Diagnostic Performance of Dengue NS1 and Antibodies by Serum Concentration Technique

Dengue infection has been a public health problem worldwide, especially in tropical areas. A lack of sensitive diagnostic methods in the early phase of the illness is one of the challenging problems in clinical practices. We, herein, analyzed 86 sera of acute febrile patients, from both dengue and non-dengue febrile illness, to study the diagnostic performance of dengue diagnostics. When compared with detection by Polymerase Chain Reaction (PCR), dengue NS1 detection by enzyme-linked immunosorbent assay (ELISA) had the highest sensitivity of 82.4% (with 94.3% specificity), while NS1 by rapid diagnostic test (RDT) had 76.5% sensitivity. IgM detection by ELISA and RDT showed only 27.5% and 17.9% sensitivity, respectively. The combination of NS1 and IgM in RDT yielded a sensitivity of 78.4%, with 97.1% specificity. One of the essential steps in making a diagnosis from patient samples is the preparation process. At present, a variety of techniques have been used to increase the number of analytes in clinical samples. In this study, we focused on the sample concentration method. The sera were concentrated three times with the ultrafiltration method using a 10 kDa molecular weight cut-off membrane. The results showed an increase in the sensitivity of RDT-NS1 detection at 80.4%, with 100% specificity. When combining NS1 and IgM detection, the concentration method granted RDT an 82.4% sensitivity, with 100% specificity. In conclusion, serum concentration by the ultrafiltration method is a simple and applicable technique. It could increase the diagnostic performance of point-of-care dengue diagnostics

Detection of SARS-CoV-2 and Variants in Hospital Wastewater in a Developing Country

Wastewater-based epidemiology (WBE) is a beneficial tool for comprehensive health information on communities, especially during the COVID-19 pandemic. In developing countries, including Thailand, the application of WBE is limited. Few SARS-CoV-2 detections and variants have been monitored in wastewater in these countries. This is because of the time-consuming, low recovery of viruses in the concentration techniques and difficulties in finding the proper primers and amplification kits. Therefore, this study aimed to quantify SARS-CoV-2 RNA concentration using a commercial clinical kit. We identified the SARS-CoV-2 variants and estimated the detection costs in the wastewater samples. One hundred and fifty hospital wastewater samples were filtered with commercial ultrafiltration (UF) and then detected for the SARS-CoV-2 concentration using a Sansure Biotech SARS-CoV-2 kit. The recovery of the virus concentration technique in UF was studied using a surrogate (porcine epidemic diarrhea virus). The virus detection in wastewater was quantified by RT-qPCR. In addition, the mutation sites in the partial spike glycoprotein (S) gene of SARS-CoV-2 were verified using short nested RT-PCR. The results showed a high recovery of the commercial UF (80.53%), and 24.6% of hospital wastewater contained SARS-CoV-2. The detection of SARS-CoV-2 in wastewater cost USD 35.43 per sample. The virus variants revealed V70del, H69del, and V144del mutations in the partial S gene of SARS-CoV-2 in B.1.1.7 (SARS-CoV-2 Alpha variant), and T95I and G142D mutations in B.1.617.2 (Delta variant)

Detection of SARS-CoV-2 and Variants in Hospital Wastewater in a Developing Country

Wastewater-based epidemiology (WBE) is a beneficial tool for comprehensive health information on communities, especially during the COVID-19 pandemic. In developing countries, including Thailand, the application of WBE is limited. Few SARS-CoV-2 detections and variants have been monitored in wastewater in these countries. This is because of the time-consuming, low recovery of viruses in the concentration techniques and difficulties in finding the proper primers and amplification kits. Therefore, this study aimed to quantify SARS-CoV-2 RNA concentration using a commercial clinical kit. We identified the SARS-CoV-2 variants and estimated the detection costs in the wastewater samples. One hundred and fifty hospital wastewater samples were filtered with commercial ultrafiltration (UF) and then detected for the SARS-CoV-2 concentration using a Sansure Biotech SARS-CoV-2 kit. The recovery of the virus concentration technique in UF was studied using a surrogate (porcine epidemic diarrhea virus). The virus detection in wastewater was quantified by RT-qPCR. In addition, the mutation sites in the partial spike glycoprotein (S) gene of SARS-CoV-2 were verified using short nested RT-PCR. The results showed a high recovery of the commercial UF (80.53%), and 24.6% of hospital wastewater contained SARS-CoV-2. The detection of SARS-CoV-2 in wastewater cost USD 35.43 per sample. The virus variants revealed V70del, H69del, and V144del mutations in the partial S gene of SARS-CoV-2 in B.1.1.7 (SARS-CoV-2 Alpha variant), and T95I and G142D mutations in B.1.617.2 (Delta variant)

False Positivity of Anti-SARS-CoV-2 Antibodies in Patients with Acute Tropical Diseases in Thailand

Serology remains a useful indirect method of diagnosing tropical diseases, especially in dengue infection. However, the current literature regarding cross-reactivity between SARS-CoV-2 and dengue serology is limited and revealed conflicting results. As a means to uncover relevant serological insight involving antibody classes against SARS-CoV-2 and cross-reactivity, anti-SARS-CoV-2 IgA, IgM, and IgG ELISA, based on spike and nucleocapsid proteins, were selected for a fever-presenting tropical disease patient investigation. The study was conducted at the Faculty of Tropical Medicine during March to December 2021. The study data source comprised (i) 170 non-COVID-19 sera from 140 adults and children presenting with acute undifferentiated febrile illness and 30 healthy volunteers, and (ii) 31 COVID-19 sera from 17 RT-PCR-confirmed COVID-19 patients. Among 170 non-COVID-19 samples, 27 were false positives (15.9%), of which IgA, IgM, and IgG cross-reactive antibody classes were detected in 18 (10.6%), 9 (5.3%), and 3 (1.8%) cases, respectively. Interestingly, one case exhibited both IgA and IgM false positivity, while two cases exhibited both IgA and IgG false positivity. The false positivity rate in anti-SARS-CoV-2 IgA and IgM was reported in adults with dengue infection (11.3% and 5%) and adults with other tropical diseases (16.7% and 13.3%). The urea dissociation method applied to mitigate false positivity resulted in significantly decreased ELISA-based false and true positives. In conclusion, the analysis of antibody against SARS-CoV-2 in sera of patients with different tropical diseases showed that high IgA and IgM false positivity thus potentially limits serological assay utility in fever-presenting patients in tropical areas

Mass spectrometry-based identification and whole-genome characterisation of the first pteropine orthoreovirus isolated from monkey faeces in Thailand

Abstract Background The pteropine orthoreovirus (PRV) was isolated from monkey (Macaca fascicularis) faecal samples collected from human-inhabited areas in Lopburi Province, Thailand. These samples were initially obtained to survey for the presence of hepatitis E virus (HEV). Results Two virus isolates were retrieved by virus culture of 55 monkey faecal samples. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was successfully used to identify the viruses as the segmented dsRNA orthoreovirus. Phylogenetic analysis of the Lopburi orthoreovirus whole-genomes revealed relationships with the well-characterised PRVs Pulau (segment L1), Cangyuan (segments L2, M3 and S3), Melaka (segments L3 and M2), Kampar (segments M1 and S2) and Sikamat (segments S1 and S4) of Southeast Asia and China with nucleotide sequence identities of 93.5–98.9%. RT-PCR showed that PRV was detected in 10.9% (6/55) and HEV was detected in 25.5% (14/55) of the monkey faecal samples. Conclusions PRV was isolated from monkey faeces for the first time in Thailand via viral culture and LC-MS/MS. The genetic diversity of the virus genome segments suggested a re-assortment within the PRV species group. The overall findings emphasise that monkey faeces can be sources of zoonotic viruses, including PRV and HEV, and suggest the need for active virus surveillance in areas of human and monkey co-habitation to prevent and control emerging zoonotic diseases in the future