GroEL Chaperonin-Based Assay for Early Diagnosis of Scrub Typhus

A point-of-care diagnostic for early and rapid diagnosis of scrub typhus caused by Orientia tsutsugamushi is required for prompt and proper treatment of patients presenting with undifferentiated febrile illnesses. In this study, an immunochromatographic antigen detection test kit (ICT AgTK) that targets the highly conserved O. tsutsugamushi 60 kDa GroEL chaperonin (heat shock protein 60) was developed. E. coli-derived recombinant GroEL expressed from DNA coding for the consensus sequence of 32 GroEL gene sequences extracted from the GenBank database was used to immunize rabbits and mice. Rabbit polyclonal antibodies (pAb) were used for preparing a gold-pAb conjugate, and the rGroEL-specific mouse monoclonal antibody was used as the antigen detection reagent at the ICT test line. In-house validation revealed that the ICT AgTK gave 85, 100 and 95% diagnostic sensitivity, specificity and accuracy, respectively, compared to the combined clinical features and standard IFA when tested on 40 frozen serum samples. The test kits correctly identified 10 scrub typhus samples out of 15 fresh plasma/buffy coat samples of patients with febrile illnesses. For independent laboratory validation, the ICT AgTK was sent to one provincial hospital. The ICT AgTK utilized by the hospital medical technologist correctly identified six scrub typhus samples out of 20 serum samples of patients with fever, as confirmed by specific IgM/IgG detection by IFA. The ICT AgTK is easy to perform with rapid turn-around time. It has the potential to be used as an important tool for on-site and early scrub typhus diagnosis by allowing testing of freshly collected samples (serum, plasma or buffy coat), especially in resource-limited healthcare settings.

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Prevalence of scrub typhus in a tertiary care centre in Telangana, south India

Iranian Journal of Microbiology ◽

10.18502/ijm.v12i3.3237 ◽

2020 ◽

Author(s):

Ram Mohan Mylavarapu Venkata Naga Lakshmi ◽

Teja Vijay Dharma ◽

Sukanya Sudhaharan ◽

Subbalaxmi Malladi Venkata Surya ◽

Rajkiran Emmadi ◽

...

Keyword(s):

Scrub Typhus ◽

Point Of Care ◽

Tertiary Care ◽

Delayed Presentation ◽

Serum Samples ◽

Tertiary Care Centre ◽

Doxycycline Treatment ◽

Clinical Records ◽

High Index Of Suspicion ◽

Point Of Care Tests

Background and Objectives: Scrub typhus is re-emerging as an important cause of acute undifferentiated fever in the last decade from various parts of India. Complexity in performing the “gold standard” immunofluorescent assay and the unre- liable nature of Weil Felix test often results in delayed or misdiagnosis in a majority of cases. The present study seeks to integrate the results of rapid diagnostic tests, clinical and laboratory features to aid the diagnosis and management of scrub typhus patients. Materials and Methods: A total of 645 serum samples with suspected scrub typhus sent to the Department of Microbiology were included in the study. Scrub typhus was tested by rapid immunochromatographic test (SD Diagnostics) and IgM ELI- SA (Inbios International, USA). Clinical features, laboratory parameters and final outcome were analysed from the clinical records of positive patients. Results: Scrub typhus was diagnosed in 13.7% of patients and majority of them were observed in the month of August. 58.6% of scrub typhus patients presented with fever of one to two weeks duration. Eschar was documented in 13.7% of patients and 24% of patients gave a history of working outdoors or exposure to vegetation. All the patients responded to Doxycycline treatment and there was no mortality. Conclusion: High index of suspicion for scrub typhus is necessary in febrile patients not responding to conventional anti- biotics especially during outbreak situations. Rapid immunochromatographic tests with excellent specificity and acceptable sensitivity can be used as potential point of care tests for quick diagnosis of scrub typhus especially in delayed presentation.

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Seroprevalence and Clinical Features of Scrub Typhus among Febrile Patients Attending a Referral Hospital in Kathmandu, Nepal

Tropical Medicine and Infectious Disease ◽

10.3390/tropicalmed6020078 ◽

2021 ◽

Vol 6 (2) ◽

pp. 78

Author(s):

Anil Pokhrel ◽

Binod Rayamajhee ◽

Saroj Khadka ◽

Sandeep Thapa ◽

Samjhana Kapali ◽

...

Keyword(s):

Blood Serum ◽

Clinical Presentation ◽

Scrub Typhus ◽

Serological Test ◽

Demographic Data ◽

Outbreak Detection ◽

Serum Samples ◽

Diagnostic Dilemma ◽

Serological Tests ◽

Test Kit

(1) Background: Scrub typhus (ST) is endemic to Nepal. It is often underdiagnosed and misdiagnosed due to non-specific clinical presentation coupled with limited microbiological facilities, leading to adverse clinical outcomes. This study aimed to assess the seroprevalence of scrub typhus in febrile patients attending Sukraraj Tropical and Infectious Disease Hospital (STIDH), Nepal, from August 2018 to April 2019. (2) Materials and Method: Blood/serum samples and clinical and demographic data of adult febrile patients (≥19 years) who attended or were referred to the hospital were collected after obtaining written informed consent from the participants excluding immunocompromised individuals. Collected blood/serum samples were subjected to hematological, biochemical, and serological tests. A serological test for scrub typhus was performed using the ImmuneMed scrub typhus rapid diagnostic test kit. Data generated were analyzed using SPSS software version 24.0. (3) Results: Amongst the 2070 febrile patients, 462 (22.3%) were seropositive to at least one etiological agent of febrile illnesses (scrub typhus: 253 cases, dengue: 101 cases, leptospirosis: 9, brucellosis: 52, malaria: 9 and kala-azar: 20 cases). Scrub typhus accounted for 12.2% (n = 253) of total febrile illnesses followed by dengue (4.9%, n = 101). Mixed seropositivity of scrub typhus with dengue, brucellosis, and typhoid was found in 12 (0.6%), 9 (0.4%), and 5 (0.2%) cases, respectively. Among 253 scrub typhus patients, 53.4% were female. Among the 154 patients, the most common symptoms were fever (100%), headache (79.2%), sweating (70.1%), breathing difficulty (51.3%), redness of the eye (43.5%), and pathognomonic eschar was observed in 9.1% patients. Fifty percent of scrub typhus patients had low platelet count and >30% of patients had an elevated level of liver enzymes (such as serum glutamic oxaloacetic transaminase (SGPT) and serum glutamic pyruvic transaminase (SGOT). (4) Conclusion: Scrub typhus is a considerable cause of febrile illness in Nepal. Females apparently have a higher chance of acquiring scrub typhus. ST presents nonspecific clinical presentation. The diagnostic dilemma of typhus patients can be minimized by the early monitoring of ST-associated symptoms. The country’s health system needs to be strengthened for early outbreak detection, and immediate response actions against scrub typhus to control the future outbreak of ST.

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Evaluation of a point-of-care immunoassay test kit ‘StrongStep’ for cryptococcal antigen detection

PLoS ONE ◽

10.1371/journal.pone.0190652 ◽

2018 ◽

Vol 13 (1) ◽

pp. e0190652 ◽

Cited By ~ 10

Author(s):

Edward Mpoza ◽

Liliane Mukaremera ◽

Didas Atwebembere Kundura ◽

Andrew Akampurira ◽

Tonny Luggya ◽

...

Keyword(s):

Point Of Care ◽

Antigen Detection ◽

Cryptococcal Antigen ◽

Test Kit

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Ferromagnetic Resonance Biosensor for Homogeneous and Volumetric Detection of DNA

10.26434/chemrxiv.5662315.v1 ◽

2017 ◽

Author(s):

Bo Tian ◽

Peter Svedlindh ◽

Mattias Strömberg ◽

Erik Wetterskog

Keyword(s):

Magnetic Nanoparticles ◽

Ferromagnetic Resonance ◽

Limit Of Detection ◽

Point Of Care ◽

Rolling Circle Amplification ◽

Resonance Field ◽

Isothermal Amplification ◽

Detection Sensitivity ◽

Serum Samples ◽

Rolling Circle

In this work, we demonstrate for the first time, a ferromagnetic resonance (FMR) based homogeneous and volumetric biosensor for magnetic label detection. Two different isothermal amplification methods, i.e., rolling circle amplification (RCA) and loop-mediated isothermal amplification (LAMP) are adopted and combined with a standard electron paramagnetic resonance (EPR) spectrometer for FMR biosensing. For RCA-based FMR biosensor, binding of RCA products of a synthetic Vibrio cholerae target DNA sequence gives rise to the formation of aggregates of magnetic nanoparticles. Immobilization of nanoparticles within the aggregates leads to a decrease of the net anisotropy of the system and a concomitant increase of the resonance field. A limit of detection of 1 pM is obtained with an average coefficient of variation of 0.16%, which is superior to the performance of other reported RCA-based magnetic biosensors. For LAMP-based sensing, a synthetic Zika virus target oligonucleotide is amplified and detected in 20% serum samples. Immobilization of magnetic nanoparticles is induced by their co-precipitation with Mg2P2O7 (a by-product of LAMP) and provides a detection sensitivity of 100 aM. The fast measurement, high sensitivity and miniaturization potential of the proposed FMR biosensing technology makes it a promising candidate for designing future point-of-care devices.

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Faculty Opinions recommendation of Point-of-care urine lipoarabinomannan antigen detection for diagnosis of tuberculosis in children.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.736725267.793565963 ◽

2019 ◽

Author(s):

Anthony Harries ◽

Kudakwashe C Takarinda

Keyword(s):

Point Of Care ◽

Antigen Detection ◽

Tuberculosis In Children

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A Label and Probe-Free Zika Virus Immunosensor Prussian Blue@carbon Nanotube-Based for Amperometric Detection of the NS2B Protein

Biosensors ◽

10.3390/bios11050157 ◽

2021 ◽

Vol 11 (5) ◽

pp. 157

Author(s):

Bárbara V. M. Silva ◽

Marli T. Cordeiro ◽

Marco A. B. Rodrigues ◽

Ernesto T. A. Marques ◽

Rosa F. Dutra

Keyword(s):

Carbon Nanotube ◽

Prussian Blue ◽

Zika Virus ◽

Point Of Care ◽

Amperometric Detection ◽

Cross Reactivity ◽

Structural Protein ◽

Serum Samples ◽

Polypyrrole Composite ◽

Congenital Disabilities

Zika virus (ZIKV) is a mosquito-borne infection, predominant in tropical and subtropical regions causing international concern due to the ZIKV disease having been associated with congenital disabilities, especially microcephaly and other congenital abnormalities in the fetus and newborns. Development of strategies that minimize the devastating impact by monitoring and preventing ZIKV transmission through sexual intercourse, especially in pregnant women, since no vaccine is yet available for the prevention or treatment, is critically important. ZIKV infection is generally asymptomatic and cross-reactivity with dengue virus (DENV) is a global concern. An innovative screen-printed electrode (SPE) was developed for amperometric detection of the non-structural protein (NS2B) of ZIKV by exploring the intrinsic redox catalytic activity of Prussian blue (PB), incorporated into a carbon nanotube–polypyrrole composite. Thus, this immunosensor has the advantage of electrochemical detection without adding any redox-probe solution (probe-less detection), allowing a point-of-care diagnosis. It was responsive to serum samples of only ZIKV positive patients and non-responsive to negative ZIKV patients, even if the sample was DENV positive, indicating a possible differential diagnosis between them by NS2B. All samples used here were confirmed by CDC protocols, and immunosensor responses were also checked in the supernatant of C6/36 and in Vero cell cultures infected with ZIKV.

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A rapid point-of-care assay accurately measures vitamin D

Journal of Endocrinological Investigation ◽

10.1007/s40618-021-01575-8 ◽

2021 ◽

Author(s):

K. Albrecht ◽

J. Lotz ◽

L. Frommer ◽

K. J. Lackner ◽

G. J. Kahaly

Keyword(s):

Vitamin D ◽

Metabolic Pathways ◽

Point Of Care ◽

Immunofluorescence Assay ◽

Laboratory Method ◽

Controlled Study ◽

Mean Deviation ◽

Serum Samples ◽

Assay Validation ◽

Accuracy And Precision

Abstract Purpose Vitamin D (VitD) is a pleiotropic hormone with effects on a multitude of systems and metabolic pathways. Consequently, the relevance of a sufficiently high VitD serum level becomes self-evident. Methods A rapid immunofluorescence assay designed for the point-of-care measurement of serum VitD3 solely was tested. Inter- and intra-assay validation, double testing and result comparison with a standardized laboratory method were performed. Results An overall linear correlation of r = 0.89 (Pearson, 95% CI 0.88–0.92, p < 0.01) between the point of care and the conventional reference assay was registered. Accuracy and precision were of special interest at cut-points (10 ng/ml [mean deviation 1.7 ng/ml, SD 1.98 ng/ml, SE 0.16 ng/ml], 12 ng/ml [MD 0.41, SD 1.89, SE 0.19] and 30 ng/ml [MD − 1.11, SD 3.89, SE 0.35]). Only a slight deviation was detected between the two assays when using fresh (r = 0.91, 95% CI 0.86–0.94, p < 0.01) and frozen serum samples (r = 0.86, 0.82–0.89, p < 0.01). Results remained steady when samples were frozen several times. Inter- and intra-assay validation according to the CLSI protocol as well as multiuser testing showed stable results. Conclusion This novel, innovative, and controlled study indicates that the evaluated rapid point of care VitD assay is reliable, accurate, and suited for clinical practice.

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Comparative application of antigen detection enzyme-linked immunosorbent assay and buffy coat parasitological technique for diagnosis of bovine trypanosomosis in Nigeria

Nigerian Veterinary Journal ◽

10.4314/nvj.v25i2.3469 ◽

2005 ◽

Vol 25 (2) ◽

Author(s):

IF Ijagbone ◽

GO Esuruoso ◽

SA Agbede

Keyword(s):

Immunosorbent Assay ◽

Buffy Coat ◽

Antigen Detection ◽

Enzyme Linked Immunosorbent Assay ◽

Bovine Trypanosomosis

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Occurrence ofBordetellaInfection in Pigs in Northern India

International Journal of Microbiology ◽

10.1155/2014/238575 ◽

2014 ◽

Vol 2014 ◽

pp. 1-6 ◽

Cited By ~ 3

Author(s):

Sandeep Kumar ◽

Bhoj R. Singh ◽

Monika Bhardwaj ◽

Vidya Singh

Keyword(s):

Plasmid Profile ◽

Antigen Detection ◽

Bordetella Bronchiseptica ◽

Atrophic Rhinitis ◽

Serum Samples ◽

Northern India ◽

Antimicrobial Sensitivity ◽

Nasal Swabs ◽

Species Specific ◽

Almost All

Bordetella bronchisepticainfection causing atrophic rhinitis in pigs is reported from almost all countries. In the present study, occurrence ofBordetellainfection in apparently healthy pigs was determined in 392 pigs sampled to collect 358 serum samples and 316 nasal swabs from Northern India by conventional bacterioscopy, detection of antigen with multiplex polymerase chain reaction (mPCR), and detection of antibodies with microagglutination test (MAT) and enzyme linked immune-sorbent assay (ELISA).Bordetella bronchisepticacould be isolated from six (1.92%) nasal swabs. Although isolates varied significantly in their antimicrobial sensitivity, they had similar plasmid profile. The genus specific and species specific amplicons were detected from 8.2% and 4.4% nasal swabs using mPCR withalcgene (genus specific) andflagene andfim2 gene (species specific) primers, respectively. Observations revealed that there may be other bordetellae infecting pigs because about 50% of the samples positive using mPCR for genus specific amplicons failed to confirm presence ofB. bronchiseptica. Of the pig sera tested with MAT and ELISA forBordetellaantibodies, 67.6% and 86.3% samples, respectively, were positive. For antigen detection mPCR was more sensitive than conventional bacterioscopy while for detection of antibodies neither of the two tests (MAT and ELISA) had specificity in relation to antigen detection. Study indicated high prevalence of infection in swine herds in Northern India.

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