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Few studies have investigated whether SARS-CoV-2 infections increase the incidence of dengue haemorrhagic fever/shock syndrome (DHF/DSS) and/or severe dengue (SD) in dengue virus (DENV)-infected patients. This study was performed on a site with high incidences of classical dengue, but relatively few DHF/DSS or SD cases as defined by the WHO 1997 or 2009 criteria, respectively. Clinical, haematological/biochemical, and viral diagnostic data were collected from febrile patients before, during, and after the COVID-19 epidemic to assess whether (a) DENV-infected patients with prior SARS-CoV-2 infections or (b) DENV-SARS-CoV-2-co-infected patients had increased incidences of SD/DHF/DSS using logistic regression and machine learning models. Higher numbers of DHF/DSS/SD occurred during the COVID-19 epidemic, particularly in males and 18–40-year-olds. Significantly increased symptoms in the DENV-SARS-CoV-2-co-infected cases were (a) haemoconcentration (p < 0.0009) and hypotension (p < 0.0005) (DHF/DSS and SD criteria), (b) thrombocytopenia and mucosal bleeding (DHF/DSS-criteria), (c) abdominal pain, persistent vomiting, mucosal bleeding, and thrombocytopenia (SD warning signs) and (d) dyspnoea, but without fluid accumulation. DENV-infected patients with prior SARS-CoV-2 infections had significantly increased incidences of thrombocytopenia (DHF/DSS-criteria) and/or abdominal pain and persistent vomiting and also thrombocytopenia (SD warning signs), but without significant haemoconcentration or hypotension. DENV-SARS-CoV-2 co-infections significantly increased the incidence of DHF/DSS/SD, while DENV-infected patients with prior SARS-CoV-2 infections displayed significantly increased incidences of thrombocytopenia (DHF/DSS-criteria) and three important SD warning signs, which are therefore very important for health workers/clinicians in assessing patients' DHF/DSS/SD risk factors and planning their optimal therapies. [ABSTRACT FROM AUTHOR]

Abstract Background Epitope-mapping of infectious agents is essential for pathogenesis studies. Since polyclonal antibodies (PAbs) and monoclonal antibodies (MAbs) are always polyspecific and can react with multiple epitopes, it is important to distinguish between specific and non-specific reactions. Relative antibody discriminating specificity (RADS) values, obtained from their relative ELISA reactions with L-amino acid peptides prepared in the natural versus reverse orientations (x-fold absorbance natural/absorbance reverse = RADS value) may be valuable for this purpose. PAbs generated against the dengue type-2 virus (DENV-2) nonstructural-1 (NS1) glycoprotein candidate vaccine also reacted with both DENV envelope (E) glycoproteins and blood-clotting proteins. New xKGSx/xSGKx amino acid motifs were identified on DENV-2 glycoproteins, HIV-1 gp41 and factor IXa. Their potential roles in DENV and HIV-1 antibody-enhanced replication (AER) and auto-immunity were assessed. In this study, a) RADS values were determined for MAbs and PAbs, generated in congeneic (H2: class II) mice against DENV NS1 glycoprotein epitopes, to account for their cross-reaction patterns, and b) MAb 1G5.3 reactions with xKGSx/xSGKx motifs present in the DENV-4 NS1, E and HIV-1 glycoproteins and factor IXa were assessed after the introduction of amino acid substitutions, deletions, or intra-/inter-cysteine (C-C) bridges. Results MAbs 1H7.4, 5H4.3, 3D1.4 and 1G5.3 had high (4.23- to 16.83-fold) RADS values against single epitopes on the DENV-2 NS1 glycoprotein, and MAb 3D1.4 defined the DENV complex-conserved LX1 epitope. In contrast, MAbs 1G5.4-A1-C3 and 1C6.3 had low (0.47- to 1.67-fold) RADS values against multiple epitopes. PAb DENV complex-reactions occurred through moderately-high (2.77- and 3.11-fold) RADS values against the LX1 epitope. MAb 1G5.3 reacted with xSGKx motifs present in DENV-4 NS1 and E glycoproteins, HIV-1 gp41 and factor IXa, while natural C-C bridge formations or certain amino acid substitutions increased its binding activity. Conclusions These results: i) were readily obtained using a standard 96-well ELISA format, ii) showed the LX1 epitope to be the immuno-dominant DENV complex determinant in the NS1 glycoprotein, iii) supported an antigenic co-evolution of the DENV NS1 and E glycoproteins, and iv) identified methods that made it possible to determine the role of anti-DENV PAb reactions in viral pathogenesis.

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Objective: To evaluate the case report forms and times elapsed between the surveillance steps for dengue virus (DENV) infection in a large Colombian city before the emergence of other arbovirus epidemics., Materials and Methods: The descriptive epidemiology of DENV infection cases was analyzed from 2009 to 2013. The completeness of the case report forms filed at the Primary Units of Data Generation (PUDG) were evaluated, as well as the accuracy and suitability of the tests (PPV: positive predictive value). The average time-lags between each step were then calculated., Results: There were 7.3, 12.38, 4.66, 6.25 and 29.9 annual cases of dengue infection per 10 000 inhabitants in 2009 to 2013, respectively. In this study, only 57.76% of the cases were classified correctly by the physicians and 26.32% of them were questioned about their home conditions and whether their family/friends had similar symptoms. Patients visited a clinic/hospital on average 4.76 days after developing symptoms and the health system was notified on average 1.75 days later, while 70.6% of them were reported within the one-day target period. There were only minor changes in case reporting times even during a DENV epidemic. Some (12.85%) of the case forms were later modified (average 16.7 days). In the period 2009-2013, the IgM confirmed PPV was 58.60%, while 20 mandatory criteria were absent on more than 25% of the forms., Conclusions: The system was accurate, simple, flexible, stable and acceptable, but a number of ways are suggested to improve this case detection and reporting system.

Background: In 2015, the Zika arbovirus (ZIKV) began circulating in the Americas, rapidly expanding its global geographic range in explosive outbreaks. Unusual among mosquito-borne diseases, ZIKV has been shown to also be sexually transmitted, although sustained autochthonous transmission due to sexual transmission alone has not been observed, indicating the reproduction number (R0) for sexual transmission alone is less than 1. Critical to the assessment of outbreak risk, estimation of the potential attack rates, and assessment of control measures, are estimates of the basic reproduction number, R0., Methods: We estimated the R0 of the 2015 ZIKV outbreak in Barranquilla, Colombia, through an analysis of the exponential rise in clinically identified ZIKV cases (n=359 to the end of November, 2015)., Findings: The rate of exponential rise in cases was ρ=0.076days -1 , with 95% CI [0.066,0.087] days -1 . We used a vector-borne disease model with additional direct transmission to estimate the R0; assuming the R0 of sexual transmission alone is less than 1, we estimated the total R0=3.8 [2.4,5.6], and that the fraction of cases due to sexual transmission was 0.23 [0.01,0.47] with 95% confidence., Interpretation: This is among the first estimates of R0 for a ZIKV outbreak in the Americas, and also among the first quantifications of the relative impact of sexual transmission., (Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Detection of dengue virus (DENV) soluble/excreted (s/e) form of the nonstructural-1 (NS1) glycoprotein in patient acute-phase sera is ideal for diagnosis. The commercially-available detection assays are, however, too expensive for routine use and have low specificity, particularly for the s/e NS1 glycoprotein of DENV-2 and DENV-4, which are important causes of lethal human disease worldwide., Methods: Mouse monoclonal antibodies (MAbs) were generated and screened against s/e NS1 glycoprotein purified from each DENV serotype to obtain those that reacted equally with each serotype, but not with yellow fever virus (YFV) s/e NS1 glycoprotein or human serum proteins. One MAb, MAb 2C4.6, was further tested against these DENV glycoproteins in human sera using simple, peroxidase-labelled secondary antibody/substrate-developed dot-blot assays., Results: Optimal quenching of endogenous human serum peroxidases was attained using 3% H(2)O(2) in H(2)0 for 5 min. MAb 2C4.6 showed an acceptable detection sensitivity of < 32 ng/ml for the s/e NS1 glycoprotein of each DENV serotype but did not cross-react with the YFV s/e NS1 glycoprotein or human serum proteins. By contrast, the LX1 epitope-specific MAb, 3D1.4, showed similar detection sensitivity against only the DENV-1 NS1 glycoprotein, consistent with results from commercial DENV s/e NS1 glycoprotein detection assays.DENV s/e NS1 glycoproteins were stable in human sera after drying on the nitrocellulose membranes and storage for one month at ambient temperature (28°C) before being processed. The total assay time was reduced to 3 h without any loss of detection sensitivity. This dot-blot format was ideal for the circulating immune complex disruption step, which is required for increased DENV s/e NS1 glycoprotein detection., Conclusions: This is the first study to determine the detection sensitivity of MAbs against known concentrations of s/e NS1 glycoprotein from each DENV serotype. The preparation of patient serum samples for dot-blot assays can be performed by staff with a basic level of training and storage at low temperatures (e.g., -80°C) is not necessary. These simple, inexpensive (US$ 0.05/sample), robust, sensitive and relatively rapid assays, using improved MAbs such as MAb 2C4.6, should be ideal for the diagnosis of all DENV serotypes in DENV endemic regions.

Background: SEVERE DENGUE DISEASE (SDD) (DHF/DSS: dengue hemorrhagic fever/dengue shock syndrome) results from either primary or secondary dengue virus (DENV) infections, which occur 4 - 6 days after the onset of fever. As yet, there are no definitive clinical or hematological criteria that can specifically identify SDD patients during the early acute febrile-phase of disease (day 0 - 3: < 72 hours). This study was performed during a SDD (DHF/DSS) epidemic to: 1) identify the DENV serotypes that caused SDD during primary or secondary DENV infections; 2) identify simple clinical and hematological criteria that could significantly discriminate between patients who subsequently developed SDD versus non-SDD (N-SDD), or had a non-DENV fever of unknown origin (FUO) during day 0 - 3 of fever; 3) assess whether DENV serotype co-infections resulted in SDD., Methods: First serum samples, with clinical and hematological criteria, were collected from 100 patients during the early acute febrile-phase (day 0 - 3: < 72 hours), assessed for DENV or FUO infections by IgM- and IgG-capture ELISAs on paired serum samples and by DENV isolations, and subsequently graded as SDD, N-SDD or FUO patients., Results: IN THIS STUDY: 1) Thirty-three patients had DENV infections, predominantly secondary DENV-2 infections, including each SDD (DHF/DSS) case; 2) Secondary DENV-2/-3 and DENV-2/-4 serotype co-infections however resulted in N-SDD; 3) Each patient who subsequently developed SDD, but none of the others, displayed three clinical criteria: abdominal pain, conjunctival injection and veni-puncture bleeding, therefore each of these criteria provided definitively significant prognostic (P < 0.001) values; 4) Petechia, positive tourniquet tests and hepatomegaly, and neutrophilia or leukopenia also significantly identified those who: a) subsequently developed SDD versus N-SDD, or had a FUO; b) subsequently developed SDD versus N-SDD; c) subsequently developed N-SDD versus FUOs, respectively., Conclusions: This is the first report of simple definitively prognostic criteria for SDD patients, including the first assessment and confirmation of conjunctival injection. The three definitive clinical criteria used alone, or supported by the other four criteria, could be essential for specifically identifying those patients needing prompt hospital-based therapies to lessen or avert SDD, without unnecessary hospitalization of the other patients., Keywords: Dengue virus; Severe dengue; Dengue fever; Diagnostic; Criteria; Hemorrhage; Shock.

Introduction: Since the methodologies used to calculate Stegomyia indices have been shown to be inadequate for assessing the risk of dengue virus transmission and targeting Aedes aegypti control strategies, new surveillance methods are needed., Objective: To evaluate the water-surface sweeping method in combination with calibration factors to estimate the total number of Ae. aegypti late larval stages (L3/L4) in large water-storage containers at different temperatures at which transmission of dengue virus occurs., Materials and Methods: Calibration factors were derived based on the proportion of L3/L4 recovered from a predetermined number of larvae using a net of specific dimensions and water-storage containers of different capacities and water levels in semi-field conditions and at four different altitudes (14, 358, 998 and 1,630 meters above sea level). The calibration factors obtained at 14 masl were then fully validated in a field study site at this altitude., Results: Four calibration factors were derived at 14 masl (28-30°C) that were used to estimate the total L3/L4 numbers in large water storage containers greater than 20 L (n=478) at 1/3, 2/3 and full water-levels. This methodology was accurate and robust within and between the 10 pairs of field workers who applied it. Different calibration factors were, however, derived to accurately estimate the total L3/L4 numbers at each of the study sites located at 358, 998 and 1,630 masl, where average temperatures were 19°C, 24°C, and 26°C respectively., Conclusions: The accurate estimates of L3/L4 numbers calculated using the water surface sweeping method can be useful for evaluating intervention strategies directed against the larval stages.

A sampling method coupled with statistical calibration factors was developed to accurately assess the numbers of larvae and pupae of Aedes aegypti in large water-storage containers of variable capacities and water levels. Aedes aegypti productivity in different types of breeding sites found in an urban study area in central Colombia was assessed and compared. In this study, water-storage tanks and drums were found to comprise 79% of the containers positive for larval Ae. aegypti, which contributed to 93 and 92% of the total production of populations of 4th-stage larvae and pupae, respectively. These main breeding sites of Ae. aegypti were found at an indoor to outdoor ratio of 2.4:1 and no correlation was found between temporal fluctuation of populations of larval Ae. aegypti and monthly rainfall. Netted lids that used inexpensive local materials were designed to prevent oviposition by Ae. aegypti. During a 6-month trial period, 56% of inspected containers had netted lids correctly in place. Of these, 78% had no mosquito larvae. Because only 37% of uncovered containers were free of mosquito larvae, a significant difference was demonstrated when these inexpensive mechanical barriers were used (chi2 = 138.7; P < 0.001). These netted lids and the improved methods described to assess the productivity of larval and pupal Ae. aegypti in this study are now being used in combination with other strategies to assess and control these populations of dengue virus vectors in the main port city on the Atlantic Coast of Colombia.