Point-of-care testing for <i>Toxoplasma gondii</i> IgG/IgM using <i>Toxoplasma</i> ICT IgG-IgM test with sera from the United States and implications for developing countries

Background: Congenital toxoplasmosis is a serious but preventable and treatable disease. Gestational screening facilitates early detection and treatment of primary acquisition. Thus, fetal infection can be promptly diagnosed and treated and outcomes can be improved. Methods: We tested 180 sera with the Toxoplasma ICT IgG-IgM point-of-care (POC) test. Sera were from 116 chronically infected persons (48 serotype II; 14 serotype I-III; 25 serotype I-IIIa; 28 serotype Atypical, haplogroup 12; 1 not typed). These represent strains of parasites infecting mothers of congenitally infected children in the U.S. 51 seronegative samples and 13 samples from recently infected persons known to be IgG/IgM positive within the prior 2.7 months also were tested. Interpretation was confirmed by two blinded observers. A comparison of costs for POC vs. commercial laboratory testing methods was performed. Results: We found that this new Toxoplasma ICT IgG-IgM POC test was highly sensitive (100%) and specific (100%) for distinguishing IgG/IgM-positive from negative sera. Use of such reliable POC tests can be cost-saving and benefit patients. Conclusions: Our work demonstrates that the Toxoplasma ICT IgG-IgM test can function reliably as a point-of-care test to diagnose Toxoplasma gondii infection in the U.S. This provides an opportunity to improve maternal-fetal care by using approaches, diagnostic tools, and medicines already available. This infection has serious, lifelong consequences for infected persons and their families. From the present study, it appears a simple, low-cost POC test is now available to help prevent morbidity/disability, decrease cost, and make gestational screening feasible. It also offers new options for improved prenatal care in low- and middle-income countries.</p

Point-of-Care Testing for Toxoplasma Gondii IgG/IgM Using Toxoplasma ICT IgG-IgM Test with Sera from the United States and Implications for Developing Countries

Background Congenital toxoplasmosis is a serious but preventable and treatable disease. Gestational screening facilitates early detection and treatment of primary acquisition. Thus, fetal infection can be promptly diagnosed and treated and outcomes can be improved. Methods We tested 180 sera with the Toxoplasma ICT IgG-IgM point-of-care (POC) test. Sera were from 116 chronically infected persons (48 serotype II; 14 serotype I-III; 25 serotype I-IIIa; 28 serotype Atypical, haplogroup 12; 1 not typed). These represent strains of parasites infecting mothers of congenitally infected children in the U.S. 51 seronegative samples and 13 samples from recently infected persons known to be IgG/IgM positive within the prior 2.7 months also were tested. Interpretation was confirmed by two blinded observers. A comparison of costs for POC vs. commercial laboratory testing methods was performed. Results We found that this new Toxoplasma ICT IgG-IgM POC test was highly sensitive (100%) and specific (100%) for distinguishing IgG/IgM-positive from negative sera. Use of such reliable POC tests can be cost-saving and benefit patients. Conclusions Our work demonstrates that the Toxoplasma ICT IgG-IgM test can function reliably as a point-of-care test to diagnose Toxoplasma gondii infection in the U.S. This provides an opportunity to improve maternal-fetal care by using approaches, diagnostic tools, and medicines already available. This infection has serious, lifelong consequences for infected persons and their families. From the present study, it appears a simple, low-cost POC test is now available to help prevent morbidity/disability, decrease cost, and make gestational screening feasible. It also offers new options for improved prenatal care in low- and middle-income countries

New paradigms for understanding and step changes in treating active and chronic, persistent apicomplexan infections

Toxoplasma gondii, the most common parasitic infection of human brain and eye, persists across lifetimes, can progressively damage sight, and is currently incurable. New, curative medicines are needed urgently. Herein, we develop novel models to facilitate drug development: EGS strain T. gondii forms cysts in vitro that induce oocysts in cats, the gold standard criterion for cysts. These cysts highly express cytochrome b. Using these models, we envisioned, and then created, novel 4-(1H)-quinolone scaffolds that target the cytochrome bc₁ complex Qi site, of which, a substituted 5,6,7,8-tetrahydroquinolin-4-one inhibits active infection (IC₅₀, 30 nM) and cysts (IC₅₀, 4 μM) in vitro, and in vivo (25 mg/kg), and drug resistant Plasmodium falciparum (IC₅₀, <30 nM), with clinically relevant synergy. Mutant yeast and co-crystallographic studies demonstrate binding to the bc₁ complex Q[subscript]i site. Our results have direct impact on improving outcomes for those with toxoplasmosis, malaria, and ~2 billion persons chronically infected with encysted bradyzoites

Novel paradigm enables accurate monthly gestational screening to prevent congenital toxoplasmosis and more

Background: Congenital toxoplasmosis is a treatable, preventable disease, but untreated causes death, prematurity, loss of sight, cognition and motor function, and substantial costs worldwide. Objectives: We asked whether high performance of an Immunochromatographic-test (ICT) could enable accurate, rapid diagnosis/treatment, establishing new, improved care-paradigms at point-of-care and clinical laboratory. Methods: Data were obtained in 12 studies/analyses addressing: 1-feasibility/efficacy; 2-false-positives; 3-acceptability; 4-pink/black-line/all studies; 5-time/cost; 6-Quick-Information/Limit-of-detection; 7, 8-acute;-chronic; 9-epidemiology; 10-ADBio; 11,12-Commentary/Cases/Chronology. Findings: ICT was compared with gold-standard or predicate-tests. Overall, ICT performance for 1093 blood/4967 sera was 99.2%/97.5% sensitive and 99.0%/99.7% specific. However, in clinical trial, FDA-cleared-predicate tests initially caused practical, costly problems due to false-positive-IgM results. For 58 persons, 3/43 seronegative and 2/15 chronically infected persons had false positive IgM predicate tests. This caused substantial anxiety, concerns, and required costly, delayed confirmation in reference centers. Absence of false positive ICT results contributes to solutions: Lyon and Paris France and USA Reference laboratories frequently receive sera with erroneously positive local laboratory IgM results impeding patient care. Therefore, thirty-two such sera referred to Lyon’s Reference laboratory were ICT-tested. We collated these with other earlier/ongoing results: 132 of 137 USA or French persons had false-positive local laboratory IgM results identified correctly as negative by ICT. Five false positive ICT results in Tunisia and Marseille, France, emphasize need to confirm positive ICT results with Sabin-Feldman-Dye-test or western blot. Separate studies demonstrated high performance in detecting acute infections, meeting FDA, CLIA, WHO REASSURED, CEMark criteria and patient and physician satisfaction with monthly-gestational-ICT-screening. Conclusions/significance: This novel paradigm using ICT identifies likely false positives or raises suspicion that a result is truly positive, rapidly needing prompt follow up and treatment. Thus, ICT enables well-accepted gestational screening programs that facilitate rapid treatment saving lives, sight, cognition and motor function. This reduces anxiety, delays, work, and cost at point-of-care and clinical laboratories. Trial registration: NCT04474132,&nbsp;https://clinicaltrials.gov/study/NCT04474132ClinicalTrials.gov</p

Rapid, inexpensive, fingerstick, whole-blood, sensitive, specific, point-of-care test for anti-<i>Toxoplasma</i> antibodies

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Rapid, inexpensive, fingerstick, whole-blood, sensitive, specific, point-of-care test for anti-Toxoplasma antibodies.

Rapid, inexpensive, fingerstick, whole-blood, sensitive, specific, point-of-care test for anti-<i>Toxoplasma</i> antibodie

Implementation of <i>Toxoplasma</i> ICT IgG-IgM POC testing with separation of serum at point of care and representative <i>Toxoplasma</i> ICT IgG-IgM test negative and positive test results for sera.

<p>This involves a lancet to obtain the sample with fingerprick ($0.13 for the lancet and a very low cost for alcohol wipes), a small centrifuge tube to separate the serum, a small Class II biosafety cabinet ($6546) for safe handling of samples, and a small centrifuge for separating serum (228), from which serum can be removed easily and be tested. The following methodology is described in Chapey [17]: Briefly: the Toxoplasma ICT IgG-IgM assay is based on a lateral flow chromatographic immunoassay (LFCI) technology that allows the simultaneous detection of T. gondii IgG or IgM antibodies in human serum/plasma [17]. A minimum sample volume of 30–50 μL of serum/plasma is required [17]. Each cassette contains: a) a nitrocellulose strip on which there are two reactive bands, one with the Toxoplasma gondii antigen (from whole cell lysate) called the “test” band (T band) and one with the rabbit gamma globulins called the “control” band (C band); b) a fiberglass support (conjugate pad) which is impregnated with red latex particles coupled with Toxoplasma antigens (“test” latex = T latex) and blue latex particles coupled with goat anti-rabbit IgG (“control” latex = C latex) [17]. The test is run by dispensing the serum/plasma and an eluting solution (eluent) in the “sample well” of the cassette [17]. With the addition of the eluent, starts the concomitant migration (chromatography) of the serum/plasma and the latex particles [17]. If anti-Toxoplasma antibodies (IgG or IgM or both) are present in the sample, a complex is formed between the T latex and the patient’s antibodies, which is then captured by the T band, and it results in the appearance of a red line (positive test) [17]. The direct capture of the C latex by the C band results in the appearance of a control blue line which indicates that the chromatography performed well [17]. The results are read 20–30 minutes after the eluent solution has been dispensed into the well [17]. Representative example of U.S. sera, negative (left) and positive (right) results. This is the new simple POC test, based on lateral-flow-chromatographic-immunoassay method, already commercially available in France, that detects simultaneously both Toxoplasma IgG and IgM antibodies and costs only 4 (the cost we were charged) per test, as opposed to a $650 cost for testing at a commercial laboratory in the U.S.</p

Summary of results with <i>Toxoplasma</i> ICT IgG-IgM test and reference tests: Test parameters of <i>Toxoplasma</i> ICT IgG-IgM POC test.

<p>Summary of results with <i>Toxoplasma</i> ICT IgG-IgM test and reference tests: Test parameters of <i>Toxoplasma</i> ICT IgG-IgM POC test.</p

Summary of results with <i>Toxoplasma</i> ICT IgG-IgM test and reference tests: Subgroup breakdown.

<p>Summary of results with <i>Toxoplasma</i> ICT IgG-IgM test and reference tests: Subgroup breakdown.</p