Longitudinal clinico-serological analysis of anti-nucleocapsid and anti-receptor binding domain of spike protein antibodies against SARS-CoV-2

Abstract

Objectives: Monitoring the antibody responses to SARS-CoV-2 infection and its correlation to clinical spectrum of disease is critical in understanding the disease progression and protection against re-infection. We assessed the nucleocapsid (N) and receptor-binding-domain of spike (SRBD) protein specific IgG and neutralizing antibody (NAb) responses in COVID-19 patients up to 8 months and its correlation with diverse disease spectrum.

Methods: During the first wave of the SARS-CoV-2 pandemic, from 284 COVID-19 patients, 608 samples were collected up to 8 months post infection. The patients were categorized as asymptomatic, symptomatic and severe. The N and SRBD IgG and NAb titers were evaluated and correlated with clinical data.

Results: A steep increase in antigen specific antibody titers was observed till 40 days post onset of the disease (POD), followed by a partial decline till 240 days. Severe disease was associated with a stronger SRBD IgG response and higher NAb titers. The persistence of antibody response was observed in 76% against N, 80% against SRBD and 80% for NAbs of cases up to 8 months POD.

Conclusion: RBD and N protein specific IgG persisted till 240 days POD which correlated with NAb response, irrespective of individual`s symptomatic status indicating overall robust protection against re-infection.

Keywords: 240 days; IgG antibody response; SARS-CoV-2; neutralizing antibody response; nucleocapsid; receptor binding domain.

Figure 1

Humoral immune response assessed in 608 samples collected from 284 COVID-19 patients. Antibody responses to SARS-CoV-2 against N and SRBD proteins (A-B); neutralizing antibody activity (C) in acute and convalescent samples; Correlation of SRBD IgG with N IgG antibody levels (D) (r = 0.6848, p < 0.0001); Correlation of SARS-CoV-2 N IgG antibody levels and neutralizing antibody titers (E) (r = 0.7207, p < 0.0001); Correlation of SARS-CoV-2 SRBD IgG antibody levels and neutralization antibody titers (F) (r = 0.7911, p < 0.0001).

Figure 2

Longitudinal responses to SARS-CoV-2 IgG and NAbs till 240 days POD. The longitudinal trends of N (blue) and SRBD (red) specific IgG and neutralizing antibodies (green) with trend line showing geometric mean titers of each parameter for 608 samples from 284 patients collected over a period of 240 days POD.

Figure 3

Seropositivity observed for N IgG (blue) and SRBD IgG (red) among 284 patients collected from 0-240 days POD. Seropositivity for both antibodies show a steep upward incline till 28 days POD and a gradual decline as the disease progressed.

Figure 4

Levels of IgG antibodies and titers of NAbs against SARS-CoV-2 of asymptomatic (blue), symptomatic (red) and severe symptomatic (green) cases among 284 patients collected from 0-240 days POD. Antibody levels are expressed as ratio values (A, B) and NAb titers are plotted (C). The median and quartiles were represented in the violin plots. (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001)

Figure 5

Immune response across diverse disease severity with respect to neutrophil to lymphocyte ratio (NLR) and IgG antibody levels (N = 125). Association between A) N IgG ratio and NLR (cut-off = 3.04); B) SRBD IgG ratio and neutrophil to lymphocyte ratio for 125 patents. (Asymptomatic = 42, Symptomatic = 68 and severe symptomatic = 15 cases.)

Figure 6

Associations of immune response and disease severity correlogram of COVID-19 patients: A) Asymptomatic cases, B) Symptomatic cases and C) Severe symptomatic cases). Spearman rank order correlation values (r) are shown from red (-1.0) to blue (1.0); r values are indicated by color and square size.