Identification of differences in the magnitude and specificity of SARS-CoV-2 nucleocapsid antibody responses in naturally infected and vaccinated individuals.

As there are limited data on B-cell epitopes for the nucleocapsid protein in SARS-CoV-2, we sought to identify the immunodominant regions within the N protein, recognized by patients with varying severity of natural infection with the Wuhan strain (WT), delta, omicron, and in those who received the Sinopharm vaccines, which is an inactivated, whole virus vaccine. Using overlapping peptides representing the N protein, with an in-house ELISA, we mapped the immunodominant regions within the N protein, in seronegative (n = 30), WT infected (n = 30), delta infected (n = 30), omicron infected + vaccinated (n = 20) and Sinopharm (BBIBP-CorV) vaccinees (n = 30). We then investigated the sensitivity and specificity of these immunodominant regions and analyzed their conservation with other SARS-CoV-2 variants of concern, seasonal human coronaviruses, and bat Sarbecoviruses. We identified four immunodominant regions aa 29-52, aa 155-178, aa 274-297, and aa 365-388, which were highly conserved within SARS-CoV-2 and the bat coronaviruses. The magnitude of responses to these regions varied based on the infecting SARS-CoV-2 variants, >80% of individuals gave responses above the positive cut-off threshold to many of the four regions, with some differences with individuals who were infected with different VoCs. These regions were found to be 100% specific, as none of the seronegative individuals gave any responses. As these regions were highly specific with high sensitivity, they have a potential to be used to develop diagnostic assays and to be used in development of vaccines.

Adipokine levels and their association with clinical disease severity in patients with dengue.

Adipokines have not been studied in acute dengue, despite their emerging role in inducing and regulating inflammation. Therefore, we sought to identify adipokine levels in patients with varying severities of acute dengue to understand their role in disease pathogenesis. We determined the levels of leptin, resistin, omentin, adiponectin, as well as IFNß, and NS1 using quantitative ELISA in patients with dengue fever (DF = 49) and dengue haemorrhagic fever (DHF = 22) at admission (febrile phase) and at the time of discharge (recovery phase). The viral loads and serotypes of all samples were quantified using quantitative real-time RT-PCR. Resistin levels (p = 0.04) and omentin (p = 0.006) levels were significantly higher in patients who developed DHF. Omentin levels in the febrile phase also correlated with the AST (Spearman's r = 0.38, p = 0.001) and ALT levels (Spearman's r = 0.24, p = 0.04); as well as serum leptin levels with both AST (Spearman's r = 0.27, p = 0.02) and ALT (Spearman's r = 0.28, p = 0.02). Serum adiponectin levels in the febrile phase did not correlate with any of the other adipokines or with liver enzymes, but inversely correlated with CRP levels (Spearman's r = -0.31, p = 0.008). Although not significant (p = 0.14) serum IFNß levels were lower in the febrile phase in those who progressed to develop DHF (median 0, IQR 0 to 39.4 pg/ml), compared to those who had DF (median 37.1, IQR 0 to 65.6 pg.ml). The data suggest that adipokines are likely to play a role in the pathogenesis of dengue, which should be further explored for the potential to be used as prognostic markers and as therapeutic targets.

Identification of differences in the magnitude and specificity of SARS-CoV-2 nucleocapsid antibody responses in naturally infected and vaccinated individuals.

Background: As there are limited data on B cell epitopes for the nucleocapsid protein in SARS-CoV-2, we sought to identify the immunodominant regions within the N protein, recognized by patients with varying severity of natural infection with the Wuhan strain (WT), delta, omicron and in those who received the Sinopharm vaccines, which is an inactivated, whole virus vaccine. Methods: Using overlapping peptides representing the N protein, with an in-house ELISA, we mapped the immunodominant regions within the N protein, in seronegative (n=30), WT infected (n=30), delta infected (n=30), omicron infected+vaccinated (n=20) and Sinopharm (BBIBP-CorV) vaccinees (n=30). We then investigated the sensitivity and specificity of these immunodominant regions and analysed their conservation with other SARS-CoV-2 variants of concern, seasonal human coronaviruses and bat Sarbecoviruses. We then investigated the kinetics of responses to these regions in those with varying severity of acute COVID-19. Results: We identified four immunodominant regions aa 29-52, aa 155-178, aa 274 to 297 and aa 365 to 388, were highly conserved within SARS-CoV-2 and the bat coronaviruses. The magnitude of responses to these regions varied based on the infecting SARS-CoV-2 variants, with WT infected individuals predominantly recognizing aa155 to 178 regions, delta infected individuals and vaccinated+omicron infected individuals predominantly recognizing regions aa 29 to 52 and aa 274 to 294 regions. Sinopharm vaccinees recognized all four regions, with the magnitude of responses significantly lower than other groups. >80% of individuals gave responses above the positive cut-off threshold to many of the four regions, with some differences with individuals who were infected with different VoCs. These regions were found to be 100% specific, as none of the seronegative individuals gave any responses. Conclusions: N-protein specific responses appear to be detectable in over 90% of those who were naturally infected or vaccinated with a whole virus inactivated vaccine, with responses mainly directed against four regions of the protein, which were highly conserved. As these regions were highly specific with high sensitivity, they have a potential to be used to develop diagnostic assays and to be used in development of vaccines.

Adipokine levels and their association with clinical disease severity in patients with dengue.

Adipokines have not been studied in acute dengue, despite their emerging role in inducing and regulating inflammation. Therefore, we sought to identify adipokine levels in patients with varying severities of acute dengue to understand their role in disease pathogenesis. We determined the levels of leptin, resistin, omentin, adiponectin, as well as IFNß, and NS1 using quantitative ELISA in patients with dengue fever (DF=49) and dengue haemorrhagic fever (DHF=22) at admission (febrile phase) and at the time of discharge (recovery phase). The viral loads and serotypes of all samples were quantified using quantitative real-time RT-PCR. Resistin levels (p =0.04) and omentin (p=0.006) levels were significantly higher in patients who developed DHF. Omentin levels in the febrile phase also correlated with the AST (Spearman's r=0.38, p=0.001) and ALT levels (Spearman's r=0.24, p=0.04); as well as serum leptin levels with both AST (Spearman's r=0.27, p=0.02) and ALT (Spearman's r=0.28, p=0.02). Serum adiponectin levels in the febrile phase did not correlate with any of the other adipokines or with liver enzymes, but inversely correlated with CRP levels (Spearman's r=-0.31, p=0.008). Although not significant (p=0.14) serum IFNß levels were lower in the febrile phase in those who progressed to develop DHF (median 0, IQR 0 to 39.4 pg/ml), compared to those who had DF (median 37.1, IQR 0 to 65.6 pg.ml). The data suggest that adipokines are likely to play a role in the pathogenesis of dengue, which should be further explored for the potential to be used as prognostic markers and as therapeutic targets.

Comparison of the kinetics and magnitude of antibody responses to different SARS-CoV-2 proteins in Sinopharm/BBIBP-CorV vaccinees following the BNT162b2 booster or natural infection.

The kinetics and magnitude of antibody responses to different proteins of the SARS-CoV-2 virus in Sinopharm/BBIBP-CorV vaccinees has not been previously studied. Therefore, we investigated antibody responses to different SARS-CoV-2 proteins at 2 weeks, 3 months, and 6 months post-second dose in previously infected (n = 20) and uninfected (n = 20) Sinopharm/BBIBP-CorV vaccinees. The IgG antibodies to the S, S1 and S2 and N were several folds higher in those who had natural infection compared to uninfected individuals at all time points. We then compared the persistence of antibody responses and effect of natural omicron infection or BNT162b2 booster in Sinopharm/BBIBP-CorV vaccinees. We measured the total antibodies to the RBD, ACE2 blocking antibodies and antibody responses to different SARS-CoV-2 proteins in Sinopharm vaccinees at 7 months post second dose, including those who remained uninfected and not boosted (n = 21), or those who had previous infection and who did not obtain the booster (n = 17), those who were not infected, but who received a BNT162b2 booster (n = 30), or those who did not receive the booster but were infected with omicron (n = 29). At 7 months post second dose uninfected (no booster) had the lowest antibody levels to the RBD, while omicron infected vaccinees showed significantly higher anti-RBD antibody levels (p = 0.04) than vaccinees who received the booster. Only 3/21 cohort A (14.3%) had ACE2 blocking antibodies, while higher frequencies were observed in naturally infected individuals (100%), those who received the booster (18/21, 85.7%), and omicron infected individuals (100%). Pre-vaccination, naturally infected had the highest antibody levels to the N protein. These data suggest that those previously infected Sinopharm/BBIBP-CorV vaccinees have a robust antibody response, 7 months post vaccination, while vaccinees who were naturally infected with omicron had a similar immune response to those who received the booster. It will be important to investigate implications for subsequent clinical protection.