Reverse transcriptase polymerase chain reaction (RT-PCR) assays for coronavirus disease 2019 (COVID-19) should be interpreted with clinical, epidemiological history and exposure risk to avoid misdiagnosis. We report a cruise-ship worker with significant travelling history, presented with acute respiratory symptoms and radiographic evidence of viral pneumonia. Initial RNA-dependent RNA polymerase (RdRp) gene confirmatory assay was negative. Use of a more robust RT-PCR assay detected ORF1ab, N and S genes for COVID-19, and the diagnosis was supported by an IgM and IgG positive COVID-19 serology. Subsequent follow up samples which reported inconsistencies in detecting RdRp gene also raise the concern of reliability of RdRp gene as the confirmatory assay for diagnosis of COVID-19. Patient later had prolonged viral shedding beyond serological recovery, with a negative viral culture reflecting non-infectivity.
Dengue haemorrhagic fever (DHF) is a severe viral illness with significant morbidity and mortality especially among children in Southeast Asia. The tourniquet test is recommended by the World Health Organisation (WHO) as an initial clinical screening procedure for patients suspected to have DHF, particularly in patients with DHF grade I. The aim of this study was to evaluate the validity of the tourniquet test as a predictor of DHF and also to assess the usefulness of repeated, serial tourniquet testing as a diagnostic indicator of DHF. Individuals included into this study were children from the Institute of Paediatrics, Kuala Lumpur who were admitted on a clinical suspicion of DHF based on the WHO criteria and who had serology for Dengue IgM performed. A standard method of tourniquet was performed on these patients on a daily basis following admission. A total number of 60 patients were considered for analysis and of these the tourniquet test was positive in 50 patients and negative in the remaining 10 patients. Sensitivity of the test was found to be 85.4% and the specificity was 25%. Further assessment of the predictive value of the test showed that the positive predictive value (PPV) was 82% while the negative predictive value (NPV) was 30%. In conclusion, a positive tourniquet test, serially performed on a daily basis was found clinically to be a useful preliminary screening tool for DHF as recommended by WHO. However its specificity was low and consequently led to a high false positive rate.
High levels of anti-dengue IgM or IgG can be detected using numerous rapid diagnostic tests (RDTs). However, the sensitivity and specificity of these tests are reduced by changes in envelope glycoprotein antigenicity that inevitably occur in limited expression systems. A novel RDT was designed to enhance diagnostic sensitivity. Dengue viruses cultured in animal cells were used as antigens to retain the native viral coat protein. Monoclonal antibodies (mAbs) were then developed, for the first time, against domain I of envelope glycoprotein (EDI). The anti-dengue EDI mAb was employed as a capturer, and EDII and EDIII, which are mainly involved in the induction of neutralizing antibodies in patients, were fully available to bind to anti-dengue IgM or IgG in patients. A one-way automatic blood separation device prevented reverse migration of plasma and maximize the capture of anti-dengue antibodies at the test lines. A clinical evaluation in the field proved that the novel RDT (sensitivities of 96.5% and 96.7% for anti-dengue IgM and IgG) is more effective in detecting anti-dengue antibodies than two major commercial tests (sensitivities of 54.8% and 82% for SD BIOLINE; 50.4% and 75.3% for PanBio). The innovative format of RDT can be applied to other infectious viral diseases.
BACKGROUND: Dengue is a mosquito-borne disease that causes a public health problem in tropical and subtropical countries. Current immunological diagnostics based on IgM and/or nonstructural protein 1 (NS1) antigen are limited for acute dengue infection due to low sensitivity and accuracy.
OBJECTIVES: This study aimed to develop a one-step multiplex real-time RT-PCR assay showing higher sensitivity and accuracy than previous approaches.
STUDY DESIGN: Serotype-specific primers and probes were designed through the multiple alignment of NS1 gene. The linearity and limit of detection (LOD) of the assay were determined. The assay was clinically validated with an evaluation panel that was immunologically tested by WHO and Malaysian specimens.
RESULTS: The LOD of the assay was 3.0 log10 RNA copies for DENV-1, 2.0 for DENV-3, and 1.0 for DENV-2 and DENV-4. The assay showed 95.2% sensitivity (20/21) in an evaluation panel, whereas NS1 antigen- and anti-dengue IgM-based immunological assays exhibited 0% and 23.8-47.6% sensitivities, respectively. The assay showed 100% sensitivity both in NS1 antigen- and anti-dengue IgM-positive Malaysian specimens (26/26). The assay provided the information of viral loads and serotype with discrimination of heterotypic mixed infection.
CONCLUSIONS: The assay could be clinically applied to early dengue diagnosis, especially during the first 5 days of illness and approximately 14 days after infection showing an anti-dengue IgM-positive response.
Rapid diagnostic tests (RDTs) can detect anti-malaria antibodies in human blood. As they can detect parasite infection at the low parasite density, they are useful in endemic areas where light infection and/or re-infection of parasites are common. Thus, malaria antibody tests can be used for screening bloods in blood banks to prevent transfusion-transmitted malaria (TTM), an emerging problem in malaria endemic areas. However, only a few malaria antibody tests are available in the microwell-based assay format and these are not suitable for field application. A novel malaria antibody (Ab)-based RDT using a differential diagnostic marker for falciparum and vivax malaria was developed as a suitable high-throughput assay that is sensitive and practical for blood screening. The marker, merozoite surface protein 1 (MSP1) was discovered by generation of a Plasmodium-specific network and the hierarchical organization of modularity in the network. Clinical evaluation revealed that the novel Malaria Pf/Pv Ab RDT shows improved sensitivity (98%) and specificity (99.7%) compared with the performance of a commercial kit, SD BioLine Malaria P.f/P.v (95.1% sensitivity and 99.1% specificity). The novel Malaria Pf/Pv Ab RDT has potential for use as a cost-effective blood-screening tool for malaria and in turn, reduces TTM risk in endemic areas.