scholarly journals Evaluation of In-House and Commercial Serological Tests for Diagnosis of Human Tularemia

2017 ◽  
Vol 56 (1) ◽  
Author(s):  
Hadjila Yanes ◽  
Aurélie Hennebique ◽  
Isabelle Pelloux ◽  
Sandrine Boisset ◽  
Dominique J. Bicout ◽  
...  
Keyword(s):  
Francisella Tularensis ◽  
Immunofluorescence Assay ◽  
Screening Tests ◽  
Clinical Samples ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serological Tests ◽  
Content Type ◽  
Microagglutination Test ◽  
Confirmatory Tests ◽  
National Reference Center

ABSTRACTTularemia is a zoonosis caused by the bacteriumFrancisella tularensis. Its specific diagnosis remains based on serological methods, whileF. tularensisis rarely detected in clinical samples by culture or PCR. The aim of the present study was to evaluate the performance of the Serion enzyme-linked immunosorbent assay (ELISA) classicFrancisella tularensisIgG and IgM tests (Virion/Serion GmbH Institute, Würzburg, Germany) and the VIRapid tularemia immunochromatographic test (ICT) (Vircell, Granada, Spain) compared to that of the in-house microagglutination test (MAT) and indirect immunofluorescence assay (IFA) currently used at the French National Reference Center forFrancisella. We evaluated 256 consecutive sera from 208 patients, including 51 confirmed and 23 probable tularemia cases, and 134 control patients not infected withF. tularensis. The IFA tests displayed 72.5% sensitivity for IgM (cutoff titer ≥80) and 74.5% for IgG (cutoff titer ≥160), and 99.3% specificity for both IgM and IgG. Using cutoffs advocated by the manufacturer, the Serion ELISAs displayed 88.2% sensitivity for IgM and 86.3% for IgG antibodies; specificity was 94.8% for IgM and 95.5% for IgG. Compared to MAT and IFA tests, the Serion ELISAs allowed earlier detection of specific antibodies (1 to 2 weeks versus 2 to 3 weeks after the onset of symptoms). The ICT sensitivity and specificity were 90% and 83.6%, respectively, when considering the cutoff advocated by the manufacturer. In conclusion, the Serion ELISAs are useful as screening tests for tularemia diagnosis, but additional confirmatory tests (such as MAT and IFA) are needed, especially in areas of low endemicity.

scholarly journals Detection of Francisella tularensis-Specific Antibodies in Patients with Tularemia by a Novel Competitive Enzyme-Linked Immunosorbent Assay

2012 ◽  
Vol 20 (1) ◽  
pp. 9-16 ◽  
Author(s):  
Neekun Sharma ◽  
Akitoyo Hotta ◽  
Yoshie Yamamoto ◽  
Osamu Fujita ◽  
Akihiko Uda ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Francisella Tularensis ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
The Novel ◽  
Serum Samples ◽  
Content Type ◽  
Microagglutination Test ◽  
Highly Sensitive ◽  
High Degree

ABSTRACTA novel competitive enzyme-linked immunosorbent assay (cELISA) was developed and evaluated for detection of antibodies againstFrancisella tularensisin humans. The assay is based on the ability of serum antibodies to inhibit the binding of monoclonal antibodies (MAbs) directed againstF. tularensislipopolysaccharide antigens. The assay was evaluated using serum samples of tularemia patients, inactivatedF. tularensis-immunized rabbits, andF. tularensis-infected mice. Antibodies againstF. tularensiswere successfully detected in serum samples of tularemia patients as well as the immunized and infected animals. The cELISA method was compared to indirect ELISA (iELISA) and the commonly used microagglutination test (MA) using serum samples of 19 tularemia patients and 50 healthy individuals. The sensitivity and specificity of cELISA were 93.9 and 96.1%, respectively, in comparison to the iELISA. MA was less sensitive than cELISA with a sensitivity and specificity of only 81.8 and 98.0%, respectively. A high degree of correlation (R2= 0.8226) was observed between cELISA and iELISA results. The novel cELISA developed in this study appears to be highly sensitive and specific for serodiagnosis of human tularemia. The potential of the MAb-based cELISA to be used in both human and animal samples emphasizes its usefulness for serological survey of tularemia among multiple animal species.

scholarly journals Development and Comparative Evaluation of a Plate Enzyme-Linked Immunosorbent Assay Based on Recombinant Outer Membrane Antigens Omp28 and Omp31 for Diagnosis of Human Brucellosis

2013 ◽  
Vol 20 (8) ◽  
pp. 1217-1222 ◽  
Author(s):  
Sapana Tiwari ◽  
Ashu Kumar ◽  
Duraipandian Thavaselvam ◽  
Smita Mangalgi ◽  
Vedika Rathod ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Immunosorbent Assay ◽  
Expression System ◽  
Disease Diagnosis ◽  
Cross Reactivity ◽  
Clinical Samples ◽  
Enzyme Linked Immunosorbent Assay ◽  
Human Brucellosis ◽  
Serological Tests ◽  
Content Type

ABSTRACTBrucellosis is an important zoonotic infectious disease of humans and livestock with worldwide distribution and is caused by bacteria of the genusBrucella. The diagnosis of brucellosis always requires laboratory confirmation by either isolation of pathogens or detection of specific antibodies. The conventional serological tests available for the diagnosis of brucellosis are less specific and show cross-reactivity with other closely related organisms. These tests also necessitate the handling ofBrucellaspecies for antigen preparation. Therefore, there is a need to develop reliable, rapid, and user-friendly systems for disease diagnosis and alternatives to vaccine approaches. Keeping in mind the importance of brucellosis as an emerging infection and the prevalence in India, we carried out the present study to compare the recombinant antigens with the native antigens (cell envelope and sonicated antigen) ofBrucellafor diagnosis of human brucellosis by an indirect plate enzyme-linked immunosorbent assay (ELISA). Recombinant outer membrane protein 28 (rOmp28) and rOmp31 antigens were cloned, expressed, and purified in the bacterial expression system, and the purified proteins were used as antigens. Indirect plate ELISAs were then performed and standardized for comparison of the reactivities of recombinant and native antigens against the 433 clinical samples submitted for brucellosis testing, 15 culture-positive samples, and 20 healthy donor samples. The samples were separated into four groups based on their positivity to rose bengal plate agglutination tests (RBPTs), standard tube agglutination tests (STATs), and 2-mercaptoethanol (2ME) tests. The sensitivities and specificities of all the antigens were calculated, and the rOmp28 antigen was found to be more suitable for the clinical diagnosis of brucellosis than the rOmp31 antigen and native antigens. The rOmp28-based ELISA showed a very high degree of agreement with the conventional agglutination tests and promising results for further use in clinical screening and serodiagnosis of human brucellosis.

scholarly journals Evaluation of a Recombinant Trypanosoma cruzi Mucin-Like Antigen for Serodiagnosis of Chagas' Disease

2011 ◽  
Vol 18 (11) ◽  
pp. 1850-1855 ◽  
Author(s):  
Claudia R. De Marchi ◽  
Javier M. Di Noia ◽  
Alberto C. C. Frasch ◽  
Vicente Amato Neto ◽  
Igor C. Almeida ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Chagas Disease ◽  
The United States ◽  
Cross Reactivity ◽  
Immunofluorescence Assay ◽  
Health Concern ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serological Tests ◽  
Small Surface ◽  
Content Type

ABSTRACTChagas' disease is caused by the protozoan parasiteTrypanosoma cruziand is one of the most important endemic problems in Latin America. Lately, it has also become a health concern in the United States and Europe. Currently, a diagnosis of Chagas' disease and the screening of blood supplies for antiparasite antibodies are achieved by conventional serological tests that show substantial variation in the reproducibility and reliability of their results. In addition, the specificity of these assays is curtailed by antigenic cross-reactivity with sera from patients affected by other endemic diseases, such as leishmaniasis. Here we used a highly sensitive chemiluminescent enzyme-linked immunosorbent assay (CL-ELISA) to evaluate a recombinant protein core of a mucin-like molecule (termed trypomastigote small surface antigen [TSSA]) for the detection of specific serum antibodies in a broad panel of human sera. The same samples were evaluated by CL-ELISA using as the antigen either a mixture of nativeT. cruzitrypomastigote mucins or an epimastigote extract and, for further comparison, by conventional serologic tests, such as an indirect hemagglutination assay and indirect immunofluorescence assay. TSSA showed ∼87% sensitivity among the seropositive Chagasic panel, a value which was increased up to >98% when only parasitologically positive samples were considered. More importantly, TSSA showed a significant increase in specificity (97.4%) compared to those of currently used assays, which averaged 80 to 90%. Overall, our data demonstrate that recombinant TSSA may be a useful antigen for the immunodiagnosis of Chagas' disease.

scholarly journals The Trypomastigote Small Surface Antigen from Trypanosoma cruzi Improves Treatment Evaluation and Diagnosis in Pediatric Chagas Disease

2017 ◽  
Vol 55 (12) ◽  
pp. 3444-3453 ◽  
Author(s):  
Virginia Balouz ◽  
Luciano J. Melli ◽  
Romina Volcovich ◽  
Guillermo Moscatelli ◽  
Samanta Moroni ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Chagas Disease ◽  
Surface Antigen ◽  
Rapid Assessment ◽  
Drug Efficacy ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serological Tests ◽  
Small Surface ◽  
Content Type

ABSTRACTChagas disease is caused by the protozoan parasiteTrypanosoma cruzi. Assessment of parasitological cure upon treatment with available drugs relies on achieving consistent negative results in conventional parasitological and serological tests, which may take years to assess. Here, we evaluated the use of a recombinantT. cruziantigen termed trypomastigote small surface antigen (TSSA) as an early serological marker of drug efficacy inT. cruzi-infected children. A cohort of 78 pediatric patients born toT. cruzi-infected mothers was included in this study. Only 39 of the children were infected withT. cruzi, and they were immediately treated with trypanocidal drugs. Serological responses against TSSA were evaluated in infected and noninfected populations during the follow-up period using an in-house enzyme-linked immunosorbent assay (ELISA) and compared to conventional serological methods. Anti-TSSA antibody titers decreased significantly faster than anti-whole parasite antibodies detected by conventional serology both inT. cruzi-infected patients undergoing effective treatment and in those not infected. The differential kinetics allowed a significant reduction in the required follow-up periods to evaluate therapeutic responses or to rule out maternal-fetal transmission. Finally, we present the case of a congenitally infected patient with an atypical course in whom TSSA provided an early marker forT. cruziinfection. In conclusion, we showed that TSSA was efficacious both for rapid assessment of treatment efficiency and for early negative diagnosis in infants at risk of congenitalT. cruziinfection. Based upon these findings we propose the inclusion of TSSA for refining the posttherapeutic cure criterion and other diagnostic needs in pediatric Chagas disease.

scholarly journals Laboratory Confirmation of Lyme Disease

1991 ◽  
Vol 2 (2) ◽  
pp. 64-69 ◽  
Author(s):  
Tom G Schwan ◽  
Warren J Simpson ◽  
Patricia A Rosa
Keyword(s):  
Lyme Disease ◽  
Immunofluorescence Assay ◽  
Detection Methods ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serological Tests ◽  
Specificity And Sensitivity ◽  
Laboratory Confirmation ◽  
Staining Methods ◽  
Alternative Detection ◽  
Species Specific

Lyme disease can be confirmed in the laboratory by isolation or detection of its causative agent, a tick-borne spirocheteBorrelia burgdorferi,or by a diagnostic change in the titre of antibodies specific to the agent.B burgdorferican be isolated and cultivated in Barbour-Stoenner-Kelly II medium. It can be detected by light microscopy in tissue sections or, rarely, in blood smears using various staining methods. There is interest in the development of alternative detection methods, including identification of specific antigens ofB burgdorferiin the urine of suspected cases and demonstration of the presence of species-specific DNA using polymerase chain reaction assays. Currently, serological tests (indirect immunofluorescence assay, enzyme-linked immunosorbent assay and Western immunoblot) are the most practical and available methods for confirming Lyme disease. The quest to improve the specificity and sensitivity of serological tests – for example, through the use of specific recombinant antigens – continues.

scholarly journals A Novel Real-Time PCR-Based Screening Test with Pooled Fecal Samples for Bovine Johne’s Disease

2020 ◽  
Vol 58 (12) ◽  
Author(s):  
Satoko Kawaji ◽  
Reiko Nagata ◽  
Yasutaka Minegishi ◽  
Yumi Saruyama ◽  
Akiko Mita ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Johne's Disease ◽  
Johne’S Disease ◽  
Enzyme Linked Immunosorbent Assay ◽  
Pcr Assay ◽  
Serological Tests ◽  
Fecal Samples ◽  
Content Type ◽  
Cattle Herds

ABSTRACT Johne’s disease (JD) is an economically important infectious disease in livestock farming caused by Mycobacterium avium subsp. paratuberculosis. As an alternative to serological tests, which are used mainly for the screening of whole herds, we developed a novel ResoLight-based real-time PCR (RL-PCR) assay with pooled fecal samples for the detection of fecal shedders in cattle herds. The RL-PCR assay included an internal amplification control (IC) which was amplified using the same primer pair as the target molecule M. avium subsp. paratuberculosis IS900 and differentiated based on melting temperatures. Individual fecal suspensions were pooled and concentrated by centrifugation to avoid a loss of sensitivity by the dilution effect. Combined with a DNA extraction kit (Johne-PureSpin; FASMAC), no inhibition of PCR amplification was observed with up to 15 fecal samples in a pool. The detection limit of RL-PCR at a pool size of 10 was 10 M. avium subsp. paratuberculosis organisms per gram of feces, which was comparable to that of individual testing. A total of 2,654 animals in 12 infected herds were screened by individual antibody-enzyme-linked immunosorbent assay (ELISA) and the RL-PCR assay using pooled feces. Fifty animals were diagnosed with JD through the screening by RL-PCR, compared with only 5 by ELISA (which were also positive in RL-PCR). In 7 JD-free herds, the results of 4 out of 327 pools (1.2%) were invalid due to the lack of IC amplification, and then animals were confirmed negative individually. Our results suggest that implementation of herd screening by pooled RL-PCR would advance the monitoring and control of JD in cattle herds.

scholarly journals Evaluation of Toxoplasma ELITe MGB Real-Time PCR Assay for Diagnosis of Toxoplasmosis

2017 ◽  
Vol 55 (5) ◽  
pp. 1369-1376 ◽  
Author(s):  
Florence Robert-Gangneux ◽  
Marie-Pierre Brenier-Pinchart ◽  
Hélène Yera ◽  
Sorya Belaz ◽  
Emmanuelle Varlet-Marie ◽  
...  
Keyword(s):  
Amniotic Fluid ◽  
Congenital Toxoplasmosis ◽  
Clinical Samples ◽  
Repeated Dna ◽  
Pcr Assay ◽  
Content Type ◽  
National Reference ◽  
Pcr Assays ◽  
National Reference Center ◽  
Commercial Kit

ABSTRACT Molecular diagnosis of congenital toxoplasmosis or disseminated toxoplasmosis is based mainly on PCR. The repeated DNA element rep529 has become the main DNA target used in most PCR methods, whether laboratory developed or commercial. In this multicenter study, we evaluated the Toxoplasma ELITe MGB (Elitech) commercial kit by comparison with three reference quantitative PCR assays (RAs) used routinely in three proficient laboratories of the French National Reference Center for Toxoplasmosis network, using Toxoplasma calibrated suspensions diluted to obtain a range of concentrations from 0.1 to 10,000 parasites/ml. These suspensions were extracted with either the DNA extraction kit (EXTRAblood; Elitech) recommended by the manufacturer or the QIAamp DNA minikit (Qiagen). The Toxoplasma ELITe MGB assay was also evaluated on a panel of 128 clinical samples, including 56 amniotic fluid samples, 55 placenta samples, and various other samples, of which 95 originated from patients with proven toxoplasmosis. The ELITe MGB assay amplified low-concentration replicates (<10 parasites/ml) of calibrated suspensions less frequently than the RAs of 2/3 laboratories. Additionally, the combination of EXTRAblood and Toxoplasma ELITe MGB yielded poorer sensitivity than the combination of QIAamp DNA minikit and ELITe MGB for low parasite concentrations ( P < 0.001 for 1 parasite/ml). On clinical samples, the sensitivity and the specificity of the commercial assay were 89% and 100%, respectively. The sensitivity ranged from 79% (placenta samples) to 100% (amniotic fluid samples). Overall, this study shows that the Toxoplasma ELITe MGB assay is suitable for the diagnosis of toxoplasmosis from non-cell-rich or non-hemoglobin-rich samples and that the EXTRAblood kit is not optimal.

scholarly journals A Simple and Rapid Chromatographic Strip Test for Detection of Antibody to Porcine Reproductive and Respiratory Syndrome Virus

2005 ◽  
Vol 17 (5) ◽  
pp. 469-473 ◽  
Author(s):  
Y. S. Lyoo ◽  
S. B. Kleiboeker ◽  
K.-Y. Jang ◽  
N. K. Shin ◽  
J.-M. Kang ◽  
...  
Keyword(s):  
Immunofluorescence Assay ◽  
Economic Problem ◽  
Clinical Samples ◽  
Respiratory Syndrome Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Laboratory Equipment ◽  
Reduction Strategies ◽  
Antibody Elisa ◽  
Strip Test

Porcine reproductive and respiratory syndrome virus (PRRSV) continues to be a major economic problem for swine industries worldwide despite several disease-reduction strategies such as age-segregated early weaning and all-in-all-out pig movement. Routine diagnosis of PRRSV is carried out by the combined use of an antibody-detecting enzyme-linked immunosorbent assay (ELISA), immunofluorescence, reverse transcription-polymerase chain reaction, and virus isolation. These assays require specialized laboratory equipment in addition to multistep sample handling and sample preparation. The objective of this study was to evaluate a simple pen-side assay (BioSign™ PRRSV) for rapid detection of PRRSV antibody based on a lateral flow chromatographic strip immunoassay system. This assay uses Escherichia coli–expressed viral nucleocapsid protein antigen for detecting antibodies against PRRSV in swine sera. In this report, the authors describe the evaluation of this assay using sera from both clinical samples and experimentally infected piglets. The results were compared with those of a standard, commercially available antibody ELISA (HerdChek®PRRS ELISA) and an indirect immunofluorescence assay using the same serum samples. The BioSign™ PRRSV assay was capable of detecting antibodies in sera known to contain antibodies to PRRSV, resulting in 93.2% sensitivity for samples from experimentally infected pigs and 98.7% sensitivity for clinical serum samples. For sera that did not contain antibodies to PRRSV, the specificity was found to be 98.5% and 99.2% for clinical and experimental serum samples, respectively.

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