Melioidosis is a fatal infectious disease caused by the environmental bacterium
It is highly endemic in Asia and northern Australia but neglected in many other tropical countries. Melioidosis ...patients have a wide range of clinical manifestations, and definitive diagnosis requires bacterial culture, which can be time-consuming. A reliable rapid serological tool is greatly needed for disease surveillance and diagnosis. We previously demonstrated by enzyme-linked immunosorbent assay (ELISA) that a hemolysin-coregulated protein (Hcp1) is a promising target for serodiagnosis of melioidosis. In this study, we developed a rapid immunochromatography test (ICT) using Hcp1 as the target antigen (Hcp1-ICT). We evaluated this test for specific antibody detection using serum samples obtained from 4 groups of human subjects, including the following: (i) 487 culture-confirmed melioidosis patients from four hospitals in northeast Thailand; (ii) 202 healthy donors from northeast Thailand; (iii) 90 U.S. healthy donors; and (iv) 207 patients infected with other organisms. Compared to culture results as a gold standard, the sensitivity of ICT for all hospitals was 88.3%. The specificities for Thai donors and U.S. donors were 86.1% and 100%, respectively, and the specificity for other infections was 91.8%. The results of the Hcp1-ICT demonstrated 92.4% agreement with the Hcp1-ELISA results with a kappa value of 0.829, indicating that the method is much improved compared with the current serological method, the indirect hemagglutination assay (IHA) (69.5% sensitivity and 67.6% specificity for Thais). The Hcp1-ICT represents a potential point-of-care (POC) test and may be used to replace the IHA for screening of melioidosis in hospitals as well as in resource-limited areas.
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the cause of Coronavirus Disease 2019 (COVID-19), poses extraordinary threats and complex challenges to global public health. ...Quantitative measurement of SARS-CoV-2 antibody titer plays an important role in understanding the patient-to-patient variability of immune response, assessing the efficacy of vaccines, and identifying donors for blood transfusion therapy. There is an urgent and ever-increasing demand for serological COVID-19 antibody tests that are highly sensitive, quantitative, rapid, simple, minimally invasive, and inexpensive. In this work, we developed a single-step, wash-free immunoassay for rapid and highly sensitive quantitative analysis of serological human IgG against SARS-CoV-2 which requires only a single droplet of serum. By simply incubating 4 μL human serum samples with antibody-functionalized gold nanoparticles, a photonic crystal optical biosensor coated with the recombinant spike protein serves as a sensing platform for the formation of sandwich immunocomplex through specific antigen–antibody interactions, upon which the detected IgG molecules can be counted with digital precision. We demonstrated a single-step 15-min assay capable of detecting as low as 100 pg mL−1 human COVID-19 IgG in serum samples. The calculated limit of detecting (LOD) and limit of quantification (LOQ) is 26.7 ± 7.7 and 32.0 ± 8.9 pg mL−1, respectively. This work represents the first utilization of the Activate Capture + Digital Counting (AC + DC)-based immunoassay for rapid and quantitative analysis of serological COVID-19 antibody, demonstrating a route toward point-of-care testing, using a portable detection instrument. On the basis of the sandwich immunoassay principle, the biosensing platform can be extended for the multiplexed detection of antigens, additional IgGs, cytokines, and other protein biomarkers.
PRAM-based Digital Immunoassay for Serological COVID-19 IgG. Display omitted
•A one-step, digital immunoassay for rapid analysis of serological COVID-19 antibody by PRAM is developed.•It employs the principle of AC+DC, enabling the quantitative detection of serological human COVID-19 IgG in 15 minutes.•A detection limit of 100 pg mL-1 and a high sensitivity was achieved.•The assay requires only a fingerstick quantity of serum and has great potential for POC testing using a portable PRAM.
La sérologie de la toxoplasmose relève en France de l’obligation légale. Son interprétation théoriquement simple se heurte au quotidien à des difficultés dont les plus courantes sont liées à la non ...standardisation des réactifs pouvant conduire à des interprétations erronées ou à des discordances de statut pour un même patient. La présence d’IgM n’est pas un critère suffisant ni même nécessaire pour conclure à une infection récente. Une bonne communication entre cliniciens et biologistes est indispensable à l’élaboration de conclusions pertinentes.
Toxoplasmosis serology is a legal obligation in France. Its theoretically simple interpretation comes up against difficulties, the most common of which is the non-standardization of reagents, which can lead to faulty interpretations or status discrepancies for the same patient. The presence of IgM is not a sufficient or even necessary criterion to conclude that a recent infection has occurred. Good communication between clinicians and biologists is essential to do relevant conclusions.
Bovine tuberculosis (bTB) is a zoonosis caused by Mycobacterium bovis. Test-and-cull protocols and gross pathological examinations of abattoir animals as well as milk pasteurization have been ...implemented to prevent the spread of tuberculosis from animals to humans worldwide. Despite the importance of precise and rapid diagnostic tests, conventional methods including intradermal skin tests and γ-interferon assays are limited by the high rate of false-negative results for cattle in the late infectious stage and due to laborious and time-consuming procedures. Therefore, antibody detection methods such as enzyme-linked immunosorbent assay (ELISA) are urgently needed to supplement the established approaches and expand the diagnostic window. This study was conducted to develop a bTB ELISA by evaluating recombinant and native proteins and various assay parameters. We produced recombinant MPB70 and SahH (M70S) and a native 20-kDa protein (20K) and optimized the ELISA protocol. The 20K ELISA showed 94.4% sensitivity and 98.2% specificity with an optimal sample-to-positive ratio cut-off of 0.531. The sensitivity and specificity of M70S ELISA were 94.4% and 97.3%, respectively, with an optimal sample-to-negative ratio cut-off of 1.696. Both assays showed acceptable diagnostic efficiency and could be used for bTB diagnosis in combination with established methods for herd screening and to expand the diagnostic window.
Background
Recently, different arboviruses became endemic in Brazil mostly causing acute febrile illnesses, however, neurological manifestations have also been reported. This study aimed to ...investigate which viruses were involved in the meningitis etiology and the contribution of the circulating arboviruses in Salvador, Bahia, Brazil.
Methods
From June 2014 to February 2016, 170 patients with suspected viral meningitis were identified in Couto Maia Hospital, Salvador-BA, Brazil. Their CSF samples were investigated for possible viral etiology by reverse transcription-PCR (RT-PCR) for different arboviruses: DENV, ZIKV and CHIKV; and for the EV; and by PCR for the HHV1-5 complex (HSV1-2, VZV, EBV and CMV). Also, ELISA was carried out in a subgroup of remaining samples for detection of DENV IgM and NS1 antigen, CHIKV IgM and ZIKV IgM.
Results
Thirty-seven patients were PCR or ELISA positive for at least one of the studied viruses (overall positivity 21.8%). EV was the agent most frequently detected (10 cases; 27.0%), along with all four DENV serotypes (10 cases; 27.0%); followed by CHIKV (6 cases; 16.2%), ZIKV (6 cases; 16.2%), and Varicella zoster virus (VZV) (1 case; 2.7%). Four cases (10.8%) presented viral co-infection detected: DENV1 + CHIKV, DENV1 + EV, DENV4 + ZIKV, and CHIKV + ZIKV. Arboviruses (DENV, CHIKV and ZIKV) accounted for the great majority of cases (26 cases; 70.3%) of all single and co-infections: DENV has been the most frequently detected arbovirus (13 cases; 35.1%). Among non-arboviral meningitis, the most common etiology was the EV (11 cases; 29.7%).
Conclusions
Arboviruses accounted for the majority of identified viruses among patients with suspected viral meningitis. In areas where they are endemic it is crucial to increase viral surveillance and consider them in the differential diagnosis of meningitis.
Scrub typhus is a mite-borne rickettsiosis caused by the intracellular bacterium Orientia tsutsugamushi (OTS), which is classified as a biosafety level-3 (BSL-3) pathogen. For serological tests of ...scrub typhus, mouse fibroblast cells infected with the five prevalent serotypes of OTS in Japan are generally used as antigens for indirect immunofluorescent assay (IFA). In this study, sf-9 insect cells infected with the recombinant type-specific antigens (rTSA)-expressing baculovirus were applied for IFA. The paired sera samples of 15 scrub typhus-patients, 10 rickettsiosis-patients, and 10 control individuals were used. Both IgM and IgG IFA titers determined by the rTSA based IFA were correlated with those determined with the OTS-infected cell-based IFA (R 2 = 0.7319 to 0.7956). Based on the criteria for serological diagnosis, such as the suitable cutoff for single serum (IgM ≥ 1:160) and/or significant increase in IgG titer between paired sera (≥ 4 times), all of the 15 scrub typhus patients diagnosed as positive with OTS-infected cell-based IFA were also diagnosed as positive by the rTSA-based IFA, whereas all 10 rickettsiosis patients and 10 control individuals were not. The rTSAs, which can be prepared in BSL-2 laboratories, are efficacious in the serological diagnosis of scrub typhus.
Lajmska borelioza (LB) je bolest koju u Europi najčešće uzrokuju borelije kompleksa Borrelia burgdorferi sensu lato, dok je u Sjevernoj Americi jedino patogena B.burgdorferi sensu stricto. Kliničke ...manifestacije LB su polimorfne. Postavljanje dijagnoze temelji se na kliničkoj slici i epidemiološkim podacima o vjerojatnosti kontakta s krpeljima uz primjenu mikrobiološke dijagnostike. Najčešća je rutinska dijagnostika serologija za određivanje i praćenje dinamike specifičnih protutijela IgM i IgG. Nakon primarnog testiranja, rezultate je potrebno potvrditi dodatnim testom visoke specifičnosti. U područjima visoke prevalencije, specifičnost rezultata visoko osjetljivog i specifičnog testa nije obvezno dodatno potvrđivati. I pozitivni i negativni nalazi moraju se interpretirati klinički. Serološka dijagnostika predstavlja dobrobit za prepoznavanje i liječenje bolesnika samo ako se rezultati interpretiraju temeljem poznavanja patogeneze, kliničke slike i imunosnog odgovora kao i karakteristika korištenog testa. Karakteristike samih borelija i prezentacije antigena, izbjegavanje imunosnog odgovora, dostupnost različitih testova kao i Interneta predstavljaju zamke, posebno ako se slijede neprovjerene informacije.
Humans can be infected by the intracellular parasite Toxoplasma gondii, which causes toxoplasmosis, a common parasitic disease. Although the infection is generally asymptomatic for most adults, ...severe complications may occur in some individuals, especially women in early pregnancy. Serologic diagnosis is used as a routine practice to determine the immune status for infection by T. gondii. In this review, we attempt to provide an overview of the serological diagnosis of toxoplasmosis, including diagnostic strategy, current problems in detection with specific antibodies, and the standardization of T. gondii serological detection.
•We attempt to provide an overview of serological diagnosis of toxoplasmosis.•We reviewed serological diagnostic strategy and current problems in detection for T. gondii infection.•We demonstrate that potential application of chimeric antibodies as the surrogate for serological detection.•The necessary of participation in EQA for clinical laboratories is highlighted.
To diagnose Lyme Borreliosis, it is advised to use an enzyme-linked immunosorbent test to check for serum antibodies specific for Lyme and all tests with positive or ambiguous enzyme-linked ...immunosorbent assay (ELISA) results being confirmed by immunoblot. This method of measuring the humoral immunity in human fluids (e.g., by ELISA) has provided robust and reproducible results for decades and similar assays have been validated for monitoring of B cell immunity. These immunological tests that detect antibodies to Borrelia burgdorferi are useful in the diagnosis of Borreliosis on a routine basis. The variety of different Borrelia species and their different geographic distributions are the main reasons why standards and recommendations are not identical across all geographic regions of the world. In contrast to humoral immunity, the T cell reaction or cellular immunity to the Borrelia infection has not been well elucidated, but over time with more studies a novel T cell-based assay (EliSpot) has been developed and validated for the sensitive detection of antigen-specific T cell responses to B. burgdorferi. The EliSpot Lyme assay can be used to study the T cell response elicited by Borrelia infections, which bridges the gap between the ability to detect humoral immunity and cellular immunity in Lyme disease. In addition, detecting cellular immunity may be a helpful laboratory diagnostic test for Lyme disease, especially for seronegative Lyme patients. Since serodiagnostic methods of the Borrelia infection frequently provide false positive and negative results, this T cell-based diagnostic test (cellular assay) may help in confirming a Lyme diagnosis. Many clinical laboratories are convinced that the cellular assay is superior to the Western Blot assay in terms of sensitivity for detecting the underlying Borrelia infection. Research also suggests that there is a dissociation between the magnitude of the humoral and the T cell-mediated cellular immune responses in the Borrelia infection. Lastly, the data implies that the EliSpot Lyme assay may be helpful to identify Borrelia infected individuals when the serology-based diagnostic fails to do so. Here in this chapter the pairing of humoral and cellular immunity is employed to evaluate the adaptive response in patients.
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