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.

Bovine enteroviruses (BEV) are members of the genus Enterovirus in the family Picornaviridae. They are predominantly isolated from cattle feces, but also are detected in feces of other animals, including goats and deer. These viruses are found in apparently healthy animals, as well as in animals with clinical signs and several studies reported recently suggest a potential role of BEV in causing disease in animals. In this study, we surveyed the presence of BEV in domestic and wild animals in Thailand, and assessed their genetic variability. Viral RNA was extracted from fecal samples of cattle, domestic goats, Indian bison (gaurs), and deer. The 5’ untranslated region (5’UTR) was amplified by nested reverse transcription-polymerase chain reaction (RT-PCR) with primers specific to BEV 5’UTR. PCR products were sequenced and analyzed phylogenetically using the neighbor-joining algorithm to observe genetic variations in regions of the bovine and bovine-like enteroviral 5’UTR found in this study. BEV and BEV-like sequences were detected in the fecal samples of cattle (40/60, 67 %), gaurs (3/30, 10 %), and goats (11/46, 24 %). Phylogenetic analyses of the partial 5’UTR sequences indicated that different BEV variants (both EV-E and EV-F species) co-circulated in the domestic cattle, whereas the sequences from gaurs and goats clustered according to the animal species, suggesting that these viruses are host species-specific. Varieties of BEV and BEV-like 5’UTR sequences were detected in fecal samples from both domestic and wild animals. To our knowledge, this is the first report of the genetic variability of BEV in Thailand.

The Malayan pit viper (Calloselasma rhodostoma) is a hemotoxic snake widely found in Southeast Asia and is responsible for the majority of poisoning cases in this region, including Thailand. However, a comprehensive knowledge of the venom protein profile and classification, as well as novel venom proteins, of this viper is still limited. Recently, the detailed composition of several snake venoms has been discovered through the use of transcriptome analysis. Therefore, the aim of this study was to employ a next-generation sequencing platform and bioinformatics analysis to undertake venom-gland de novo transcriptomics of Malayan pit vipers. Furthermore, 21,272 functional coding genes were identified from 36,577 transcripts, of which 314 transcripts were identified as toxin proteins, accounting for 61.41% of total FPKM, which can be categorized into 22 toxin gene families. The most abundant are snake venom metalloproteinase kistomin (P0CB14) and zinc metalloproteinase/disintegrin (P30403), which account for 60.47% of total toxin FPKM and belong to the SVMP toxin family, followed by snake venom serine protease 1 (O13059) and Snaclec rhodocetin subunit beta (P81398), which account for 6.84% and 5.50% of total toxin FPKM and belong to the snake venom serine protease (SVSP) and Snaclec toxin family, respectively. Amino acid sequences of the aforementioned toxins were compared with those identified in other important medical hemotoxic snakes from Southeast Asia, including the Siamese Russell's viper (Daboia siamensis) and green pit viper (Trimeresurus albolabris), in order to analyze their protein homology. The results demonstrated that ranges of 58%–62%, 31%–60%, and 48%–59% identity was observed among the SVMP, Snaclec, and SVSP toxin families, respectively. Understanding the venom protein profile and classification is essential in interpreting clinical symptoms during human envenomation and developing potential therapeutic applications. Moreover, the variability of toxin families and amino acid sequences among related hemotoxic snakes found in this study suggests the use and development of universal antivenom for the treatment of envenomating patients is still challenging.

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.

• Anti-rabies IgY was extracted from the egg yolks of immunized hens. • The isolation of IgY by PEG precipitation showed the highest amount and purity. • IgY affinity chromatography can increase the purity of IgY up to 98 %. • Both crude extracted and purified IgY could neutralize rabies virus according to RFFIT. • ∼8 eggs are required to produce 1 vial of RIG with the same potency as equine RIG.