spots (DBS) and liquid whole blood (WB), with parallel sample processing of 94 samples per run. The system was evaluated using 699 samples of known infection status pre-determined by gold standard nested PCR. Results The sensitivity and specificity of WB-HTP-LAMP was 98.6% (95% CI, 95.7–100), and 99.7% (95% CI, 99.2–100); sensitivity of DBS-HTP-LAMP was 97.1% (95% CI, 93.1–100), and specificity 100% against PCR. At parasite densities greater or equal to 2 parasites/μL, WB and DBS HTP-LAMP showed 100% sensitivity and specificity against PCR. At densities less than 2 p/μL, WB-HTP-LAMP sensitivity was 88.9% (95% CI, 77.1–100) and specificity was 99.7% (95% CI, 99.2–100); sensitivity and specificity of DBS-HTP-LAMP was 77.8% (95% CI, 54.3–99.5) and 100% respectively. Conclusions The HTP-LAMP system is a highly sensitive diagnostic test, with the potential to allow large scale population screening in malaria elimination campaigns. PMID:28166235

Accuracy of a Rapid Diagnostic Test (Cypress Chagas Quick Test®) for the Diagnosis of Chronic Chagas Disease in a Nonendemic Area: A Retrospective Longitudinal Study.

PubMed

Angheben, Andrea; Staffolani, Silvia; Anselmi, Mariella; Tais, Stefano; Degani, Monica; Gobbi, Federico; Buonfrate, Dora; Gobbo, Maria; Bisoffi, Zeno

2017-11-01

We analyzed the accuracy of Chagas Quick Test ® , a rapid diagnostic test, for the diagnosis of chronic Chagas disease through a retrospective study on a cohort of 669 patients consecutively examined at a single reference center in Italy, during a 7-year period. We observed high concordance with serological reference standard but low accuracy for screening purposes (sensitivity/specificity: 82.8%/98.7%) at least in our nonendemic context.

[Assessment of a rapid diagnostic test for malaria in rural health care facilities in Senegal].

PubMed

Munier, A; Diallo, A; Sokhna, C; Chippaux, J P

2009-10-01

The aim of the study was to determine the accuracy of a rapid diagnostic test in confirming presumptive malaria diagnosis in a rural zone of Senegal. Thick blood smear was used as the reference technique for comparison. METHOHDOLOGY: Testing was conducted on children between the ages of 1 and 14 years at three health care facilities located in the Niakhar are from August 2006 to June 2007. If malaria was suspected by the nurse based on clinical findings, two thick smears and one rapid diagnostic test (Core Malaria Pf) were performed. Blood slides were stained in Niakhar and read in Dakar. A total of 474 patients were examined. Three-fourths (75%) of these patients were seen during the rainy season. Malaria was suspected in 335 patients (71%). Rapid tests and thick smears were obtained in 330 of these patients with positive results in 194 (59%) and 180 (55%) respectively. The sensitivity, specificity, positive predictive value and negative predictive value of the rapid test were 96%, 87%, 90% and 95% respectively. Our data show that the rapid diagnostic test used in this study exhibits good sensitivity and positive predictive value. Despite its cost this test could be helpful in confirming malaria diagnosis in outlying health care facilities without the necessary resources to perform blood smears. Confirmation is necessary to avoid unwarranted prescription of malaria treatment due to inaccurate clinical diagnosis

Development of a multiplex PCR assay for rapid and simultaneous detection of four genera of fish pathogenic bacteria.

PubMed

Zhang, D F; Zhang, Q Q; Li, A H

2014-11-01

Species of genus Aeromonas, Vibrio, Edwardsiella and Streptococcus are the most common fish pathogenic bacteria that cause economically devastating losses in aquaculture. A multiplex polymerase chain reaction (mPCR) was developed for the simultaneous detection and differentiation of the four genera of fish pathogenic bacteria. Through the use of genus-specific primers instead of species-specific ones, the current mPCR covered much more target bacterial species compared with previously reported species-specific mPCR methods. The specificity of the four putative genus-specific primers was validated experimentally while used exclusively (uniplex PCR) or combined (mPCR) against bacterial genomic DNA templates of the target bacteria and nontarget bacteria. The PCR amplicons for the following genera were obtained as expected: Aeromonas (875 bp), Vibrio (524 bp), Edwardsiella (302 bp) and Streptococcus (197 bp), and the fragments could be separated clearly on the agarose gel electrophoresis. The mPCR did not produce nonspecific amplification products when used to amplify 21 nontarget species of bacteria. The mPCR detection limits for each target bacterial genera were 50 colony-forming units (CFU) in pure culture and 100 CFU in fish tissue samples. In conclusion, the mPCR assay was proven to be a powerful alternative to the conventional culture-based method, given its rapid, specific, sensitive and reliable detection of target pathogens. The fish pathogenic bacteria of genus Aeromonas, Vibrio, Edwardsiella and Streptococcus frequently cause severe outbreaks of diseases in cultured fish, and the genus-specific multiplex PCR assay developed in this study can detect the bacteria of the four genera when present in the samples either alone or mixed. The mPCR assay is expected to identify the causative agents more efficiently than uniplex PCR or species-specific multiplex PCR for clinical diagnosis, resulting in the earlier implementation of control measures. This mPCR assay provides a rapid, specific and sensitive tool for the detection or identification of common fish pathogenic bacteria in aquaculture practice. © 2014 The Society for Applied Microbiology.

Population-based assessment of sensitivity and specificity of a pinhole for detection of significant refractive errors in the community.

PubMed

Marmamula, Srinivas; Keeffe, Jill E; Narsaiah, Saggam; Khanna, Rohit C; Rao, Gullapalli N

2014-11-01

Measurements of refractive errors through subjective or automated refraction are not always possible in rapid assessment studies and community vision screening programs; however, measurements of vision with habitual correction and with a pinhole can easily be made. Although improvements in vision with a pinhole are assumed to mean that a refractive error is present, no studies have investigated the magnitude of improvement in vision with pinhole that is predictive of refractive error. The aim was to measure the sensitivity and specificity of 'vision improvement with pinhole' in predicting the presence of refractive error in a community setting. Vision and vision with pinhole were measured using a logMAR chart for 488 of 582 individuals aged 15 to 50 years. Refractive errors were measured using non-cycloplegic autorefraction and subjective refraction. The presence of refractive error was defined using spherical equivalent refraction (SER) at two levels: SER greater than ± 0.50 D sphere (DS) and SER greater than ±1.00 DS. Three definitions for significant improvement in vision with a pinhole were used: 1. Presenting vision less than 6/12 and improving to 6/12 or better, 2. Improvement in vision of more than one logMAR line and 3. Improvement in vision of more than two logMAR lines. For refractive error defined as spherical equivalent refraction greater than ± 0.50 DS, the sensitivities and specificities for the pinhole test predicting the presence of refractive error were 83.9 per cent (95% CI: 74.5 to 90.9) and 98.8 per cent (95% CI: 97.1 to 99.6), respectively for definition 1. Definition 2 had a sensitivity 89.7 per cent (95% CI: 81.3 to 95.2) and specificity 88.0 per cent (95% CI: 4.4 to 91.0). Definition 3 had a sensitivity of 75.9 per cent (95% CI: 65.5 to 84.4) and specificity of 97.8 per cent (95% CI: 95.8 to 99.0). Similar results were found with spherical equivalent refraction greater than ±1.00 DS, when tested against the three pinhole-based definitions. Refractive error definitions based on improvement in vision with the pinhole shows good sensitivity and specificity at predicting the presence of significant refractive errors. These definitions can be used in rapid assessment surveys and community-based vision screenings. © 2014 The Authors. Clinical and Experimental Optometry © 2014 Optometrists Association Australia.

Diagnostic value of sputum adenosine deaminase (ADA) level in pulmonary tuberculosis.

PubMed

Binesh, Fariba; Jalali, Hadi; Zare, Mohammad Reza; Behravan, Farhad; Tafti, Arefeh Dehghani; Behnaz, Fatemah; Tabatabaee, Mohammad; Shahcheraghi, Seyed Hossein

2016-06-01

Tuberculosis is still a considerable health problem in many countries. Rapid diagnosis of this disease is important, and adenosine deaminase (ADA) has been used as a diagnostic test. The aim of this study was to assess the diagnostic value of ADA in the sputum of patients with pulmonary tuberculosis. The current study included 40 patients with pulmonary tuberculosis (culture positive, smear ±) and 42 patients with non tuberculosis pulmonary diseases (culture negative). ADA was measured on all of the samples. The median value of ADA in non-tuberculosis patients was 2.94 (4.2) U/L and 4.01 (6.54) U/L in tuberculosis patients, but this difference was not statistically significant (p=0.100). The cut-off point of 3.1 U/L had a sensitivity of 61% and a specificity of 53%, the cut-off point of 2.81 U/L had a sensitivity of 64% and a specificity of 50% and the cut-off point of 2.78 U/L had a sensitivity of 65% and a specificity of 48%. The positive predictive values for cut-off points of 3.1, 2.81 and 2.78 U/L were 55.7%, 57.44% and 69.23%, respectively. The negative predictive values for the abovementioned cut-off points were 56.75%, 57.14% and 55.88%, respectively. Our results showed that sputum ADA test is neither specific nor sensitive. Because of its low sensitivity and specificity, determination of sputum ADA for the diagnosis of pulmonary tuberculosis is not recommended.

Evaluation of a New Immunochromatography Technology Test (LDBio Diagnostics) To Detect Toxoplasma IgG and IgM: Comparison with the Routine Architect Technique

PubMed Central

Flori, Pierre; Delaunay, Edouard; Guillerme, Cécile; Charaoui, Sana; Raberin, Hélène; Hafid, Jamal; L'Ollivier, Coralie

2017-01-01

ABSTRACT A study comparing the ICT (immunochromatography technology) Toxoplasma IgG and IgM rapid diagnostic test (LDBio Diagnostics, France) with a fully automated system, Architect, was performed on samples from university hospitals of Marseille and Saint-Etienne. A total of 767 prospective sera and 235 selected sera were collected. The panels were selected to test various IgG and IgM parameters. The reference technique, Toxoplasma IgGII Western blot analysis (LDBio Diagnostics), was used to confirm the IgG results, and commercial kits Platelia Toxo IgM (Bio-Rad) and Toxo-ISAgA (bioMérieux) were used in Saint-Etienne and Marseille, respectively, as the IgM reference techniques. Sensitivity and specificity of the ICT and the Architect IgG assays were compared using a prospective panel. Sensitivity was 100% for the ICT test and 92.1% for Architect (cutoff at 1.6 IU/ml). The low-IgG-titer serum results confirmed that ICT sensitivity was superior to that of Architect. Specificity was 98.7% (ICT) and 99.8% (Architect IgG). The ICT test is also useful for detecting IgM without IgG and is both sensitive (100%) and specific (100%), as it can distinguish nonspecific IgM from specific Toxoplasma IgM. In comparison, IgM sensitivity and specificity on Architect are 96.1% and 99.6%, respectively (cutoff at 0.5 arbitrary units [AU]/ml). To conclude, this new test overcomes the limitations of automated screening techniques, which are not sensitive enough for IgG and lack specificity for IgM (rare IgM false-positive cases). PMID:28954897

[Validity of expired carbon monoxide and urine cotinine using dipstick method to assess smoking status].

PubMed

Park, Su San; Lee, Ju Yul; Cho, Sung-Il

2007-07-01

We investigated the validity of the dipstick method (Mossman Associates Inc. USA) and the expired CO method to distinguish between smokers and nonsmokers. We also elucidated the related factors of the two methods. This study included 244 smokers and 50 ex-smokers, recruited from smoking cessation clinics at 4 local public health centers, who had quit for over 4 weeks. We calculated the sensitivity, specificity and Kappa coefficient of each method for validity. We obtained ROC curve, predictive value and agreement to determine the cutoff of expired air CO method. Finally, we elucidated the related factors and compared their effect powers using the standardized regression coefficient. The dipstick method showed a sensitivity of 92.6%, specificity of 96.0% and Kappa coefficient of 0.79. The best cutoff value to distinguish smokers was 5-6 ppm. At 5 ppm, the expired CO method showed a sensitivity of 94.3%, specificity of 82.0% and Kappa coefficient of 0.73. And at 6 ppm, sensitivity, specificity and Kappa coefficient were 88.5%, 86.0% and 0.64, respectively. Therefore, the dipstick method had higher sensitivity and specificity than the expired CO method. The dipstick and expired CO methods were significantly increased with increasing smoking amount. With longer time since the last smoking, expired CO showed a rapid decrease after 4 hours, whereas the dipstick method showed relatively stable levels for more than 4 hours. The dipstick and expired CO methods were both good indicators for assessing smoking status. However, the former showed higher sensitivity and specificity and stable levels over longer hours after smoking, compared to the expired CO method.

Rapid identification and differentiation of Fasciola hepatica and Fasciola gigantica by a loop-mediated isothermal amplification (LAMP) assay.

PubMed

Ai, L; Li, C; Elsheikha, H M; Hong, S J; Chen, J X; Chen, S H; Li, X; Cai, X Q; Chen, M X; Zhu, X Q

2010-12-15

The present study developed and validated a species-specific loop-mediated isothermal amplification (LAMP) assay for the rapid detection and discrimination of Fasciola hepatica and Fasciola gigantica. The LAMP assay is inexpensive, easy to perform and shows rapid reaction, wherein the amplification can be obtained in 45 min under isothermal conditions of 61 °C or 62 °C by employing a set of four species-specific primer mixtures and results can be checked through naked-eye visualization. The optimal assay conditions with no cross-reaction with other closely related trematodes (Clonorchis sinensis, Opisthorchis viverrini, Orientobilharzia turkestanicum and Schistosoma japonicum) as well as within the two Fasciola species were established. The assay was validated by examining F. gigantica DNA in the intermediate host snails and in faecal samples. The results indicated that the LAMP assay is approximately 10(4) times more sensitive than the conventional specific PCR assays. These findings indicate that this Fasciola species-specific LAMP assay may have a potential clinical application for detection and differentiation of Fasciola species, especially in endemic countries. Copyright © 2010 Elsevier B.V. All rights reserved.

Evaluation of the OnSite (Pf/Pan) rapid diagnostic test for diagnosis of clinical malaria

PubMed Central

2012-01-01

Background Accurate diagnosis of malaria is an essential prerequisite for proper treatment and drug resistance monitoring. Microscopy is considered the gold standard for malaria diagnosis but has limitations. ELISA, PCR, and Real Time PCR are also used to diagnose malaria in reference laboratories, although their application at the field level is currently not feasible. Rapid diagnostic tests (RDTs) however, have been brought into field operation and widely adopted in recent days. This study evaluates OnSite (Pf/Pan) antigen test, a new RDT introduced by CTK Biotech Inc, USA for malaria diagnosis in a reference setting. Methods Blood samples were collected from febrile patients referred for malaria diagnosis by clinicians. Subjects were included in this study from two different Upazila Health Complexes (UHCs) situated in two malaria endemic districts of Bangladesh. Microscopy and nested PCR were considered the gold standard in this study. OnSite (Pf/Pan) RDT was performed on preserved whole blood samples. Results In total, 372 febrile subjects were included in this study. Of these subjects, 229 (61.6%) tested positive for Plasmodium infection detected by microscopy and nested PCR. OnSite (Pf/Pan) RDT was 94.2% sensitive (95% CI, 89.3-97.3) and 99.5% specific (95% CI, 97.4-00.0) for Plasmodium falciparum diagnosis and 97.3% sensitive (95% CI, 90.5-99.7) and 98.7% specific (95% CI, 96.6-99.6) for Plasmodium vivax diagnosis. Sensitivity varied with differential parasite count for both P. falciparum and P. vivax. The highest sensitivity was observed in febrile patients with parasitaemia that ranged from 501–1,000 parasites/μL regardless of the Plasmodium species. Conclusion The new OnSite (Pf/Pan) RDT is both sensitive and specific for symptomatic malaria diagnosis in standard laboratory conditions. PMID:23234579

Comparison of the Diagnostic Accuracy of Three Rapid Tests for the Serodiagnosis of Hepatic Cystic Echinococcosis in Humans

PubMed Central

Tamarozzi, Francesca; Covini, Ilaria; Mariconti, Mara; Narra, Roberta; Tinelli, Carmine; De Silvestri, Annalisa; Manzoni, Federica; Casulli, Adriano; Ito, Akira; Neumayr, Andreas; Brunetti, Enrico

2016-01-01

Background The diagnosis of cystic echinococcosis (CE) is based primarily on imaging, in particular with ultrasound for abdominal CE, complemented by serology when imaging results are unclear. In rural endemic areas, where expertise in ultrasound may be scant and conventional serology techniques are unavailable due to lack of laboratory equipment, Rapid Diagnostic Tests (RDTs) are appealing. Methodology/Principal Findings We evaluated the diagnostic accuracy of 3 commercial RDTs for the diagnosis of hepatic CE. Sera from 59 patients with single hepatic CE cysts in well-defined ultrasound stages (gold standard) and 25 patients with non-parasitic cysts were analyzed by RDTs VIRapid HYDATIDOSIS (Vircell, Spain), Echinococcus DIGFA (Unibiotest, China), ADAMU-CE (ICST, Japan), and by RIDASCREEN Echinococcus IgG ELISA (R-Biopharm, Germany). Sensitivity, specificity and ROC curves were compared with McNemar and t-test. For VIRapid and DIGFA, correlation between semiquantitative results and ELISA OD values were evaluated by Spearman’s coefficient. Reproducibility was assessed on 16 randomly selected sera with Cohen’s Kappa coefficient. Sensitivity and Specificity of VIRapid (74%, 96%) and ADAMU-CE (57%, 100%) did not differ from ELISA (69%, 96%) while DIGFA (72%, 72%) did (p = 0.045). ADAMU-CE was significantly less sensitive in the diagnosis of active cysts (p = 0.019) while DIGFA was significantly less specific (p = 0.014) compared to ELISA. All tests were poorly sensitive in diagnosing inactive cysts (33.3% ELISA and ADAMU-CE, 42.8% DIGFA, 47.6% VIRapid). The reproducibility of all RDTs was good-very good. Band intensity of VIRapid and DIGFA correlated with ELISA OD values (r = 0.76 and r = 0.79 respectively, p<0.001). Conclusions/Significance RDTs may be useful in resource-poor settings to complement ultrasound diagnosis of CE in uncertain cases. VIRapid test appears to perform best among the examined kits, but all tests are poorly sensitive in the presence of inactive cysts, which may pose problems with accurate diagnosis. PMID:26871432

Evaluation of Seegene Allplex Respiratory Panel 1 kit for the detection of influenza virus and human respiratory syncytial virus.

PubMed

Gimferrer, Laura; Andrés, Cristina; Rando, Ariadna; Piñana, Maria; Codina, Maria Gema; Martin, Maria Del Carmen; Fuentes, Francisco; Rubio, Susana; Alcubilla, Pilar; Pumarola, Tomàs; Antón, Andrés

2018-05-25

Influenza (FLUV) and human respiratory syncytial (HRSV) viruses are etiological agents of respiratory infections that cause a significant morbidity and mortality worldwide. A rapid and accurate diagnosis of these respiratory viruses is essential for an appropriate patient management. Molecular tests are the best detection option due to their high sensitivity and specificity. Seegene's Allplex™ Respiratory Panel 1 (Allplex RP1) is a real-time one-step RT-PCR assay for the simultaneous detection of FLUAV, FLUBV, HRSV-A and HRSV-B. In addition, it allows the determination of FLUAV subtype (H1, H3 and H1pdm09). This study aims to evaluate Allplex RP1 as a rapid molecular test for the detection of FLUAV, FLUBV, HRSV-A and HRSV-B viruses. The Allplex RP1 assay will be compared with other two commercial molecular assays, Prodesse ProFlu+ and ProFAST+ (Hologic, Madison, WI, USA), and GeneXpert Flu/RSV XC (Cepheid, USA). Allplex RP1, ProFlu+ and GeneXpert tests showed 95%, 91% and 96% of accuracy; and 94%, 88% and 95% of sensitivity, respectively. Moreover, Allplex RP1 showed a FLUAV subtype sensitivity of 91% and 88% for FLUAV-H1pdm09 and FLUAV-H3 respectively, and ProFAST+ assay showed sensitivities of 100% for both targets. The three assays showed a 100% of specificity and PPV, while the NPV were 84%, 73% and 86% for Allplex RP1, Prodesse and GeneXpert, respectively. In this study, Seegene's Allplex RP1 assay showed to be highly sensitive, specific, and suitable for detection of FLUV and HRSV, including FLUAV subtyping. In addition, it is also a hands-on-time saving assay due to the automated nucleic acid extraction and PCR setup. Copyright © 2018. Published by Elsevier B.V.

Performance of two rapid diagnostic tests for malaria diagnosis at the China-Myanmar border area

PubMed Central

2013-01-01

Background Rapid diagnostic tests (RDTs) have become an essential tool in the contemporary malaria control and management programmes in the world. This study aims to evaluate the performance of two commonly used RDTs for malaria diagnosis in the China-Myanmar border area. Methods A total 606 febrile patients in the China-Myanmar border were recruited to this study and were diagnosed for malaria infections by microscopy, two RDTs tests (Pf/Pan device, and Pv/Pf device) and nested PCR. Results Malaria parasites were found in 143 patients by microscopy, of which 51, 73, and 19 were Plasmodium falciparum, Plasmodium vivax and P. falciparum/P. vivax mixed infections, respectively. Compared to microscopy, the sensitivity of the Pf/Pan device was 88.6% for P. falciparum and 69.9% for P. vivax with the specificity of 90.4%. For a subset of 350 patients, the sensitivity of the Pf/Pan device and Pv/Pf device for detection of P. falciparum was 87.5% and 91.7%, respectively; and for detection of P. vivax was 72.0% and 73.8%, respectively. The specificity of the Pf/Pan device and Pv/Pf device was 94.3% and 96.5%, respectively. Nested PCR detected malaria parasites in 174 of 606 samples, of which 67, 79, two and 26 were P. falciparum, P. vivax, P. ovale and P. falciparum/P. vivax mixed infections, respectively. Compared to nested PCR, all other methods had sensitivity below 80%, suggesting that a significant number of cases were missed. Conclusions Compared to PCR, both microscopy and RDTs had lower sensitivities. RDTs had similar performance to microscopy for P. falciparum diagnosis, but performed worse for P. vivax diagnosis. Other RDT products should be selected with higher sensitivity (and good specificity) for both P. falciparum and P. vivax diagnosis. PMID:23433230

Microscopic-observation drug susceptibility and thin layer agar assays for the detection of drug resistant tuberculosis: a systematic review and meta-analysis.

PubMed

Minion, Jessica; Leung, Erika; Menzies, Dick; Pai, Madhukar

2010-10-01

Simple, rapid, and affordable tests are needed to detect drug resistance in Mycobacterium tuberculosis. We did a systematic review and meta-analysis to investigate the accuracy of microscopic-observation drug susceptibility (MODS) and thin layer agar (TLA) assays for rapid screening of patients at risk of drug-resistant tuberculosis. In accordance with protocols and methods recommended by the Cochrane Diagnostic Test Accuracy Working Group, we systematically searched PubMed, Embase, and Biosis for reports published between January, 1990, and February, 2009. We included studies investigating detection of drug resistance in M tuberculosis with the MODS or TLA assay, and in which an accepted reference standard was used. Data extracted from the studies were combined by use of bivariate random-effects regression models and hierarchical summary receiver operating characteristic curves to estimate sensitivity and specificity for detection of resistance to specific drugs. We identified 12 studies, of which nine investigated the MODS assay and three investigated the TLA assay. For the MODS assay of rifampicin resistance, pooled estimates were 98·0% (95% CI 94·5-99·3) for sensitivity and 99·4% (95·7-99·9) for specificity. For the MODS assay of isoniazid resistance with a 0·1 μg/mL cutoff, pooled sensitivity was 97·7% (94·4-99·1) and pooled specificity was 95·8% (88·1-98·6), but with a 0·4 μg/mL cutoff, sensitivity decreased to 90·0% (84·5-93·7) and specificity increased to 98·6% (96·9-99·4). All assessments of rifampicin and isoniazid resistance with the TLA assay yielded 100% accuracy. Mean turnaround time was 9·9 days (95% CI 4·1-15·8) for the MODS assay and 11·1 days (10·1-12·0) for the TLA assay. MODS and TLA assays are inexpensive, rapid alternatives to conventional methods for drug susceptibility testing of M tuberculosis. Our data and expert opinion informed WHO's recommendation for use of selected non-commercial drug susceptibility tests, including MODS, as an interim solution until capacity for genotypic or automated liquid culture drug susceptibility testing is developed. Stop TB Department of WHO. Copyright © 2010 Elsevier Ltd. All rights reserved.

Recombinant Phage Probes for Salmonella Typhimurium Detection

DTIC Science & Technology

2005-03-23

food safety analysis that are slower, labor-intensive, and cost-inefficient. Confirmation of presence in food products can take as long as 48 hours by conventional culture. Current rapid detection initiatives include biosensors that routinely incorporate antibodies as the biorecognition unit. Although sensitive and specific, antibodies are costly and may degrade under unfavorable environmental conditions. We believe that a stable, inexpensive substitute for antibodies is filamentous phage manipulated through phage display technique then affinity selected for specificity to

Effectiveness of real-time polymerase chain reaction assay for the detection of Mycobacterium tuberculosis in pathological samples: a systematic review and meta-analysis.

PubMed

Babafemi, Emmanuel O; Cherian, Benny P; Banting, Lee; Mills, Graham A; Ngianga, Kandala

2017-10-25

Rapid and accurate diagnosis of tuberculosis (TB) is key to manage the disease and to control and prevent its transmission. Many established diagnostic methods suffer from low sensitivity or delay of timely results and are inadequate for rapid detection of Mycobacterium tuberculosis (MTB) in pulmonary and extra-pulmonary clinical samples. This study examined whether a real-time polymerase chain reaction (RT-PCR) assay, with a turn-a-round time of 2 h, would prove effective for routine detection of MTB by clinical microbiology laboratories. A systematic literature search was performed for publications in any language on the detection of MTB in pathological samples by RT-PCR assay. The following sources were used MEDLINE via PubMed, EMBASE, BIOSIS Citation Index, Web of Science, SCOPUS, ISI Web of Knowledge and Cochrane Infectious Diseases Group Specialised Register, grey literature, World Health Organization and Centres for Disease Control and Prevention websites. Forty-six studies met set inclusion criteria. Generated pooled summary estimates (95% CIs) were calculated for overall accuracy and bivariate meta-regression model was used for meta-analysis. Summary estimates for pulmonary TB (31 studies) were as follows: sensitivity 0.82 (95% CI 0.81-0.83), specificity 0.99 (95% CI 0.99-0.99), positive likelihood ratio 43.00 (28.23-64.81), negative likelihood ratio 0.16 (0.12-0.20), diagnostic odds ratio 324.26 (95% CI 189.08-556.09) and area under curve 0.99. Summary estimates for extra-pulmonary TB (25 studies) were as follows: sensitivity 0.70 (95% CI 0.67-0.72), specificity 0.99 (95% CI 0.99-0.99), positive likelihood ratio 29.82 (17.86-49.78), negative likelihood ratio 0.33 (0.26-0.42), diagnostic odds ratio 125.20 (95% CI 65.75-238.36) and area under curve 0.96. RT-PCR assay demonstrated a high degree of sensitivity for pulmonary TB and good sensitivity for extra-pulmonary TB. It indicated a high degree of specificity for ruling in TB infection from sampling regimes. This was acceptable, but may better as a rule out add-on diagnostic test. RT-PCR assays demonstrate both a high degree of sensitivity in pulmonary samples and rapidity of detection of TB which is an important factor in achieving effective global control and for patient management in terms of initiating early and appropriate anti-tubercular therapy. PROSPERO CRD42015027534 .

Can we rely on the multiplex ligation-dependent probe amplification method (MLPA) for prenatal diagnosis?

PubMed Central

Omrani, Mir Davood; Azizi, Faezeh; Rajabibazl, Masoumeh; Safavi Naini, Niloufar; Omrani, Sara; Abbasi, Arezo Mona; Saleh Gargari, Soraya

2014-01-01

Background: The major aneuploidies that are diagnosed prenatally involve the autosomal chromosomes 13, 18, and 21, as well as sex chromosomes, X and Y. Because multiplex ligation-dependent probe amplification (MLPA) is rapid and non-invasive, it has replaced traditional culture methods for the screening and diagnosis of common aneuploidies in some countries. Objective: To evaluate the sensitivity and specificity of MLPA in a cross-sectional descriptive study for the detection of chromosomal aneuploidies in comparison to other methods. Materials and Methods: Genomic DNA was extracted from the peripheral blood samples of 10 normal controls and the amniotic fluid of 55 patients. Aneuploidies screening of chromosomes 13, 18, 21, X and Y were carried out using specific MLPA probe mixes (P095-A2). For comparison purposes, samples were also tested by Quantitative Fluorescent-PCR (QF-PCR) and routine chromosomal culture method. Results: Using this specific MLPA technique and data-analyzing software (Genemarker v1.85), one case was diagnosed with 45, X (e.g. Monosomy X or Turner’s Syndrome), and the remaining 54 cases revealed normal karyotypes. These results were concordant with routine chromosomal culture and QF-PCR findings. Conclusion: The experiment demonstrates that MLPA can provide a rapid and accurate clinical method for prenatal identification of common chromosomal aneuploidies with 100% sensitivity and 100% specificity. PMID:24976821

Optical imaging of RNAi-mediated silencing of cancer

NASA Astrophysics Data System (ADS)

Ochiya, Takahiro; Honma, Kimi; Takeshita, Fumitaka; Nagahara, Shunji

2008-02-01

RNAi has rapidly become a powerful tool for drug target discovery and validation in an in vitro culture system and, consequently, interest is rapidly growing for extension of its application to in vivo systems, such as animal disease models and human therapeutics. Cancer is one obvious application for RNAi therapeutics, because abnormal gene expression is thought to contribute to the pathogenesis and maintenance of the malignant phenotype of cancer and thereby many oncogenes and cell-signaling molecules present enticing drug target possibilities. RNAi, potent and specific, could silence tumor-related genes and would appear to be a rational approach to inhibit tumor growth. In subsequent in vivo studies, the appropriate cancer model must be developed for an evaluation of siRNA effects on tumors. How to evaluate the effect of siRNA in an in vivo therapeutic model is also important. Accelerating the analyses of these models and improving their predictive value through whole animal imaging methods, which provide cancer inhibition in real time and are sensitive to subtle changes, are crucial for rapid advancement of these approaches. Bioluminescent imaging is one of these optically based imaging methods that enable rapid in vivo analyses of a variety of cellular and molecular events with extreme sensitivity.

Evaluation of Commercially Available Diagnostic Tests for the Detection of Dengue Virus NS1 Antigen and Anti-Dengue Virus IgM Antibody

PubMed Central

Hunsperger, Elizabeth A.; Yoksan, Sutee; Buchy, Philippe; Nguyen, Vinh Chau; Sekaran, Shamala Devi; Enria, Delia A.; Vazquez, Susana; Cartozian, Elizabeth; Pelegrino, Jose L.; Artsob, Harvey; Guzman, Maria G.; Olliaro, Piero; Zwang, Julien; Guillerm, Martine; Kliks, Susie; Halstead, Scott; Peeling, Rosanna W.; Margolis, Harold S.

2014-01-01

Commercially available diagnostic test kits for detection of dengue virus (DENV) non-structural protein 1 (NS1) and anti-DENV IgM were evaluated for their sensitivity and specificity and other performance characteristics by a diagnostic laboratory network developed by World Health Organization (WHO), the UNICEF/UNDP/World Bank/WHO Special Programme for Research and Training in Tropical Diseases (TDR) and the Pediatric Dengue Vaccine Initiative (PDVI). Each network laboratory contributed characterized serum specimens for the panels used in the evaluation. Microplate enzyme-linked immunosorbent assay (ELISA) and rapid diagnostic test (RDT formats) were represented by the kits. Each ELISA was evaluated by 2 laboratories and RDTs were evaluated by at least 3 laboratories. The reference tests for IgM anti-DENV were laboratory developed assays produced by the Armed Forces Research Institute for Medical Science (AFRIMS) and the Centers for Disease Control and Prevention (CDC), and the NS1 reference test was reverse transcriptase polymerase chain reaction (RT-PCR). Results were analyzed to determine sensitivity, specificity, inter-laboratory and inter-reader agreement, lot-to-lot variation and ease-of-use. NS1 ELISA sensitivity was 60–75% and specificity 71–80%; NS1 RDT sensitivity was 38–71% and specificity 76–80%; the IgM anti-DENV RDTs sensitivity was 30–96%, with a specificity of 86–92%, and IgM anti-DENV ELISA sensitivity was 96–98% and specificity 78–91%. NS1 tests were generally more sensitive in specimens from the acute phase of dengue and in primary DENV infection, whereas IgM anti-DENV tests were less sensitive in secondary DENV infections. The reproducibility of the NS1 RDTs ranged from 92-99% and the IgM anti-DENV RDTs from 88–94%. PMID:25330157

The Sensitivity and Specificity of Loop-Mediated Isothermal Amplification (LAMP) Assay for Tuberculosis Diagnosis in Adults with Chronic Cough in Malawi.

PubMed

Nliwasa, Marriott; MacPherson, Peter; Chisala, Palesa; Kamdolozi, Mercy; Khundi, McEwen; Kaswaswa, Kruger; Mwapasa, Mphatso; Msefula, Chisomo; Sohn, Hojoon; Flach, Clare; Corbett, Elizabeth L

2016-01-01

Current tuberculosis diagnostics lack sensitivity, and are expensive. Highly accurate, rapid and cheaper diagnostic tests are required for point of care use in low resource settings with high HIV prevalence. To investigate the sensitivity and specificity, and cost of loop-mediated isothermal amplification (LAMP) assay for tuberculosis diagnosis in adults with chronic cough compared to Xpert® MTB/RIF, fluorescence smear microscopy. Between October 2013 and March 2014, consecutive adults at a primary care clinic were screened for cough, offered HIV testing and assessed for tuberculosis using LAMP, Xpert® MTB/RIF and fluorescence smear microscopy. Sensitivity and specificity (with culture as reference standard), and costs were estimated. Of 273 adults recruited, 44.3% (121/273) were HIV-positive and 19.4% (53/273) had bacteriogically confirmed tuberculosis. The sensitivity of LAMP compared to culture was 65.0% (95% CI: 48.3% to 79.4%) with 100% (95% CI: 98.0% to 100%) specificity. The sensitivity of Xpert® MTB/RIF (77.5%, 95% CI: 61.5% to 89.2%) was similar to that of LAMP, p = 0.132. The sensitivity of concentrated fluorescence smear microscopy with routine double reading (87.5%, 95% CI: 73.2% to 95.8%) was higher than that of LAMP, p = 0.020. All three tests had high specificity. The lowest cost per test of LAMP was at batch size of 14 samples (US$ 9.98); this was lower than Xpert® MTB/RIF (US$ 13.38) but higher than fluorescence smear microscopy (US$ 0.65). The sensitivity of LAMP was similar to Xpert® MTB/RIF but lower than fluorescence smear microscopy; all three tests had high specificity. These findings support the Malawi policy that recommends a combination of fluorescence smear microscopy and Xpert® MTB/RIF prioritised for people living with HIV, already found to be smear-negative, or being considered for retreatment of tuberculosis.

The Sensitivity and Specificity of Loop-Mediated Isothermal Amplification (LAMP) Assay for Tuberculosis Diagnosis in Adults with Chronic Cough in Malawi

PubMed Central

Nliwasa, Marriott; MacPherson, Peter; Chisala, Palesa; Kamdolozi, Mercy; Khundi, McEwen; Kaswaswa, Kruger; Mwapasa, Mphatso; Msefula, Chisomo; Sohn, Hojoon; Flach, Clare; Corbett, Elizabeth L.

2016-01-01

Background Current tuberculosis diagnostics lack sensitivity, and are expensive. Highly accurate, rapid and cheaper diagnostic tests are required for point of care use in low resource settings with high HIV prevalence. Objective To investigate the sensitivity and specificity, and cost of loop-mediated isothermal amplification (LAMP) assay for tuberculosis diagnosis in adults with chronic cough compared to Xpert® MTB/RIF, fluorescence smear microscopy. Methods Between October 2013 and March 2014, consecutive adults at a primary care clinic were screened for cough, offered HIV testing and assessed for tuberculosis using LAMP, Xpert® MTB/RIF and fluorescence smear microscopy. Sensitivity and specificity (with culture as reference standard), and costs were estimated. Results Of 273 adults recruited, 44.3% (121/273) were HIV-positive and 19.4% (53/273) had bacteriogically confirmed tuberculosis. The sensitivity of LAMP compared to culture was 65.0% (95% CI: 48.3% to 79.4%) with 100% (95% CI: 98.0% to 100%) specificity. The sensitivity of Xpert® MTB/RIF (77.5%, 95% CI: 61.5% to 89.2%) was similar to that of LAMP, p = 0.132. The sensitivity of concentrated fluorescence smear microscopy with routine double reading (87.5%, 95% CI: 73.2% to 95.8%) was higher than that of LAMP, p = 0.020. All three tests had high specificity. The lowest cost per test of LAMP was at batch size of 14 samples (US$ 9.98); this was lower than Xpert® MTB/RIF (US$ 13.38) but higher than fluorescence smear microscopy (US$ 0.65). Conclusion The sensitivity of LAMP was similar to Xpert® MTB/RIF but lower than fluorescence smear microscopy; all three tests had high specificity. These findings support the Malawi policy that recommends a combination of fluorescence smear microscopy and Xpert® MTB/RIF prioritised for people living with HIV, already found to be smear-negative, or being considered for retreatment of tuberculosis. PMID:27171380

Microfluidic biosensor for β-Hydroxybutyrate (βHBA) determination of subclinical ketosis diagnosis.

PubMed

Weng, Xuan; Zhao, Wenting; Neethirajan, Suresh; Duffield, Todd

2015-02-12

Determination of β-hydroxybutyrate (βHBA) is a gold standard for diagnosis of Subclinical Ketosis (SCK), a common disease in dairy cows that causes significant economic loss. Early detection of SCK can help reduce the risk of the disease progressing into clinical stage, thus minimizing economic losses on dairy cattle. Conventional laboratory methods are time consuming and labor-intensive, requiring expensive and bulky equipment. Development of portable and robust devices for rapid on-site SCK diagnosis is an effective way to prevent and control ketosis and can significantly aid in the management of dairy animal health. Microfluidic technology provides a rapid, cost-effective way to develop handheld devices for on-farm detection of sub-clinical ketosis. In this study, a highly sensitive microfluidics-based biosensor for on-site SCK diagnosis has been developed. A rapid, low-cost microfluidic biosensor with high sensitivity and specificity was developed for SCK diagnosis. Determination of βHBA was employed as the indicator in the diagnosis of SCK. On-chip detection using miniaturized and cost-effective optical sensor can be finished in 1 minute with a detection limit of 0.05 mM concentration. Developed microfluidic biosensor was successfully tested with the serum samples from dairy cows affected by SCK. The results of the developed biosensor agreed well with two other laboratory methods. The biosensor was characterized by high sensitivity and specificity towards βHBA with a detection limit of 0.05 mM. The developed microfluidic biosensor provides a promising prototype for a cost-effective handheld meter for on-site SCK diagnosis. By using microfluidic method, the detection time is significantly decreased compared to other laboratory methods. Here, we demonstrate a field-deployable device to precisely identify and measure subclinical ketosis by specific labeling and quantification of β-hydroxybutyate in cow blood samples. A real-time on-site detection system will maximize convenience for the farmers.

Univariate and bivariate likelihood-based meta-analysis methods performed comparably when marginal sensitivity and specificity were the targets of inference.

PubMed

Dahabreh, Issa J; Trikalinos, Thomas A; Lau, Joseph; Schmid, Christopher H

2017-03-01

To compare statistical methods for meta-analysis of sensitivity and specificity of medical tests (e.g., diagnostic or screening tests). We constructed a database of PubMed-indexed meta-analyses of test performance from which 2 × 2 tables for each included study could be extracted. We reanalyzed the data using univariate and bivariate random effects models fit with inverse variance and maximum likelihood methods. Analyses were performed using both normal and binomial likelihoods to describe within-study variability. The bivariate model using the binomial likelihood was also fit using a fully Bayesian approach. We use two worked examples-thoracic computerized tomography to detect aortic injury and rapid prescreening of Papanicolaou smears to detect cytological abnormalities-to highlight that different meta-analysis approaches can produce different results. We also present results from reanalysis of 308 meta-analyses of sensitivity and specificity. Models using the normal approximation produced sensitivity and specificity estimates closer to 50% and smaller standard errors compared to models using the binomial likelihood; absolute differences of 5% or greater were observed in 12% and 5% of meta-analyses for sensitivity and specificity, respectively. Results from univariate and bivariate random effects models were similar, regardless of estimation method. Maximum likelihood and Bayesian methods produced almost identical summary estimates under the bivariate model; however, Bayesian analyses indicated greater uncertainty around those estimates. Bivariate models produced imprecise estimates of the between-study correlation of sensitivity and specificity. Differences between methods were larger with increasing proportion of studies that were small or required a continuity correction. The binomial likelihood should be used to model within-study variability. Univariate and bivariate models give similar estimates of the marginal distributions for sensitivity and specificity. Bayesian methods fully quantify uncertainty and their ability to incorporate external evidence may be useful for imprecisely estimated parameters. Copyright © 2017 Elsevier Inc. All rights reserved.

Validation of the MDS research criteria for prodromal Parkinson's disease: Longitudinal assessment in a REM sleep behavior disorder (RBD) cohort.

PubMed

Fereshtehnejad, Seyed-Mohammad; Montplaisir, Jacques Y; Pelletier, Amelie; Gagnon, Jean-François; Berg, Daniela; Postuma, Ronald B

2017-06-01

Recently, the International Parkinson and Movement Disorder Society introduced the prodromal criteria for PD. Objectives Our study aimed to examine diagnostic accuracy of the criteria as well as the independence of prodromal markers to predict conversion to PD or dementia with Lewy bodies. This prospective cohort study was performed on 121 individuals with rapid eye movement sleep behavior disorder who were followed annually for 1 to 12 years. Using data from a comprehensive panel of prodromal markers, likelihood ratio and post-test probability of the criteria were calculated at baseline and during each follow-up visit. Forty-eight (39.7%) individuals with rapid eye movement sleep behavior disorder converted to PD/dementia with Lewy bodies. The prodromal criteria had 81.3% sensitivity and 67.9% specificity for conversion to PD/dementia with Lewy bodies at 4-year follow-up. One year before conversion, sensitivity was 100%. The criteria predicted dementia with Lewy bodies with even higher accuracy than PD without dementia at onset. Those who met the threshold of prodromal criteria at baseline had significantly more rapid conversion into a neurodegenerative state (4.8 vs. 9.1 years; P < 0.001). Pair-wise combinations of different prodromal markers showed that markers were independent of one another. The prodromal criteria are a promising tool for predicting incidence of PD/dementia with Lewy bodies and conversion time in a rapid eye movement sleep behavior disorder cohort, with high sensitivity and high specificity with long follow-up. Prodromal markers influence the overall likelihood ratio independently, allowing them to be reliably multiplied. Defining additional markers with high likelihood ratio, further studies with longitudinal assessment and testing thresholds in different target populations will improve the criteria. © 2017 International Parkinson and Movement Disorder Society. © 2017 International Parkinson and Movement Disorder Society.

Prospective Evaluation of Light Scatter Technology Paired with Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry for Rapid Diagnosis of Urinary Tract Infections

PubMed Central

Montgomery, Sandra; Roman, Kiana; Ngyuen, Lan; Cardenas, Ana Maria; Knox, James; Tomaras, Andrew P.

2017-01-01

ABSTRACT Urinary tract infections are one of the most common reasons for health care visits. Diagnosis and optimal treatment often require a urine culture, which takes an average of 1.5 to 2 days from urine collection to results, delaying optimal therapy. Faster, but accurate, alternatives are needed. Light scatter technology has been proposed for several years as a rapid screening tool, whereby negative specimens are excluded from culture. A commercially available light scatter device, BacterioScan 216Dx (BacterioScan, Inc.), has recently been advertised for this application. Paired use of mass spectrometry (MS) for bacterial identification and automated-system-based susceptibility testing straight from the light scatter suspension might provide dramatic improvement in times to a result. The present study prospectively evaluated the BacterioScan device, with culture as the reference standard. Positive light scatter specimens were used for downstream rapid matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) MS organism identification and automated-system-based antimicrobial susceptibility testing. Prospective evaluation of 439 urine samples showed a sensitivity of 96.5%, a specificity of 71.4%, and positive and negative predictive values of 45.1% and 98.8%, respectively. MALDI-TOF MS analysis of the suspension after density-based selection yielded a sensitivity of 72.1% and a specificity of 96.9%. Antimicrobial susceptibility testing of the samples identified by MALDI-TOF MS produced an overall categorical agreement of 99.2%. Given the high sensitivity and negative predictive value of results obtained, BacterioScan 216Dx is a reasonable approach for urine screening and might produce negative results in as few as 3 h, with no downstream workup. Paired rapid identification and susceptibility testing might be useful when MALDI-TOF MS results in an organism identification, and it might decrease the time to a result by more than 24 h. PMID:28356414

Prospective Evaluation of Light Scatter Technology Paired with Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Rapid Diagnosis of Urinary Tract Infections.

PubMed

Montgomery, Sandra; Roman, Kiana; Ngyuen, Lan; Cardenas, Ana Maria; Knox, James; Tomaras, Andrew P; Graf, Erin H

2017-06-01

Urinary tract infections are one of the most common reasons for health care visits. Diagnosis and optimal treatment often require a urine culture, which takes an average of 1.5 to 2 days from urine collection to results, delaying optimal therapy. Faster, but accurate, alternatives are needed. Light scatter technology has been proposed for several years as a rapid screening tool, whereby negative specimens are excluded from culture. A commercially available light scatter device, BacterioScan 216Dx (BacterioScan, Inc.), has recently been advertised for this application. Paired use of mass spectrometry (MS) for bacterial identification and automated-system-based susceptibility testing straight from the light scatter suspension might provide dramatic improvement in times to a result. The present study prospectively evaluated the BacterioScan device, with culture as the reference standard. Positive light scatter specimens were used for downstream rapid matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) MS organism identification and automated-system-based antimicrobial susceptibility testing. Prospective evaluation of 439 urine samples showed a sensitivity of 96.5%, a specificity of 71.4%, and positive and negative predictive values of 45.1% and 98.8%, respectively. MALDI-TOF MS analysis of the suspension after density-based selection yielded a sensitivity of 72.1% and a specificity of 96.9%. Antimicrobial susceptibility testing of the samples identified by MALDI-TOF MS produced an overall categorical agreement of 99.2%. Given the high sensitivity and negative predictive value of results obtained, BacterioScan 216Dx is a reasonable approach for urine screening and might produce negative results in as few as 3 h, with no downstream workup. Paired rapid identification and susceptibility testing might be useful when MALDI-TOF MS results in an organism identification, and it might decrease the time to a result by more than 24 h. Copyright © 2017 American Society for Microbiology.

Xpert Flu for point-of-care diagnosis of human influenza in industrialized countries.

PubMed

Salez, Nicolas; Nougairede, Antoine; Ninove, Laetitia; Zandotti, Christine; de Lamballerie, Xavier; Charrel, Rémi N

2014-05-01

Respiratory infections, particularly those caused by influenza viruses, represent the third-most important cause of death in the world due to infectious diseases. Nevertheless, despite the enormous publicity attracted by epidemics due to these viruses, laboratory diagnosis, documentation and recording of respiratory diseases is still unsatisfactory. Available diagnostic tests capable of providing results rapidly are either limited and insufficiently sensitive or highly sensitive and specific but insufficiently rapid. Considerable investment and research efforts have been made towards the development of new diagnostics for influenza A and B viruses and the Xpert(®) Flu assay (Cepheid(®), CA, USA) has emerged as one of the most promising. In this article, we review current knowledge of the Xpert Flu test, discuss its potential value as a point-of-care test and outline the potential leads for future development.

Point of Care Tuberculosis Sero-Diagnosis Kit for Wild Animals: Combination of Proteins for Improving the Diagnostic Sensitivity and Specificity.

PubMed

Veerasami, Maroudam; Venkataraman, K; Karuppannan, Chitra; Shanmugam, Arun Attur; Prudhvi, Mallepaddi Chand; Holder, Thomas; Rathnagiri, Polavarapu; Arunmozhivarman, K; Raj, Gopal Dhinakar; Vordermeier, Martin; Mohana Subramanian, B

2018-03-01

Tuberculosis is a significant problem globally for domestic animals as well as captive and free ranging wild life. Rapid point of care (POC) serology kits are well suited for the diagnosis of TB in wild animals. However, wild animals are invariably exposed to environmental non-pathogenic mycobacterium species with the development of cross reacting antibodies. In the present study, POC TB diagnosis kit was developed using a combination of pathogenic Mycobacteria specific recombinant antigens and purified protein derivatives of pathogenic and non-pathogenic Mycobacteria . To benchmark the TB antibody detection kit, particularly in respect to specificity which could not be determined in wildlife due to the lack of samples from confirmed uninfected animals, we first tested well-characterized sera from 100 M. bovis infected and 100 uninfected cattle. Then we investigated the kit's performance using sera samples from wildlife, namely Sloth Bears (n = 74), Elephants (n = 9), Cervidae (n = 14), Felidae (n = 21), Cape buffalo (n = 2), Wild bear (n = 1) and Wild dog (n = 1).In cattle, a sensitivity of 81% and a specificity of 90% were obtained. The diagnostic sensitivity of the kit was 94% when the kit was tested using known TB positive sloth bear sera samples. 47.4% of the in-contact sloth bears turned seropositive using the rapid POC TB diagnostic kit. Seropositivity in other wild animals was 25% when the sera samples were tested using the kit. A point of care TB sero-diagnostic kit with the combination of proteins was developed and the kit was validated using the sera samples of wild animals.

[Value of Immunohistochemical Methods in Detecting EML4-ALK Fusion Mutations: A Meta-analysis].

PubMed

Liu, Chang; Cai, Lu; Zhong, Diansheng; Wang, Jing

2016-01-01

The fusion between echinoderm microtubule-associated protein 4 (EML4) and anaplastic lymphatic tumor kinase (ALK) rearrangement is present in approximately 5% of non-small cell lung cancer (NSCLC) patients. It has been regarded as another new target gene after epidermal growth factor receptor (EGFR) and K-ras. Figures showed that the disease control rate could reach up to 80% in NSCLC patients with EML4-ALK fusion gene after treated with ALK inhibitors. Thus, exploring an accurate and rapid detecting method is the key in screening NSCLC patients with EML4-ALK expressions. The aim of this study is to analyze the specificity and sensitivity of IHC in detecting EML4-ALK fusion mutations. To evaluate the accuracy and clinical value of this method, and then provide basis for individual molecular therapy of NSCLC patients. Using Pubmed database to search all documents required. The deadline of retrieval was February 25, 2015. Then further screening the articles according to the inclusion and exclusion criteria. Using diagnostic test meta-analysis methods to analyze the sensitivity and specificity of the immunohistochemistry (IHC) method compared with fluorescence in situ hybridization (FISH) method. Eleven literatures were added into the meta analysis, there were 3,234 of total cases. The diagnostic odds ratio (DOR) was 1,135.00 (95%CI: 337.10-3,821.46); the area under curve (AUC) of summary receiver operating characteristic curve (SROC) curve was 0.992,3 (SEAUC=0.003,2), the Q* was 0.964,4 (SEQ*=0.008,7). Immunohistochemical detection of EML4-ALK fusion gene mutation with specific antibody is feasible. It has high sensitivity and specificity. IHC can be a simple and rapid way in screening EML4-ALK fusion gene mutation and exhibits important clinical values.

Evaluation of HIV/AIDS diagnostics kits and formulation of a testing strategy for Pakistan.

PubMed

Waheed, Usman; Hayat, Khizar; Ahmad, Bashir; Waheed, Yasir; Zaheer, Hasan Abbas

2013-04-01

Rapid diagnosis of HIV/AIDS enables the development of prevention and treatment programmes but accurate, reliable and cost effective testing strategies should be used for testing of HIV/AIDS from a large population. To evaluate the performance and effectiveness of three assays for the diagnosis of HIV in comparison with Western blot and to formulate an alternative cost-effective confirmatory approach for HIV diagnosis. 472 specimens (serum) from a Pakistani population were evaluated. Two rapid HIV testing kits (Capillus, SD Bioline) and one ELISA (Vironostika Ag/Ab) kit were used to detect HIV. Results were compared with Western blot against which sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of all HIV assays were assessed. 280/472 (59.3%) of the samples were positive for antibodies against purified HIV-1 viral proteins. The sensitivity of SD Bioline and Vironostika ELISA was 100% (95% CI; 98-100) while that of anti-HIV Capillus™ kit was 94.6% (95% CI; 91-96.8). The specificity of the Vironostika ELISA and anti-HIV Capillus™ kit was 100% (95% CI; 97-100) while specificity of SD Bioline was 98.4% (95% CI; 95-99). PPV was 100% (95% CI; 98-100%) for the anti-HIV Capillus™ and Vironostika ELISA and 98.9% (95% CI; 96-99%) for SD Bioline. NPV for SD Bioline and Vironostika ELISA was 100% (95% CI; 98-100%) and 92.7% for anti-HIV Capillus™ (95% CI; 88-96%). The sensitivity and specificity of all three kits were satisfactory compared to Western blot and could be used for effective diagnosis of HIV/AIDS in Pakistani population. Copyright © 2013 Elsevier B.V. All rights reserved.

Molecular Diagnosis of Invasive Aspergillosis and Detection of Azole Resistance by a Newly Commercialized PCR Kit

PubMed Central

Gabriel, Frédéric; Gaboyard, Manuel; Lagardere, Gaëlle; Audebert, Lucile; Quesne, Gilles; Godichaud, Sandrine; Verweij, Paul E.; Accoceberry, Isabelle

2017-01-01

ABSTRACT Aspergillus fumigatus is the main species responsible for aspergillosis in humans. The diagnosis of aspergillosis remains difficult, and the rapid emergence of azole resistance in A. fumigatus is worrisome. The aim of this study was to validate the new MycoGENIE A. fumigatus real-time PCR kit and to evaluate its performance on clinical samples for the detection of A. fumigatus and its azole resistance. This multiplex assay detects DNA from the A. fumigatus species complex by targeting the multicopy 28S rRNA gene and specific TR34 and L98H mutations in the single-copy-number cyp51A gene of A. fumigatus. The specificity of cyp51A mutation detection was assessed by testing DNA samples from 25 wild-type or mutated clinical A. fumigatus isolates. Clinical validation was performed on 88 respiratory samples obtained from 62 patients and on 69 serum samples obtained from 16 patients with proven or probable aspergillosis and 13 patients without aspergillosis. The limit of detection was <1 copy for the Aspergillus 28S rRNA gene and 6 copies for the cyp51A gene harboring the TR34 and L98H alterations. No cross-reactivity was detected with various fungi and bacteria. All isolates harboring the TR34 and L98H mutations were accurately detected by quantitative PCR (qPCR) analysis. With respiratory samples, qPCR results showed a sensitivity and specificity of 92.9% and 90.1%, respectively, while with serum samples, the sensitivity and specificity were 100% and 84.6%, respectively. Our study demonstrated that this new real-time PCR kit enables sensitive and rapid detection of A. fumigatus DNA and azole resistance due to TR34 and L98H mutations in clinical samples. PMID:28814586

Comprehensive Multiplex One-Step Real-Time TaqMan qRT-PCR Assays for Detection and Quantification of Hemorrhagic Fever Viruses

PubMed Central

Li, Jiandong; Qu, Jing; He, Chengcheng; Zhang, Shuo; Li, Chuan; Zhang, Quanfu; Liang, Mifang; Li, Dexin

2014-01-01

Background Viral hemorrhagic fevers (VHFs) are a group of animal and human illnesses that are mostly caused by several distinct families of viruses including bunyaviruses, flaviviruses, filoviruses and arenaviruses. Although specific signs and symptoms vary by the type of VHF, initial signs and symptoms are very similar. Therefore rapid immunologic and molecular tools for differential diagnosis of hemorrhagic fever viruses (HFVs) are important for effective case management and control of the spread of VHFs. Real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) assay is one of the reliable and desirable methods for specific detection and quantification of virus load. Multiplex PCR assay has the potential to produce considerable savings in time and resources in the laboratory detection. Results Primers/probe sets were designed based on appropriate specific genes for each of 28 HFVs which nearly covered all the HFVs, and identified with good specificity and sensitivity using monoplex assays. Seven groups of multiplex one-step real-time qRT-PCR assays in a universal experimental system were then developed by combining all primers/probe sets into 4-plex reactions and evaluated with serial dilutions of synthesized viral RNAs. For all the multiplex assays, no cross-reactivity with other HFVs was observed, and the limits of detection were mainly between 45 and 150 copies/PCR. The reproducibility was satisfactory, since the coefficient of variation of Ct values were all less than 5% in each dilution of synthesized viral RNAs for both intra-assays and inter-assays. Evaluation of the method with available clinical serum samples collected from HFRS patients, SFTS patients and Dengue fever patients showed high sensitivity and specificity of the related multiplex assays on the clinical specimens. Conclusions Overall, the comprehensive multiplex one-step real-time qRT-PCR assays were established in this study, and proved to be specific, sensitive, stable and easy to serve as a useful tool for rapid detection of HFVs. PMID:24752452

Prospective evaluation of commercial antibody-based rapid tests in combination with a loop-mediated isothermal amplification PCR assay for detection of Orientia tsutsugamushi during the acute phase of scrub typhus infection.

PubMed

Blacksell, Stuart D; Paris, Daniel H; Chierakul, Wirongrong; Wuthiekanun, Vanaporn; Teeratakul, Achara; Kantipong, Pacharee; Day, Nicholas P J

2012-03-01

Samples from 160 prospectively recruited febrile patients with typhus-like illness in an area of Thailand (Chiang Rai, northern Thailand) where scrub typhus is endemic were used to evaluate the diagnostic capabilities of four rapid immunochromatographic tests (ICTs) for the detection of Orientia tsutsugamushi IgM and total antibodies during acute scrub typhus infection. Of the 160 cases, 54 (34%) had been confirmed to have scrub typhus using the reference scrub typhus infection criteria (STIC), i.e., positive cell culture isolation, an admission IgM antibody titer of ≥1:12,800, a 4-fold rising IgM antibody titer, and/or positivity for ≥2 out of 3 PCR gene targets). The ICTs gave the following sensitivities and specificities: the Panbio IgM ICT, 46% (95% confidence interval [CI], 33 to 60) and 95% (95% CI, 89 to 98), respectively; the Standard Diagnostics IgM ICT, 68% (95% CI, 60 to 75) and 73% (95% CI, 68 to 78), respectively; the AccessBio IgM ICT, 56% (95% CI, 48 to 63) and 90% (95% CI, 87 to 94), respectively; and the AccessBio total antibody ABt ICT, 61% (95% CI, 53 to 68) and 68% (95% CI, 63 to 73), respectively. An isothermal loop amplification (LAMP) PCR assay for scrub typhus demonstrated a sensitivity of 52% (95% CI, 38 to 66) and a specificity of 94% (95% CI, 88 to 98). This study has revealed the diagnostic limitations of antibody-based assays in an acute care setting. However, the combination of ICTs with LAMP usually increased sensitivity with a minimal reduction in specificity. The best combination, the Panbio IgM ICT and LAMP, resulted in a sensitivity of 67% (95% CI, 53 to 79) and a specificity of 91% (95% CI, 83 to 95). The combination of antibody-based assays with DNA- or antigen-based tests shows promise for improved diagnostic sensitivity.

Loop-Mediated Isothermal Amplification Label-Based Gold Nanoparticles Lateral Flow Biosensor for Detection of Enterococcus faecalis and Staphylococcus aureus

PubMed Central

Wang, Yi; Li, Hui; Wang, Yan; Zhang, Lu; Xu, Jianguo; Ye, Changyun

2017-01-01

The report describes a simple, rapid and sensitive assay for visual and multiplex detection of Enterococcus faecalis and Staphylococcus aureus based on multiple loop-mediated isothermal amplification (mLAMP) and lateral flow biosensor (LFB). Detection and differentiation of the Ef0027 gene (E. faecalis-specific gene) and nuc gene (S. aureus-specific gene) were determined using fluorescein (FITC)-and digoxin-modified primers in the mLAMP process. In the presence of biotin- and FITC-/digoxin-modified primers, the mLAMP yielded numerous biotin- and FITC-/digoxin-attached duplex products, which were detected by LFB through biotin/streptavidin interaction (biotin on the duplex and streptavidin on the gold nanoparticle) and immunoreactions (FITC/digoxin on the duplex and anti-FITC/digoxin on the LFB test line). The accumulation of gold nanoparticles generated a characteristic red line, enabling visual and multiplex detection of target pathogens without instrumentation. The limit of detection (LoD), analytical specificity and feasibility of LAMP-LFB technique were successfully examined in pure culture and blood samples. The entire procedure, including specimen (blood samples) processing (30 min), isothermal reaction (40 min) and result reporting (within 2 min), could be completed within 75 min. Thus, this assay offers a simple, rapid, sensitive and specific test for multiplex detection of E. faecalis and S. aureus strains. Furthermore, the LAMP-LFB strategy is a universal technique, which can be extended to detect various target sequences by re-designing the specific LAMP primers. PMID:28239371

Prospective validation of rapid plasma genotyping as a sensitive and specific tool for guiding lung cancer care

PubMed Central

Sacher, Adrian G.; Paweletz, Cloud; Dahlberg, Suzanne E.; Alden, Ryan S.; O’Connell, Allison; Feeney, Nora; Mach, Stacy L.; Jänne, Pasi A.; Oxnard, Geoffrey R.

2016-01-01

Importance Plasma genotyping of cell-free DNA (cfDNA) has the potential to allow for rapid noninvasive genotyping while avoiding the inherent shortcomings of tissue genotyping and repeat biopsies. Objective To prospectively validate plasma droplet digital PCR (ddPCR) for the rapid detection of common EGFR and KRAS mutations as well as the EGFR T790M acquired resistance mutation. Design Eligible patients underwent an initial blood draw and immediate plasma ddPCR for EGFR exon 19 del, L858R, T790M and/or KRAS G12X between July 2014 and June 2015. All patients underwent biopsy for tissue genotyping which was used as the reference standard for comparison; rebiopsy was required for patients with acquired resistance to EGFR kinase inhibitors. Test turnaround time (TAT) was measured in business days from blood draw until test reporting. Setting National Cancer Institute (NCI) designated comprehensive cancer center. Participants Advanced non-squamous NSCLC patients that are either (i) newly diagnosed and planned for initial therapy or (ii) have developed acquired resistance to an EGFR kinase inhibitor and are planned for re-biopsy. Main Outcome Measure Plasma ddPCR assay sensitivity, specificity and TAT. Results 180 patients were enrolled in the study (120 newly diagnosed, 60 with acquired resistance). Tumor genotype included 80 EGFR exon 19/L858R mutants, 35 EGFR T790M, 25 KRAS G12X mutants. Median TAT for plasma ddPCR was 3 days. Tissue genotyping median TAT was 12 days for newly diagnosed patients and 27 days for acquired resistance patients. Plasma ddPCR exhibited a PPV of 100% (95%CI 91-100%) for EGFR 19 del, 100% (95%CI 85-100%) L858R and 100% (95%CI 79-100%) for KRAS, but lower for T790M at 79% (95%CI 62-91%). Sensitivity of plasma ddPCR was 82% (95%CI 69-91%) for EGFR 19 del, 74% (95%CI 55-88%) for L858R and 77% (95%CI 60-90%) for T790M but lower for KRAS at 64% (95%CI 43-82%). Sensitivity for EGFR or KRAS was higher in patients with multiple metastatic sites (p=0.001) and those with hepatic (p=0.001) or bone metastases (p=0.004), specifically. Conclusion Plasma ddPCR detects EGFR and KRAS mutations rapidly with the high specificity needed to select therapy and avoid repeat biopsies. This assay may also detect EGFR T790M missed by tissue genotyping due to tumor heterogeneity in resistant disease. This is the first prospective study to demonstrate the utility of ddPCR-based plasma genotyping in advanced NSCLC. PMID:27055085

Molecular beacon probes-base multiplex NASBA Real-time for detection of HIV-1 and HCV.

PubMed

Mohammadi-Yeganeh, S; Paryan, M; Mirab Samiee, S; Kia, V; Rezvan, H

2012-06-01

Developed in 1991, nucleic acid sequence-based amplification (NASBA) has been introduced as a rapid molecular diagnostic technique, where it has been shown to give quicker results than PCR, and it can also be more sensitive. This paper describes the development of a molecular beacon-based multiplex NASBA assay for simultaneous detection of HIV-1 and HCV in plasma samples. A well-conserved region in the HIV-1 pol gene and 5'-NCR of HCV genome were used for primers and molecular beacon design. The performance features of HCV/HIV-1 multiplex NASBA assay including analytical sensitivity and specificity, clinical sensitivity and clinical specificity were evaluated. The analysis of scalar concentrations of the samples indicated that the limit of quantification of the assay was <1000 copies/ml for HIV-1 and <500 copies/ml for HCV with 95% confidence interval. Multiplex NASBA assay showed a 98% sensitivity and 100% specificity. The analytical specificity study with BLAST software demonstrated that the primers do not attach to any other sequences except for that of HIV-1 or HCV. The primers and molecular beacon probes detected all HCV genotypes and all major variants of HIV-1. This method may represent a relatively inexpensive isothermal method for detection of HIV-1/HCV co-infection in monitoring of patients.

Meat authentication: a new HPLC-MS/MS based method for the fast and sensitive detection of horse and pork in highly processed food.

PubMed

von Bargen, Christoph; Brockmeyer, Jens; Humpf, Hans-Ulrich

2014-10-01

Fraudulent blending of food products with meat from undeclared species is a problem on a global scale, as exemplified by the European horse meat scandal in 2013. Routinely used methods such as ELISA and PCR can suffer from limited sensitivity or specificity when processed food samples are analyzed. In this study, we have developed an optimized method for the detection of horse and pork in different processed food matrices using MRM and MRM(3) detection of species-specific tryptic marker peptides. Identified marker peptides were sufficiently stable to resist thermal processing of different meat products and thus allow the sensitive and specific detection of pork or horse in processed food down to 0.24% in a beef matrix system. In addition, we were able to establish a rapid 2-min extraction protocol for the efficient protein extraction from processed food using high molar urea and thiourea buffers. Together, we present here the specific and sensitive detection of horse and pork meat in different processed food matrices using MRM-based detection of marker peptides. Notably, prefractionation of proteins using 2D-PAGE or off-gel fractionation is not necessary. The presented method is therefore easily applicable in analytical routine laboratories without dedicated proteomics background.

Evaluation of Wondfo influenza A&B fast test based on immunochromatography assay for rapid diagnosis of influenza A H1N1.

PubMed

Peng, Yunping; Wu, Junlin; Liu, Xiaoyun; Wang, Jihua; Li, Wenmei

2013-01-01

Influenza viruses cause significant morbidity and mortality in both children and adults during local outbreaks or epidemics. Therefore, a rapid test for influenza A&B would be useful. This study was conducted to evaluate the clinical performance of the Wondfo influenza A&B test for rapid diagnosis of influenza A H1N1 Infection. The rapid testing assay could distinguish infection of influenza A and B virus. The reference viral strains were cultured in MDCK cells while TCID50 if the viruses were determined. The analytical sensitivity of the Wondfo kit was 100TCID50/ml. The Wondfo kit did not show cross reactivity with other common viruses. 1928 suspected cases of influenza A (H1N1) virus infection were analyzed in the Wondfo influenza A&B test and other commercially available products. Inconsistent results were further confirmed by virus isolation in cell culture. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were 100%, 98.23%, 92.45%, and 100% for flu A, and 96.39%, 99.95%, 98.77%, and 99.84% for flu B respectively. 766 suspected cases of influenza A (H1N1) virus infection were analyzed in the Wondfo influenza A&B test and RT-PCR. The sensitivity, specificity, PPV and NPV were 56.5%, 99.75%, 99.52% and 71.04% for flu A, 25.45%, 99.86%, 93.33% and 94.54% for flu B respectively. These results indicate that the Wondfo influenza A&B test has high positive and negative detection rates. One hundred fifty-six specimens of influenza A (H1N1) confirmed by RT-PCR were analyzed by the Wondfo influenza A&B test and 66.67% were positive while only 18.59% were positive by the reference kit. These results indicate that our rapid diagnostic assay may be useful for analyzing influenza A H1N1 infections in patient specimen. Copyright © 2013 Elsevier Editora Ltda. All rights reserved.

Sensitivity and Specificity of a Prototype Rapid Diagnostic Test for the Detection of Trypanosoma brucei gambiense Infection: A Multi-centric Prospective Study.

PubMed

Bisser, Sylvie; Lumbala, Crispin; Nguertoum, Etienne; Kande, Victor; Flevaud, Laurence; Vatunga, Gedeao; Boelaert, Marleen; Büscher, Philippe; Josenando, Theophile; Bessell, Paul R; Biéler, Sylvain; Ndung'u, Joseph M

2016-04-01

A major challenge in the control of human African trypanosomiasis (HAT) is lack of reliable diagnostic tests that are rapid and easy to use in remote areas where the disease occurs. In Trypanosoma brucei gambiense HAT, the Card Agglutination Test for Trypanosomiasis (CATT) has been the reference screening test since 1978, usually on whole blood, but also in a 1/8 dilution (CATT 1/8) to enhance specificity. However, the CATT is not available in a single format, requires a cold chain for storage, and uses equipment that requires electricity. A solution to these challenges has been provided by rapid diagnostic tests (RDT), which have recently become available. A prototype immunochromatographic test, the SD BIOLINE HAT, based on two native trypanosomal antigens (VSG LiTat 1.3 and VSG LiTat 1.5) has been developed. We carried out a non-inferiority study comparing this prototype to the CATT 1/8 in field settings. The prototype SD BIOLINE HAT, the CATT Whole Blood and CATT 1/8 were systematically applied on fresh blood samples obtained from 14,818 subjects, who were prospectively enrolled through active and passive screening in clinical studies in three endemic countries of central Africa: Angola, the Democratic Republic of the Congo and the Central African Republic. One hundred and forty nine HAT cases were confirmed by parasitology. The sensitivity and specificity of the prototype SD BIOLINE HAT was 89.26% (95% confidence interval (CI) = 83.27-93.28) and 94.58% (95% CI = 94.20-94.94) respectively. The sensitivity and specificity of the CATT on whole blood were 93.96% (95% CI = 88.92-96.79) and 95.91% (95% CI = 95.58-96.22), and of the CATT 1/8 were 89.26% (95% CI = 83.27-93.28) and 98.88% (95% CI = 98.70-99.04) respectively. After further optimization, the prototype SD BIOLINE HAT could become an alternative to current screening methods in primary healthcare settings in remote, resource-limited regions where HAT typically occurs.

Development and testing of monoclonal antibody-based rapid immunodiagnostic test kits for direct detection of Vibrio cholerae O139 synonym Bengal.

PubMed

Hasan, J A; Huq, A; Nair, G B; Garg, S; Mukhopadhyay, A K; Loomis, L; Bernstein, D; Colwell, R R

1995-11-01

We report on the development and testing of two monoclonal antibody-based rapid immunodiagnostic test kits, BengalScreen, a coagglutination test, and Bengal DFA, a direct fluorescent-antibody test, for direct detection of Vibrio cholerae O139 synonym Bengal in clinical and environmental specimens. The BengalScreen test requires less than 5 min to complete and can be used in the field. Bengal DFA, being more sensitive than BengalScreen, requires only one reagent and less than 20 min for detection and enumeration of V. cholerae O139 synonym Bengal. In tests for specificity, all 40 strains of V. cholerae O139 reacted with both test kits, whereas 157 strains of heterologous species examined did not, yielding 100% specificity in this study. A field trial was conducted in with both BengalScreen and Bengal DFA, and the results were compared with those obtained by conventional culture methods. BengalScreen demonstrated a sensitivity of 95%, a specificity of 100%, a positive predictive value of 100%, and a negative predictive value of 94%. Results obtained by Bengal DFA, on the other hand, were 100% sensitive and 100% specific and yielded 100% positive and negative predictive values compared with culture methods. In a second evaluation, 93 stool specimens from Mexico that were negative for V. cholerae O139 by culture were also tested with both the BengalScreen and Bengal DFA kits. None of the 93 specimens were positive for V. cholerae O139 by both tests. A concentration method was optimized for screening of environmental water samples for V. cholerae O139 synonym Bengal with rapid test kits. BengalScreen results were unequivocally positive when water samples contained at least 2.0 x 10(3) CFU/ml, whereas Bengal DFA demonstrated an unequivocally positive reaction when the water sample contained at least 1.5 x 10(2) CFU/ml. When Bengal DFA was compared with conventional culture methods for enumeration of V. cholerae O139 synonym Bengal organisms, no difference was observed.