Development and Diagnosis Performance of IgM-Based Rapid Antigen Test for Early Detection of SARS-CoV-2 Infection in a Large Cohort of Suspected COVID-19 Cases—USA,Poland,and Sweden,2021-2022

Introduction:Antigen testing has been crucial in effectively managing the coronavirus disease 2019(COVID-19)pandemic.This study evaluated the clinical performance of a nasopharyngeal swab(NPS)-based antigen rapid diagnostic test(Ag-RDT)compared to the gold standard real-time reverse transcription-polymerase chain reaction(RT-PCR)for early detection of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).Methods:We developed an IgM-based rapid antigen test for early detection of SARS-CoV-2 infection.Between July 2021 and January 2022,we analyzed 1,030 NPS samples from participants at three centers in different countries,using both antigen rapid diagnostic tests(Ag-RDT)and RT-PCR.Results:The Ag-RDT demonstrated minimal detection limits as low as 0.1 ng/mL for recombinant N antigen and 100 TCID50/mL for heat-inactivated SARS-CoV-2 virus.Specificity assessments involving four human coronaviruses and 13 other respiratory viruses showed no cross-reactivity.The Ag-RDT assay(ALLtest)exhibited high sensitivity(93.18%-100%)and specificity(99.67%-100%)across all centers.Factors such as cycle threshold(Ct)values and the timing of symptoms since onset were influential,with sensitivity increasing at lower Ct values(<30)and within the first week of symptoms.Conclusion:The ALLtest Ag-RDT demonstrated high reliability and significant potential for diagnosing suspected COVID-19 cases.

Toxoplasma gondii infection induces cell apoptosis via multiple pathways revealed by transcriptome analysis

Toxoplasma gondii is a worldwide parasite that can infect almost all kinds of mammals and cause fatal toxoplasmosis in immunocompromised patients.Apoptosis is one of the principal strategies of host cells to clear pathogens and maintain organismal homeostasis,but the mechanism of cell apoptosis induced by T.gondii remains obscure.To explore the apoptosis influenced by T.gondii,Vero cells infected or uninfected with the parasite were subjected to apoptosis detection and subsequent dual RNA sequencing(RNA-seq).Using high-throughput Illumina sequencing and bioinformatics analysis,we found that pro-apoptosis genes such as DNA damage-inducible transcript 3(DDIT3),growth arrest and DNA damage-inducibleα(GADD45 A),caspase-3(CASP3),and high-temperature requirement protease A2(Htr A2)were upregulated,and anti-apoptosis genes such as poly(adenosine diphosphate(ADP)-ribose)polymerase family member 3(PARP3),B-cell lymphoma 2(Bcl-2),and baculoviral inhibitor of apoptosis protein(IAP)repeat containing 5(BIRC5)were downregulated.Besides,tumor necrosis factor(TNF)receptor-associated factor 1(TRAF1),TRAF2,TNF receptor superfamily member 10 b(TNFRSF10 b),disabled homolog2(DAB2)-interacting protein(DAB2 IP),and inositol 1,4,5-trisphosphate receptor type 3(ITPR3)were enriched in the upstream of TNF,TNF-related apoptosis-inducing ligand(TRAIL),and endoplasmic reticulum(ER)stress pathways,and TRAIL-receptor2(TRAIL-R2)was regarded as an important membrane receptor influenced by T.gondii that had not been previously considered.In conclusion,the T.gondii RH strain could promote and mediate apoptosis through multiple pathways mentioned above in Vero cells.Our findings improve the understanding of the T.gondii infection process through providing new insights into the related cellular apoptosis mechanisms.

A live attenuated RHΔompdcΔuprt mutant of Toxoplasma gondii induces strong protective immunity against toxoplasmosis in mice and cats

Background Toxoplasma gondii is an obligate intracellular apicomplexan parasite and is responsible for zoonotic toxoplasmosis.It is essential to develop an effective anti-T.gondii vaccine for the control of toxoplasmosis,and this study is to explore the immunoprotective effects of a live attenuated vaccine in mice and cats.Methods First,theompdc anduprt genes of T.gondii were deleted through the CRISPR-Cas9 system.Then,the intracellular proliferation and virulence of this mutant strain were evaluated.Subsequently,the immune responses induced by this mutant in mice and cats were detected,including antibody titers,cytokine levels,and subsets of T lymphocytes.Finally,the immunoprotective effects were evaluated by challenge with tachyzoites of different strains in mice or cysts of the ME49 strain in cats.Furthermore,to discover the effective immune element against toxoplasmosis,passive immunizations were carried out.GraphPad Prism software was used to conduct the log-rank(Mantel–Cox)test,Student’st test and one-way ANOVA.Results The RHΔompdcΔuprt were constructed by the CRISPR-Cas9 system.Compared with the wild-type strain,the mutant notably reduced proliferation(P<0.05).In addition,the mutant exhibited virulence attenuation in both murine(BALB/c and BALB/c-nu)and cat models.Notably,limited pathological changes were found in tissues from RHΔompdcΔuprt-injected mice.Furthermore,compared with nonimmunized group,high levels of IgG(IgG1 and IgG2a)antibodies and cytokines(IFN-γ,IL-4,IL-10,IL-2 and IL-12)in mice were detected by the mutant(P<0.05).Remarkably,all RHΔompdcΔuprt-vaccinated mice survived a lethal challenge with RHΔku80 and ME49 and WH6 strains.The immunized sera and splenocytes,especially CD8^(+)T cells,could significantly extend(P<0.05)the survival time of mice challenged with the RHΔku80 strain compared with naïve mice.In addition,compared with nonimmunized cats,cats immunized with the mutant produced high levels of antibodies and cytokines(P<0.05),and notably decreased the shedding numbers of oocysts in feces(95.3%).Conclusions The avirulent RHΔompdcΔuprt strain can provide strong anti-T.gondii immune responses,and is a promising candidate for developing a safe and effective live attenuated vaccine.