IgG Enzyme-linked Immunosorbent Assay Research Articles

Incidence of Zika Virus Infection from a Dengue Epidemiological Study of Children in Ratchaburi Province, Thailand.

Zika virus (ZIKV) is the mosquito-transmitted virus that the WHO declared a Public Health Emergency of International Concern in 2016 due to the consequence of microcephaly from infected pregnancies. The incidence of Zika infection has been unclear in many countries because most infected people have nonspecific febrile illnesses. This study’s aim is to investigate the incidence of symptomatic Zika virus infections from the archived samples of a dengue cohort study of children in central Thailand from 2006 to 2009. We performed Zika NS1 immunoglobulin (Ig)G enzyme-linked immunosorbent assay (ELISA) screening to identify symptomatic Zika infections in paired acute/convalescent serum samples. Symptomatic Zika infections were confirmed by reverse transcription polymerase chain reactions (RT-PCR) of acute serum samples. The comparison of the Zika NS1 IgG ELISA results between acute and convalescent samples showed 290/955 (30.4%) seropositive cases. Zika RT-PCR results were positive in 28 febrile cases (15 females, 13 males). Zika RT-PCR showed that symptomatic Zika infection occurred in children aged 4–11 years in Ratchaburi province, Thailand (2007–2009, first case in April 2007), and the symptomatic Zika:dengue infection ratio was 28 Zika:394 dengue (1:14). Phylogenetic analysis showed that all Zika viruses were of Asian lineage. Zika NS1 IgG ELISA identified Zika-infected patients and showed a low Zika:dengue ratio.

Evaluation of a SARS-CoV-2 Capture IgM Antibody Assay in Convalescent Sera.

ABSTRACTSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for a global pandemic with over 152 million cases and 3.19 million deaths reported by early May 2021. Understanding the serological response to SARS-CoV-2 is critical to determining the burden of infection and disease (coronavirus disease 2019 [COVID-19]) and transmission dynamics. We developed a capture IgM assay because it should have better sensitivity and specificity than the commonly used indirect assay. Here, we report the development and performance of a capture IgM enzyme-linked immunosorbent assay (ELISA) and a companion indirect IgG ELISA for the spike (S) and nucleocapsid (N) proteins and the receptor-binding domain (RBD) of S. We found that among the IgM ELISAs, the S ELISA was positive in 76% of 55 serum samples from SARS-CoV-2 PCR-positive patients, the RBD ELISA was positive in 55% of samples, and the N ELISA was positive in 15% of samples. The companion indirect IgG ELISAs were positive for S in 89% of the 55 serum samples, RBD in 78%, and N in 85%. While the specificities for IgM RBD, S, and N ELISAs and IgG S and RBD ELISAs were 97% to 100%, the specificity of the N IgG ELISA was lower (89%). RBD-specific IgM antibodies became undetectable by 3 to 6 months, and S IgM reached low levels at 6 months. The corresponding IgG S, RBD, and N antibodies persisted with some decreases in levels over this time period. These capture IgM ELISAs and the companion indirect IgG ELISAs should enhance serologic studies of SARS-CoV-2 infections.IMPORTANCE Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has inflicted tremendous loss of lives, overwhelmed health care systems, and disrupted all aspects of life worldwide since its emergence in Wuhan, China, in December 2019. Detecting current and past infection by PCR or serology is important to understanding and controlling SARS-CoV-2. With increasing prevalence of past infection or vaccination, IgG antibodies are less helpful in diagnosing a current infection. IgM antibodies indicate a more recent infection and can supplement PCR diagnosis. We report an alternative method, capture IgM, to detect serum IgM antibodies, which should be more sensitive and specific than most currently used methods. We describe this capture IgM assay and a companion indirect IgG assay for the SARS-CoV-2 spike (S), nucleocapsid (N), and receptor-binding domain (RBD) proteins. These assays can add value to diagnostic and serologic studies of coronavirus disease 2019 (COVID-19).

A Hemagglutination-Based Semiquantitative Test for Point-of-Care Determination of SARS-CoV-2 Antibody Levels.

ABSTRACTSerologic point-of-care tests to detect antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are an important tool in the COVID-19 pandemic. The majority of current point-of-care antibody tests developed for SARS-CoV-2 rely on lateral flow assays, but these do not offer quantitative information. To address this, we developed a novel antibody test leveraging hemagglutination, employing a dry card format currently used for typing ABO blood groups. Two hundred COVID-19 patient and 200 control plasma samples were reconstituted with O-negative red blood cells (RBCs) to form whole blood and added to dried viral-antibody fusion protein, followed by a stirring step and a tilting step, 3-min incubation, and a second tilting step. The sensitivities of the hemagglutination test, Euroimmun IgG enzyme-linked immunosorbent assay (ELISA), and receptor binding domain (RBD)-based CoronaChek lateral flow assay were 87.0%, 86.5%, and 84.5%, respectively, using samples obtained from recovered COVID-19 individuals. Testing prepandemic samples, the hemagglutination test had a specificity of 95.5%, compared to 97.3% and 98.9% for the ELISA and CoronaChek, respectively. A distribution of agglutination strengths was observed in COVID-19 convalescent-phase plasma samples, with the highest agglutination score (4) exhibiting significantly higher neutralizing antibody titers than weak positives (2) (P < 0.0001). Strong agglutinations were observed within 1 min of testing, and this shorter assay time also increased specificity to 98.5%. In conclusion, we developed a novel rapid, point-of-care RBC agglutination test for the detection of SARS-CoV-2 antibodies that can yield semiquantitative information on neutralizing antibody titer in patients. The 5-min test may find use in determination of serostatus prior to vaccination, postvaccination surveillance, and travel screening.

Susceptibility of cadets and recruits to chickenpox: A seroprevalence study

BackgroundRegular outbreaks of chickenpox are reported among cadets and recruits in the army undergoing training, resulting in morbidity and loss in terms of training days. Vaccination policy in the army mandates that all cadets and recruits be vaccinated against chickenpox on joining the training centers. However, 100% vaccination of trainees is not achieved. Hence, seroprevalence data among cadets and recruits were collected with the aim to assess the need for vaccination against chickenpox. MethodsThe data were collected from cadets and recruits undergoing training in military training centers by simple random sampling. Blood samples were collected from all study participants for IgG enzyme-linked immunosorbent assay for varicella zoster. A structured questionnaire was also used to collect information from all participants. ResultsA total of 2041 cadets and recruits between the ages of 19–29 years were studied of which 96.1% were males. Of all participants, 49.9% were seropositive showing varicella zoster virus (VZV) IgG antibody in the serum, and 29.1% study participants gave positive history of having had varicella in the past. The sensitivity of definite positive history of varicella when compared with presence of VZV IgG antibodies was found to be 55.50% while specificity was found to be 97.17%. ConclusionA very large majority of cadets and recruits that joined the army are susceptible to chickenpox. Hence, the risk of repeated chickenpox outbreaks in these training centers remains very high. As the sensitivity of history of past infection of chickenpox is relatively low, it cannot be used as a tool to determine the susceptibility of the trainees to the infection.

Diagnostic accuracy of a novel enzyme-linked immunoassay for the detection of IgG and IgG4 against Strongyloides stercoralis based on the recombinant antigens NIE/SsIR

BackgroundThe diagnosis of strongyloidiasis is challenging. Serological tests are acknowledged to have high sensitivity, but issues due to cross-reactions with other parasites, native parasite antigen supply and intrinsic test variability do occur. Assays based on recombinant antigens could represent an improvement. The aim of this study was to assess the sensitivity and specificity of two novel immunoglobulin (Ig)G and IgG4 enzyme-linked immunosorbent assays (ELISAs) based on the recombinant antigens NIE/SsIR for the diagnosis of strongyloidiasis.MethodsThis was a retrospective diagnostic accuracy study. We included serum samples collected from immigrants from strongyloidiasis endemic areas for whom there was a matched result for Strongyloides stercoralis on agar plate culture and/or PCR assay, or a positive microscopy for S. stercoralis larvae. For the included samples, results were also available from an in-house indirect fluorescent antibody test (IFAT) and a commercial (Bordier ELISA; Bordier Affinity Products SA) ELISA. We excluded: (i) samples with insufficient serum volume; (ii) samples from patients treated with ivermectin in the previous 6 months; and (iii) sera from patients for whom only routine coproparasitology was performed after formol–ether concentration, if negative for S. stercoralis larvae. The performance of the novel assays was assessed against: (i) a primary reference standard, with samples classified as negative/positive on the basis of the results of fecal tests; (ii) a composite reference standard (CRS), which also considered patients to be positive who had concordant positive results for the IFAT and Bordier ELISA or with a single “high titer” positive result for the IFAT or Bordier ELISA. Samples with a single positive test, either for the IFAT or Bordier ELISA, at low titer, were considered to be “indeterminate,” and analyses were carried out with and without their inclusion.ResultsWhen assessed against the primary reference standard, the sensitivities of the IgG and IgG4 ELISAs were 92% (95% confidence interval [CI]: 88–97%) and 81% (95% CI: 74–87%), respectively, and the specificities were 91% (95% CI: 88–95%) and 94% (95% CI: 91–97%), respectively. When tested against the CRS, the IgG ELISA performed best, with 78% sensitivity (95% CI: 72–83%) and 98% specificity (95% CI: 96–100%), when a cut-off of 0.675 was applied and the indeterminate samples were excluded from the analysis.ConclusionThe NIE-SsIR IgG ELISA demonstrated better accuracy than the IgG4 assay and was deemed promising particularly for serosurveys in endemic areas.Graphical abstract

Seroepidemiologic survey of emerging vector-borne infections in South Korean forest/field workers.

With global warming and lush forest change, vector-borne infections are expected to increase in the number and diversity of agents. Since the first report of severe fever with thrombocytopenia syndrome (SFTS) in 2013, the number of reported cases has increased annually in South Korea. However, although tick-borne encephalitis virus (TBEV) was detected from ticks and wild rodents, there is no human TBE case report in South Korea. This study aimed to determine the seroprevalence of TBEV and SFTS virus (SFTSV) among forest and field workers in South Korea. From January 2017 to August 2018, a total 583 sera were obtained from the forest and field workers in South Korea. IgG enzyme-linked immunosorbent assay (ELISA) and neutralization assay were conducted for TBEV, and indirect immunofluorescence assay (IFA) and neutralization assay were performed for SFTSV. Seroprevalence of TBEV was 0.9% (5/583) by IgG ELISA, and 0.3% (2/583) by neutralization assay. Neutralizing antibody against TBEV was detected in a forest worker in Jeju (1:113) and Hongcheon (1:10). Only 1 (0.2%) forest worker in Yeongju was seropositive for SFTSV by IFA (1:2,048) and neutralizing antibody was detected also. In conclusion, this study shows that it is necessary to raise the awareness of physicians about TBEV infection and to make efforts to survey and diagnose vector-borne diseases in South Korea.

Characterization and evaluation of a recombinant multiepitope peptide antigen MAG in the serological diagnosis of Toxoplasma gondii infection in pigs

BackgroundToxoplasmosis caused by Toxoplasma gondii is a serious disease threatening human and animal health. People can be infected with T. gondii by ingesting raw pork contaminated with cysts or oocysts. Serological test is a sensitive and specific method usually used for large-scale diagnosis of T. gondii infection in humans and animals (such as pigs). Commercial pig Toxoplasma antibody ELISA diagnostic kits are expensive, which limits their use; moreover, the wide antigen composition used in these diagnostic kits is still unclear and difficult to standardize. The multiepitope peptide antigen is a novel diagnostic marker, and it has potential to be developed into more accurate and inexpensive diagnostic kits.MethodsThe synthetic multiepitope antigen (MAG) cDNA encoding a protein with epitopes from five T. gondii-dominant antigens (SAG1, GRA1, ROP2, GRA4, and MIC3) was designed, synthesized, and expressed in Escherichia coli BL21 (DE3) strain. The recombinant protein was detected through western blot with pig anti-T. gondii-positive and -negative serum, and then IgG enzyme-linked immunosorbent assay (ELISA) named MAG-ELISA was designed. The MAG-ELISA was evaluated in terms of specificity, sensitivity, and stability. The MAG-ELISA was also compared with a commercial PrioCHECK®Toxoplasma Ab porcine ELISA (PrioCHECK ELISA). Finally, the trend of pig anti-T. gondii IgG levels after artificial infection with RH tachyzoites was evaluated using MAG-ELISA and two other ELISA methods (rMIC3-ELISA and PrioCHECK ELISA).ResultsMAG antigen could be specifically recognized by pig anti-T. gondii-positive but not -negative serum. MAG-ELISA showed high diagnostic performance in terms of specificity (88.6%) and sensitivity (79.1%). MAG-ELISA could be used for detecting anti-T. gondii IgG in the early stage of T. gondii infection in pigs (at least 7 days after artificial infection).ConclusionsOur results suggest that MAG antigen can be applied to specifically recognize anti-T. gondii IgG in pig, and MAG-ELISA has the potential for large-scale screening tests of T. gondii infection in pig farms and intensive industries.Graphical abstract

The Potential of O’nyong-nyong Virus Strain SG650 Murine Monoclonal Antibodies for Detection of O’nyong-nyong and Chikungunya Viruses

O’nyong-nyong virus (ONNV) and Chikungunya virus (CHIKV) are antigenically related alphaviruses responsible for febrile illnesses common to the tropics and associated with relatively high morbidity and mortality. Murine monoclonal antibodies (mAbs) targeting alphaviruses like Chikungunya have been developed and used to make commercially available kits. However, few studies have been conducted to develop antibodies specific to ONNV and no commercial kits are available for use in endemic regions where outbreak potential is high. We demonstrate the potential of in-house generated monoclonal antibodies against ONNV to detect both ONNV and CHIKV. The objective of this study was to generate mAbs using hybridoma technology, characterize the developed mAbs, determine their specificity against selected alphaviruses and check their diagnostic potential using an indirect IgG enzyme-linked immunosorbent assay (ELISA) and focus neutralization assay (FRNT50). BALB/c mice were immunized with ONNV purified proteins from ONNV infectious culture fluid. After four rounds of booster injections, the mice were sacrificed, spleen cells harvested and fused with parental myeloma cells then cultured in selective media and the successful hybrid clones with antibody-producing ability purified to yield the desired mAbs. Five monoclonal antibodies targeting the ONNV E1 protein of isotypes IgG2a/kappa, IgG2b/kappa and IgM/kappa (P1B12, P1E9, P1G11, P1B4 and P1G6) demonstrated a potential to detect both ONNV and CHIKV isolates by indirect IgG ELISA but no potential for neutralization of the viruses by FRNT50. This study demonstrates the potential efficacy of in-house serological tools as an alternative in the absence of commercial assays in screening and diagnosis of ONN and CHIK viruses which are often co-circulating. It is our recommendation that this work may be pursued further to design and optimize ELISA assays, using the developed mAbs, for the detection of both ONN and CHIK viruses in the research laboratory set-up

Evaluation of VIDAS® Diagnostic Assay Prototypes Detecting Dengue Virus NS1 Antigen and Anti-Dengue Virus IgM and IgG Antibodies.

Dengue is a serious tropical disease caused by the mosquito-borne dengue virus (DENV). Performant, rapid, and easy-to-use assays are needed for the accurate diagnosis of acute DENV infection. We evaluated the performance of three prototype assays developed for the VIDAS® automated platform to detect dengue NS1 antigen and anti-dengue IgM and IgG antibodies. Positive and negative agreement with competitor enzyme-linked immunosorbent assays (ELISA) and rapid diagnostic tests (RDT) was evaluated in 91 Lao patients (57 adults, 34 children) with acute DENV infection. The VIDAS® NS1 assay showed the best overall agreement (95.6%) with the competitor NS1 ELISA. Both VIDAS® NS1 and NS1 ELISA assays also demonstrated high sensitivity relative to DENV RNA RT-PCR set as gold standard (85.7% and 83.9%, respectively). In contrast, NS1 RDT was less sensitive relative to DENV RNA RT-PCR (72.7%). The overall agreement of VIDAS® IgM and IgG assays with the competitor assays was moderate (72.5% for IgM ELISA, 76.9% for IgG ELISA, and 68.7% for IgM and IgG RDT). In most analyses, test agreements of the VIDAS® assays were comparable in adults and children. Altogether, the VIDAS® dengue prototypes performed very well and appear to be suitable for routine detection of dengue NS1 antigen and anti-dengue IgM/IgG antibodies.

Use of COVID-19 Serologic Testing in Healthcare Workers with Acute Respiratory Tract Infection

Background: Diagnostic tests for COVID-19 are in high demand. Serologic assays are of interest as diagnostic adjuncts to SARS-CoV-2 quantitative polymerase chain reaction (PCR); however, many of the commercially available assays have limited validation data and clinical utility is unknown. We describe the utilization of SARS-CoV-2 IgG enzyme-linked immunosorbent assay (ELISA) for healthcare workers with acute respiratory tract infection (ARTI) who underwent SARS-CoV-2 PCR testing. Methods: The MetroHealth System is the largest public hospital system in Ohio, employing ~8,000 staff. COVID-19 detection began in early March 2020. EDI novel coronavirus COVID-19 IgG ELISA (KT-1032) targeting antibody response to viral nucleocapsid was obtained for diagnostic and seroprevalence analyses. Manufacturer reports of sensitivity and specificity of the assay are 100% and 99%, respectively. A 2-part test strategy for employees with symptoms of ARTI was implemented. Qualifying symptoms for SARS-CoV-2 PCR testing included fever and either cough or shortness of breath. Additional symptoms were included to reflect expanding knowledge of COVID-19. Employees who underwent SARS-CoV-2 PCR testing (Luminex ARIES) were offered serologic testing on day 14 following PCR result. Education accompanied the offer for serologic testing as well as the receipt of test result to aide interpretation. Results: From April 16, 2020, through July 6, 2020, 588 employees underwent PCR testing. Overall, 70 cases of COVID-19 were detected. Of the 197 employees who opted for serologic testing, IgG positivity was 12.6%. The mean time to IgG collection following PCR result was 30 days (range, 10–79). Using PCR results obtained in the clinical setting of ARTI as the diagnostic gold standard, IgG was 84.6% sensitive and 98.2% specific (Figure 1). Conclusions: In a population of symptomatic healthcare workers, SARS-CoV2 IgG testing was specific for COVID-19 diagnosis. Sensitivity was inadequate compared to the positive predictive agreement of 90% or greater required for US Food and Drug Administration emergency use authorization. In a low-prevalence environment for COVID-19 (&lt;5%), a positive SARS-CoV-2 IgG has a low positive predictive value, which may falsely imply immunity and may negatively affect infection prevention practices.Funding: NoDisclosures: None

Filariasis

Introduction: Expatriate workers usually contribute to the origination of some infectious diseases in the countries they reside. The aim of this study was to identify and analyze available information on filariasis imported into Najran, Saudi Arabia, by expatriate workers. Methodology: Serum (n = 497) were acquired from expatriate workers of both genders from different nationalities and various ages and also from randomly chosen parts and at different time periods of Najran. The serum samples were stored in a deep freezer at a temperature of − 65°C until utilized for the serological diagnosis of filariasis by using filariasis immunoglobulin G/immunoglobulin M (IgG/IgM) combo rapid test and human filariasis antibody (IgG4) enzyme-linked immunosorbent assay (ELISA). Results: Based on IgG/IgM rapid test, the prevalence of filariasis IgG was 3.8% (19 out of 497), whereas IgG4 ELISA showed that the prevalence of filariasis was 10.9% (10 out of 92). The highest prevalence of filariasis occurred in Indian workers, in adulthood (30–49 years), and in male workers. There was no significant agreement in results between IgG/IgM rapid test and ELISA. The percentage specificity of IgG/IgM rapid test, relative to filariasis IgG4 ELISA was 80.5%. Conclusions: The possibility of health hazard due to imported filariasis and preeminently the initiation of a novel carrier focal point of the disease are likely to relied upon the abundance of carriers of microfilaria and the density of vector mosquitoes who feed on the carriers. Furthermore, the results of this study underscore the need for screening and therapeutic strategies that guarantee the control and prevention of parasitic infectious diseases.

The seroprevalence of the newly identified porcine circovirus type 4 in China investigated by an enzymed-linked immunosorbent assay.

Porcine circovirus 4 (PCV4) is the fourth porcine circovirus newly identified in China, and it could be detected in diseased and healthy pigs. To date, the prevalence of PCV4 DNA in pig herds has been investigated in many provinces from both China and Korea, with positive rates varied from 3.28% to 25.4% in samples from different regions. However, up to now no serological data have been reported to evaluate the prevalence of PCV4 in pig herds. In this study, an indirect anti-PCV4 IgG enzyme-linked immunosorbent assay (ELISA) based on replicase protein (Rep) was developed and utilized to investigate the seroprevalence of PCV4 in pig herds of China. A total of 1790 swine serum samples from 17 provinces of China were tested including samples confirmed positive for PCV4 DNA. There was no cross-reactivity of this ELISA with PCV1, PCV2 and PCV3. PCV4 Rep antibodies have been detected in serum samples from 16 out of 17 provinces in China. The PCV4 overall seroprevalence was 43.97%, with the highest of 67.8% been detected in sows, followed by fattening and suckling pigs with positive rates of 35.0% and 14.56%, respectively, and the lowest of 12.61% been detected in nursery pigs. Moreover, from the present data, the earliest positive sample could be retrieved to at least 2008. The present study provides an overall seroprevalence of PCV4 in China, and is helpful to understand the prevalence of PCV4 in the pig herds since it was discovered.

Spinal fluid IgG antibodies from patients with demyelinating diseases bind multiple sclerosis-associated bacteria

A panel of 10 IgG enzyme-linked immunosorbent assays (ELISAs) were developed for the detection of anti-microbial immune responses in the cerebrospinal fluid (CSF) of patients with demyelinating diseases (DD). The anti-microbial ELISA assays follow on prior human brain tissue RNA sequencing studies that established multiple sclerosis (MS) microbial candidates. Lysates included in the ELISA panel were derived from Akkermansia muciniphila, Atopobium vaginae, Bacteroides fragilis, Lactobacillus paracasei, Odoribacter splanchnicus, Pseudomonas aeruginosa, Cutibacterium (Propionibacterium) acnes, Fusobacterium necrophorum, Porphyromonas gingivalis, and Streptococcus mutans. CSF responses from patients with demyelinating diseases (DD, N = 14) were compared to those with other neurological diseases (OND, N = 8) and controls (N = 13). Commercial positive and negative control CSF specimens were run with each assay. ELISA index values were derived for each specimen against each of the 10 bacterial lysates. CSF reactivity was significantly higher in the DD group compared to the controls against Akkermansia, Atopobium, Bacteroides, Lactobacillus, Odoribacter, and Fusobacterium. Four of the 11 tested DD group subjects had elevated antibody indexes against at least one of the 10 bacterial species, suggesting intrathecal antibody production. This CSF serological study supports the hypothesis that several of the previously identified MS candidate microbes contribute to demyelination in some patients.Key messagesA panel of 10 IgG enzyme-linked immunosorbent assays (ELISAs) were developed for the detection of anti-microbial immune responses in the cerebrospinal fluid (CSF) of patients with demyelinating diseases, including multiple sclerosis and acute disseminated encephalomyelitis.CSF reactivity was significantly higher in the demyelination group compared to the controls against the bacteria Akkermansia, Atopobium, Bacteroides, Lactobacillus, Odoribacter, and Fusobacterium.Several of the demyelination subjects had elevated antibody indexes against at least one of the 10 antigens, suggesting at least limited intrathecal production of anti-bacterial antibodies.This CSF serological study supports the hypothesis that several of the previously identified MS candidate microbes contribute to demyelination in some patients.

Exposure to tick-borne encephalitis virus among nature management workers in the Netherlands

Tick-borne encephalitis virus (TBEV) has only recently been detected in the Netherlands. With still few autochthonous tick-borne encephalitis (TBE) patients, human exposure to TBEV is expected to be very low among the general population. We aimed to assess the exposure to TBEV among persons with an occupationally high risk of tick bites in the Netherlands.In our cross-sectional serological survey, employees and volunteers of nature management organizations provided a single blood sample and completed an online questionnaire in 2017. The sera were screened in the anti-TBEV IgG Enzyme-Linked Immunosorbent Assay (ELISA), after which a TBEV-specific virus neutralization test (VNT) was applied to confirm positive ELISA outcomes.Ten sera tested positive for IgG antibodies in the TBEV ELISA, among 556 participants who did not report vaccination against TBEV. Through confirmation in VNT, TBEV-specific IgG antibodies were detected among 0.5% (3/556, 95%CI 0.1%–1.6%). During the five years prior to the questionnaire, 87% reported tick bites. Half of the participants considered that most of their tick bites (75% to 100%) had been acquired while being at work.A very low seroprevalence of TBEV exposure was observed among these nature management workers, even though they report a six times higher exposure to tick bites, compared to our general population. Nonetheless, the emergence of TBEV in the Netherlands reaffirms the need for education and preventative measures against tick bites and tick-borne diseases.

Rapid Point-of-Care COVID-19 Diagnosis with a Gold-Nanoarchitecture-Assisted Laser-Scribed Graphene Biosensor.

The global pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus has revealed the urgent need for accurate, rapid, and affordable diagnostic tests for epidemic understanding and management by monitoring the population worldwide. Though current diagnostic methods including real-time polymerase chain reaction (RT-PCR) provide sensitive detection of SARS-CoV-2, they require relatively long processing time, equipped laboratory facilities, and highly skilled personnel. Laser-scribed graphene (LSG)-based biosensing platforms have gained enormous attention as miniaturized electrochemical systems, holding an enormous potential as point-of-care (POC) diagnostic tools. We describe here a miniaturized LSG-based electrochemical sensing scheme for coronavirus disease 2019 (COVID-19) diagnosis combined with three-dimensional (3D) gold nanostructures. This electrode was modified with the SARS-CoV-2 spike protein antibody following the proper surface modifications proved by X-ray photoelectron spectroscopy (XPS) and scanning electron microscopy (SEM) characterizations as well as electrochemical techniques. The system was integrated into a handheld POC detection system operated using a custom smartphone application, providing a user-friendly diagnostic platform due to its ease of operation, accessibility, and systematic data management. The analytical features of the electrochemical immunoassay were evaluated using the standard solution of S-protein in the range of 5.0–500 ng/mL with a detection limit of 2.9 ng/mL. A clinical study was carried out on 23 patient blood serum samples with successful COVID-19 diagnosis, compared to the commercial RT-PCR, antibody blood test, and enzyme-linked immunosorbent assay (ELISA) IgG and IgA test results. Our test provides faster results compared to commercial diagnostic tools and offers a promising alternative solution for next-generation POC applications.

Field Performance of a Rapid Diagnostic Test for the Serodiagnosis of Abdominal Cystic Echinococcosis in the Peruvian Highlands.

We evaluated the performance of a commercial rapid diagnostic test (RDT) in a field setting for the diagnosis of abdominal cystic echinococcosis (CE) using sera collected during an ultrasound population screening in a highly endemic region of the Peruvian Andes. Abdominal CE was investigated by ultrasonography. Sera collected from individuals with abdominal CE (cases) and age- and gender-matched volunteers with no abdominal CE (controls) were tested independently in two laboratories (Peru and Italy) using the VIRapid® HYDATIDOSIS RDT and RIDASCREEN® Echinococcus IgG enzyme-linked immunosorbent assay. Performance indexes of single and serially combined tests were calculated and applied to hypothetical screening and clinical scenarios. Test concordance was also evaluated. Prevalence of abdominal CE was 6.00% (33 of 546) by ultrasound. Serum was obtained from 33 cases and 81 controls. The VIRapid test showed similar sensitivity (76% versus 74%) and lower specificity (79% versus 96%) than results obtained in a hospital setting. RDTs showed better performance when excluding subjects reporting surgery for CE and if weak bands were considered negative. Concordance between tests was moderate to very good. In hypothetical screening scenarios, ultrasound alone or confirmed by RDTs provided more reliable prevalence figures than serology alone, which overestimated it by 5 to 20 times. In a simulation of case diagnosis with pre-test probability of CE of 50%, positive and negative post-test probabilities of the VIRapid test were 78% and 22%, respectively. The application of the VIRapid test alone would not be reliable for the assessment of population prevalence of CE, but could help clinical decision making in resource-limited settings.

POS1195 ASSESSING ANTIBODY STATUS FOR SARS-CoV-2 IN PEOPLE WITH CORONAVIRUS INFECTION: A TIME COURSE STUDY IN PEOPLE WITH AUTOIMMUNE CONDITIONS

Background:With the emergence of the global coronavirus pandemic, increasing concerns have been raised about the course of SARS-CoV-2 infection in people with immune-mediated disorders.Objectives:In this study we aimed to assess the time course of proven SARS-CoV-2 infection, development of humoral immunity with detectable antibodies to the virus and evaluate any changes in antibody titres over time.Methods:We recruited 114 participants in total who had potential symptoms of Covid-19 infection. Participants were recruited from rheumatology or inflammatory bowel disease (IBD) clinics from their records who attended a London Teaching hospital for care. Ethical Approval was in place for the study. Age- and gender- matched control participants without any underlying rheumatological condition/IBD were recruited as a comparator group. Clinical symptoms for Covid-19 infection were assessed using the Covid-19 Rheumatology Global Alliance assessment criteria (https://rheum-covid.org/). Information about Disease Modifying Anti-Rheumatic Drug (DMARD) drug use was also recorded. Participants’ serum samples were collected for quantitative serological assessment of antibodies to SARS-CoV-2 using the Mologic Enzyme-Linked ImmunoSorbent Assay (ELISA) IgG kit. The optical density values were plotted aganist time using a normalisation as previously described (1). A cut-off above 0 indicated positive serology.Results:A total of 114 subjects were recruited into the study. Subjects were recruited if they had suspected Covid symptoms. Of the population recruited, the total number subsequently testing positive for SARS-CoV-2 was n=59 (either on serology or PCR). The number of subjects with autoimmune conditions diagnosed with Covid-19 was 32, with 30 subjects being symptomatic (the asymptomatic 2 subjects were excluded from further analyses). Of the 30 symptomatic subjects, the average age was 56, with a female to male ratio of 19:11. The most prevalent diagnoses were RA (30%), Psoriatic Arthritis (16.7%), SLE (10%), sarcoidosis 10%), Ulcerative Colitis (10%), Crohn’s disease (10%), seronegative inflammatory arthritis (6.7%), Sjogren’s (3.3%) and Juvenile Idiopathic Arthritis (3.3%). The majority were on csDMARDs (60%), biologic DMARDs (20%), tacrolimus (3.3%) and the remaining 16.7% were not on DMARDs. Most subjects required treatment in hospital (56.7%), a smaller number required high dependency care (6.7%) and the rest were treated at home (36.6%). The majority required no oxygen (63.3%), with a further 30% requiring oxygen and 6.7% needed more supportive care i.e. CPAP/ventilation. We also had a matched control group (n=29) of subjects who developed SARS-CoV-2 but had no underlying autoimmune conditions. These subjects had a mean age of 55, female: male ratio 18: 11. Of these 31% had home management, 55.2% had ward level treatments and 13.8% had intensive care treatment. Of the controls, 48.3% did not require oxygen, 37.9% needed simple oxygen and 13.8% were on CPAP. A total of 35 subjects from the original study have attended for follow-up visits. Antibody titres for IgG using the Mologic ELISA detection assay were compared at two visits for the control and autoimmune group. Results showed that subjects with autoimmune conditions and those without who developed SARS-CoV-2 showed very similar antibody titres by optical density (OD) (Figure 1) and maintained antibody responses beyond 6 months in most cases.Conclusion:Serological assays can assist in the understanding of disease severity of SAR-CoV-2 infection. They can be a useful tool for patient surveillance, especially in people who are on immunomodulatory drugs and are being seen in Rheumatology services. Future work is needed to assess the duration and potential protective nature of humoral antibody responses to SARS-CoV-2.

Comparable Accuracies of Nonstructural Protein 1- and Envelope Protein-Based Enzyme-Linked Immunosorbent Assays in Detecting Anti-Dengue Immunoglobulin G Antibodies.

Background: Dengue virus (DENV) infection remains a global public health concern. Enzyme-linked immunosorbent assays (ELISAs), which detect antibodies targeting the envelope (E) protein of DENV, serve as the front-line serological test for presumptive dengue diagnosis. Very few studies have determined the serostatus by detecting antibodies targeting the nonstructural protein 1 (NS1), which can function as diagnostic biomarkers to distinguish natural immunity from vaccine-induced immunity. Methods: We used community-acquired human serum specimens, with the serostatus confirmed by focus reduction microneutralization test (FRμNT), to evaluate the diagnostic performances of two NS1-based ELISA methods, namely, immunoglobulin G antibody-capture ELISA (NS1 GAC–ELISA) and indirect NS1 IgG ELISA, and compared the results with an E-based virus-like particle (VLP) GAC–ELISA. Results: NS1-based methods had comparable accuracies as VLP GAC–ELISA. Although the sensitivity in detecting anti-NS1 IgM was poor, indirect NS1 IgG ELISA showed similar limits of detection (~1–2 ng/mL) as NS1 GAC–ELISA in detecting anti-NS1 IgG. Combining the results from two or more tests as a composite reference standard can determine the DENV serostatus with a specificity reaching 100%. Conclusion: NS1-based ELISAs have comparable accuracies as VLP GAC–ELISA in determining dengue serostatus, which could effectively assist clinicians during assessments of vaccine eligibility.

Toxoplasmosis in Pregnant Women and Non-pregnant Female Volunteers in Port Harcourt, Rivers State, Nigeria

Background: Toxoplasmosis is caused by Toxoplasma gondii, a protozoan parasite, and is among the diseases generally referred to as neglected tropical disease (NTD). It can be a reason for impulsive abortion in pregnant women or hereditary chorioretinitis in neonates. Objectives: The present study examined seroprevalence and related risk issues of toxoplasmosis in pregnant women (PTW) and non-pregnant females (NPF) of two health facilities in Port Harcourt. Methods: Venous blood was collected from 380 women and analyzed by immunodiagnostic techniques using ELISA (Enzyme-Linked Immunosorbent assay) IgG and IgM tests and real-time polymerase chain reaction (PCR). Data were collected using a well-structured questionnaire on the socio-demographic risk issues linked with toxoplasmosis. Results: Among the study population, PTW and NPF had seroprevalence of 33.0% and 19.4%, respectively (P &lt; 0.05). PTW recorded a higher overall seroprevalence than NPF with 16.1%, 1.3%, and 8.4%, 0.8% for Toxoplasma gondii ELISA IgG, and IgM tests, respectively. IgM-positive cases were all authenticated by the real-time PCR technique and were all negative. Age group 35-39 years had the highest IgG seroprevalence of 24.3% for PTW, while the age group &gt; 40 years had the highest IgG seroprevalence of 18.2% for NPF. Traders recorded the uppermost seroprevalence of 20.9% and 2.6% for IgG and IgM among PTW, respectively, while teachers and traders recorded the maximum seroprevalence of 15.6% and 1.7% for IgG and IgM among NPF, respectively. Statistical analysis showed that there was no association between the age group, occupation, trimester, and toxoplasmosis. Conclusions: The toxoplasmosis was significantly high in pregnant women. It is recommended that Toxoplasma gondii tests should be included as a routine test during antenatal.

Impact of recent climate extremes on mosquito-borne disease transmission in Kenya.

Climate change and variability influence temperature and rainfall, which impact vector abundance and the dynamics of vector-borne disease transmission. Climate change is projected to increase the frequency and intensity of extreme climate events. Mosquito-borne diseases, such as dengue fever, are primarily transmitted by Aedes aegypti mosquitoes. Freshwater availability and temperature affect dengue vector populations via a variety of biological processes and thus influence the ability of mosquitoes to effectively transmit disease. However, the effect of droughts, floods, heat waves, and cold waves is not well understood. Using vector, climate, and dengue disease data collected between 2013 and 2019 in Kenya, this retrospective cohort study aims to elucidate the impact of extreme rainfall and temperature on mosquito abundance and the risk of arboviral infections. To define extreme periods of rainfall and land surface temperature (LST), we calculated monthly anomalies as deviations from long-term means (1983-2019 for rainfall, 2000-2019 for LST) across four study locations in Kenya. We classified extreme climate events as the upper and lower 10% of these calculated LST or rainfall deviations. Monthly Ae. aegypti abundance was recorded in Kenya using four trapping methods. Blood samples were also collected from children with febrile illness presenting to four field sites and tested for dengue virus using an IgG enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR). We found that mosquito eggs and adults were significantly more abundant one month following an abnormally wet month. The relationship between mosquito abundance and dengue risk follows a non-linear association. Our findings suggest that early warnings and targeted interventions during periods of abnormal rainfall and temperature, especially flooding, can potentially contribute to reductions in risk of viral transmission.