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The effect of RNA silencing in plants can be amplified if the production of secondary small interfering RNAs (siRNAs) is triggered by the interaction of microRNAs (miRNAs) or siRNAs with a long target RNA. miRNA and siRNA interactions are not all equivalent, however; most of them do not trigger secondary siRNA production. Here we use bioinformatics to show that the secondary siRNA triggers are miRNAs and transacting siRNAs of 22 nt, rather than the more typical 21-nt length. Agrobacterium-mediated transient expression in Nicotiana benthamiana confirms that the siRNA-initiating miRNAs, miR173 and miR828, are effective as triggers only if expressed in a 22-nt form and, conversely, that increasing the length of miR319 from 21 to 22 nt converts it to an siRNA trigger. We also predicted and validated that the 22-nt miR771 is a secondary siRNA trigger. Our data demonstrate that the function of small RNAs is influenced by size, and that a length of 22 nt facilitates the triggering of secondary siRNA production.

Early diagnosis is important for the clinical management of diseases caused by dengue virus (DENV) infections. We investigated the performance of three commercially available DENV nonstructural protein 1 (NS1) rapid diagnostic tests (RDTs) using 173 acute-phase sera collected from dengue fever-suspected patients during the 2012-2013 DENV outbreak in Taiwan. The results of the NS1 RDTs were compared with those of qRT-PCR to calculate the sensitivity and specificity of the NS1 RDTs. The anti-DENV IgM and IgG RDT results were included to increase the probability of detecting acute DENV infection. The anti-DENV IgM/IgG RDT results were also compared with those of IgM/IgG captured ELISA. The sera from DENV qRT-PCR-positive patients were subjected to NS1 RDTs, as well as IgM/IgG captured ELISA. These results suggested that there was no significant difference in the sensitivities of the three commercially available DNEV NS1 RDTs; the SD NS1 RDT results showed the highest agreement with the qRT-PCR reference results, followed in order by the Bio-Rad and CTK NS1 RDT results when the specificity was considered. Inclusion of the IgM or IgG RDT results increased the likelihood of diagnosing either a primary or secondary DENV infection. NS1 RDTs were more sensitive for the detection of primary infections than secondary infections, related to DENV viremia levels determined by qRT-PCR. These results suggested that anti-DENV antibodies reduced the sensitivity of NS1 rapid tests. We also analyzed the sensitivity for the detection of different DENV serotypes, and the results suggested that the NS1 RDTs used in this study were valuable for rapid screening of acute DENV infection with DENV-1, DENV-2 and DENV-3. Our results suggest that the NS1 RDT is a good alternative to qRT-PCR analysis for timely dengue disease management and prevention in dengue-endemic regions where medical resources are lacking or during large dengue outbreaks. However, the relatively low sensitivity for DENV-4 might miss the detection of DENV-4-infected cases.

Background Dengue virus serotype 2 (DENV-2) was the major serotype in the 2015 dengue outbreak in Taiwan, while DENV-1 and DENV-3 were dominant between 2005 and 2014. We aimed to investigate whether DENV-2 contributed to disease severity and mortality in the outbreak in Kaohsiung city, Taiwan. Methods We collected serum samples from dengue patients to detect the presence of DENV and determine the serotypes by using quantitative reverse transcription-polymerase chain reaction. Our cohorts comprised 105 DENV-1-infected cases and 1,550 DENV-2-infected cases. Demographic data, DENV serotype, and comorbidities were covariates for univariate and multivariate analyses to explore the association with severity and mortality. Results The results suggested that DENV-1 persisted and circulated, while DENV-2 was dominant during the dengue outbreak that occurred between September and December 2015. However, DENV-2 did not directly contribute to either severity or mortality. Aged patients and patients with diabetes mellitus (DM) or moderate to severe chronic kidney disease (CKD) had a higher risk of developing severe dengue. The mortality of dengue patients was related to a higher Charlson comorbidity index score and severe dengue. Among DENV-2-infected patients and older patients, preexisting anti-dengue IgG, DM, and moderate to severe CKD were associated with severe dengue. Moreover, female sex and severe dengue were associated with a significantly higher risk of death. Conclusions Our findings highlight the importance of timely serological testing in elderly patients to identify potential secondary infections and focus on the meticulous management of elderly patients with DM or moderate to severe CKD to reduce dengue-related death.

In Taiwan, coronavirus disease 2019 (COVID‐19) involving the delta variant occurred after that involving the alpha variant in 2021. In this study, we aimed to analyze the Delta variant. A total of 318 patients in Taiwan infected with delta variants were identified. The case fatality rate (CFR) of patients infected with delta variants was 0.94% in Taiwan compared with that of those infected with alpha variants (5.95%). The possible reasons for the low CFR might be hybrid immunity due to infection and rapid promotion of the COVID‐19 vaccination program during the alpha variant outbreak. We identified three 21J delta variants. Two long gene deletions were detected in these severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2) isolates: ORF7aΔ91 in KMUH‐8 and SpikeΔ30 in KMUH‐9. Protein structure prediction indicates that ORF7aΔ91 results in malfunction of NS7a as an interferon antagonist and that SpikeΔ30 results in a truncated spike protein (N679–A688del), resulting in a lower infection rate compared with the delta variant without these deletions. The impact of these two deletions on SARS‐CoV‐2‐associated pathogenesis deserves further investigation. Delta variants still exist in many regions in the omicron era, and the backbone of the delta variant genome possibly spread worldwide in the form of delta‐omicron hybrids (deltacron; e.g., XBC.1 and XAY.2), which casts a potential threat to public health. Our study further highlighted the importance of more understanding of the delta variants.

Background Reactive oxygen species are important mediators exerting toxic effects on various organs during ischemia-reperfusion (IR) injury. We hypothesized that adipose-derived mesenchymal stem cells (ADMSCs) protect the kidney against oxidative stress and inflammatory stimuli in rat during renal IR injury. Methods Adult male Sprague-Dawley (SD) rats (n = 24) were equally randomized into group 1 (sham control), group 2 (IR plus culture medium only), and group 3 (IR plus immediate intra-renal administration of 1.0 × 10 6 autologous ADMSCs, followed by intravenous ADMSCs at 6 h and 24 h after IR). The duration of ischemia was 1 h, followed by 72 hours of reperfusion before the animals were sacrificed. Results Serum creatinine and blood urea nitrogen levels and the degree of histological abnormalities were markedly lower in group 3 than in group 2 (all p < 0.03). The mRNA expressions of inflammatory, oxidative stress, and apoptotic biomarkers were lower, whereas the anti-inflammatory, anti-oxidative, and anti-apoptotic biomarkers were higher in group 3 than in group 2 (all p < 0.03). Immunofluorescent staining showed a higher number of CD31+, von Willebrand Factor+, and heme oxygenase (HO)-1+ cells in group 3 than in group 2 (all p < 0.05). Western blot showed notably higher NAD(P)H quinone oxidoreductase 1 and HO-1 activities, two indicators of anti-oxidative capacity, in group 3 than those in group 2 (all p < 0.04). Immunohistochemical staining showed higher glutathione peroxidase and glutathione reductase activities in group 3 than in group 2 (all p < 0.02) Conclusion ADMSC therapy minimized kidney damage after IR injury through suppressing oxidative stress and inflammatory response.

After the previous major dengue fever (DF) outbreaks in 2014 and 2015 in Taiwan, the second-largest DF outbreak re-emerged in 2023. A total of 178 patients with laboratory-confirmed dengue virus (DENV) infection, including 92 DENV-1 and 86 DENV-2 cases, were enrolled in this study conducted during the 2023 dengue outbreak in Kaohsiung City, Taiwan. This study aimed to analyze epidemiological characteristics, clinical severity, and risk factors for severe dengue (SD), as well as the diagnostic implications of the non-structural protein 1 (NS1) antigen rapid test. Patients infected with DENV-2 exhibited significantly older age, higher incidence of secondary infections, diabetes mellitus (DM), hypertension (HT), and longer hospital stays than patients infected with DENV-1. Multivariate analysis revealed that older age (age ≥65), secondary dengue infection, DM, and HT were significant independent predictors of SD. Compared with non-SD cases, SD patients were significantly more likely to be older (age ≥65), to exhibit a higher incidence of secondary infections and a greater prevalence of chronic diseases, including DM and HT. Notably, dengue-confirmed patients with negative NS1 results had a shorter duration since symptom onset (p < 0.001). Our DENV-1 and DENV-2 isolates are related to strains from neighboring Asian countries. Our findings emphasize the important factors of old age, secondary infections, and chronic diseases that contributed to dengue severity. We should meticulously manage these high-risk groups to prevent dengue progression. Screening incoming travelers for DF during the epidemic season will be an important measure to prevent the introduction of DENV into Taiwan.

In late July of 2016, a temperature mooring deployed north of Dongsha Atoll has recorded the enhancement of nonlinear internal waves (NIWs) associated with regionally environmental variability. Observations revealed large daily temperature fluctuations of 10–14 °C at depths of 80–100 m, with peak variations occurring after spring tides and showing an increasing trend, indicative of intensified NIW activity. This study investigates the uncertainties regarding the influence of river flux, precipitation, and other environmental factors on the short-term intensification of NIWs. Through the analysis of multiple independent oceanic and atmospheric datasets, we identified significant variations in zonal oceanic currents and the advection of low-salinity water discharged from the Pearl River to the Dongsha mooring (DSM), which enhanced in situ stratification. The analysis of buoyancy frequency squared (N 2 ) indicates favorable conditions for westward NIW propagation from DSM toward the Pearl River Estuary. These findings highlight the critical role of river flux in shaping NIW dynamics by creating a pathway that enhances the concentration and propagation of NIWs. Given the rising frequency of extreme summer precipitation events associated with global warming and climate change, this study suggests an increased likelihood of NIWs in the northern South China Sea directly reaching the southeastern mainland coast from the Luzon Strait.

Background Rapid molecular diagnostics are critical for timely dengue virus (DENV) detection, especially in resource-limited settings. Reverse transcription-insulated isothermal polymerase chain reaction (RT-iiPCR) enables sample-to-answer amplification without thermocyclers. While this platform was previously validated, its diagnostic performance against contemporary viral genotypes and, critically, across different host immune statuses (primary vs. secondary infection) remains largely undefined. We assessed a DENV iiPCR assay against quantitative real-time reverse transcription PCR (qRT-PCR) during the 2023 outbreak in southern Taiwan. Methods Acute-phase sera (n = 185) from febrile patients were tested by qRT-PCR, pan-DENV iiPCR, IgM/IgG ELISAs, and serotype-specific RT-PCR/iiPCR. Diagnostic performance was evaluated with qRT-PCR as a reference. Subgroup analyses examined effects of days post-symptom onset (PSO), immune status (primary vs secondary infection), and serotype on iiPCR sensitivity. Results Pan-iiPCR achieved 88.4 % sensitivity and 100 % specificity versus qRT-PCR. Sensitivity peaked within 3 days PSO (96.8 %) and in primary infections (95.0 %) but declined after day 3 (53.3 %) and in secondary infections (82.2 %). Combining pan-iiPCR with IgM testing raised sensitivity to ≥99 % beyond the viremic window. iiPCR correctly typed 83.5 % (71/85) of DENV-1 and 94.3 % (66/70) of DENV-2. Because pan-iiPCR served as the screening step, 18 RT-PCR–positive specimens were pan-iiPCR negative and therefore did not proceed to iiPCR serotyping. Conclusions iiPCR enables rapid early-phase detection and is useful when paired with serology. However, reduced sensitivity in low-viremia and secondary infections, as well as incomplete serotyping, limit standalone use. Primer optimization and integration with IgM testing may enhance outbreak utility.

The Omicron variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), first detected in October 2021, possessed many mutations compared to previous variants. We aimed to identify and analyze SARS-CoV-2 Omicron subvariants among coronavirus disease 2019 (COVID-19) patients between January 2022 and September 2022 in Taiwan. The results revealed that BA.2.3.7, featuring K97E and G1251V in the spike protein compared with BA.2, emerged in March 2022 and persistently dominated between April 2022 and August 2022, resulting in the largest COVID-19 outbreak since 2020. The accumulation of amino acid (AA) variations, mainly AA substitution, in the spike protein was accompanied by increasing severity in Omicron-related COVID-19 between April 2022 and January 2023. Older patients were more likely to have severe COVID-19, and comorbidity was a risk factor for COVID-19-related mortality. The accumulated case fatality rate (CFR) dropped drastically after Omicron variants, mainly BA.2.3.7, entered Taiwan after April 2022, and the CFR was 0.16% in Taiwan, which was lower than that worldwide (0.31%) between April 2021 and January 2023. The relatively low CFR in Omicron-related COVID-19 patients can be attributed to adjustments to public health policies, promotion of vaccination programs, effective antiviral drugs, and the lower severity of the Omicron variant.

Dengue fever, a mosquito-borne disease, is caused by dengue virus (DENV) which includes four major serotypes (DENV-1, -2, -3, and -4). Some serotypes cause more severe diseases than the other; severe dengue is associated with secondary infections by a different serotype. Timely serotyping can provide early warning of dengue epidemics to improve management of patients and outbreaks. A mobile insulated isothermal PCR (iiPCR) system is available to allow molecular detection of pathogens near points of need. In this study, side-by-side comparison with the CDC DENV-1-4 Real Time RT-PCR (qRT-PCR) was performed to evaluate the performance of four singleplex DENV-1-4 serotyping reverse transcription-iiPCR (RT-iiPCR) reagents for DENV subtyping on the mobile PCR system. The four RT-iiPCRs did not react with Zika virus and chikungunya virus; tests with serial dilutions of the four DENV serotypes made in human serum showed they had detection endpoints comparable to those of the reference method, indicating great analytical sensitivity and specificity. Clinical performance of the RT-iiPCR reagents was evaluated by testing 40 serum samples each (around 20 target serotype-positive and 20 DENV-negative); all four reagents had high agreement (97.5-100%) with the reference qRT-PCR. Moreover, testing of mosquitoes separately infected experimentally with each serotype showed that the four reagents detected specifically their target DENV serotypes in mosquito. With analytical and clinical performance comparable to the reference qRT-PCR assay, the four index RT-iiPCR reagents on the field-deployable PCR system can serve as a useful tool for DENV detection near points of needs.