Lack of cross-reactivity between anti-A IgG isoagglutinins and anti-SARS-CoV-2 IgG antibodies

To the Editor,

The ABO blood group is the most important among human blood group systems and consists of complex carbohydrate moieties at the extracellular surface of red blood cell (RBC) membrane [1]. Along with their expression on RBCs, ABO antigens (namely, A, B, AB, and O) are also highly expressed on the surface of a variety of human cells and tissues [2]. Although the physiologic role of ABO antigens and their related anti-A and anti-B natural isoagglutinins is still largely unknown, they play a prominent role in blood transfusion and cell, tissue, and organ transplantation [2]. In addition, several studies have documented over the last 50 years a close link between ABO blood groups and a wide array of diseases, including cancers and cardiovascular disorders [3]. The latter association is particularly relevant, considering the profound influence of ABO antigens on hemostasis, particularly in modulating von Willebrand factor (WVF) and factor VIII (FVIII) circulating levels [4]. Also the ABO blood group-related susceptibility to various types of viral infections, including HIV, hepatitis B, dengue and influenza viruses, has been consistently reported by several investigators over the last 20 years [5]. This issue has recently gained a renewed interest thanks to the first observations on the association between ABO blood type and Coronavirus Disease 2019 (COVID-19) from China, where the infection began and quickly spread around the world [6]. In particular, it has been hypothesized that individuals belonging to O blood type are less susceptible to Severe Acute Respiratory Syndrome Coronavirus 2 (SARS‐CoV‐2) infection than those belonging to non‐O blood groups or that have a milder disease [7]. The hypothesized reason for this phenomenon lies in the presence in O blood group subjects of anti‐A isoagglutinins, predominantly of IgG class, which would prevent the binding of SARS‐CoV‐2 to its angiotensin converting enzyme (ACE)-2 receptor thereby inhibiting the virus entry into the targeted human cells [8]. Given such association, in this study we wanted to check if there is a cross-reactivity between anti-A IgG isoagglutinins and anti-SARS-CoV-2 IgG antibodies in patients recovered from COVID-19 selected for convalescent plasma donation.

Serum samples from 12 consecutive convalescent plasma donors belonging to blood group O (group I-cases) were collected and anti-A IgG antibodies were titrated according to standard technique (dilution phosphate-buffered saline-treated control serum and dithiothreitol-treated serum) using a micro-column technology (Ortho Clinical Diagnostics, Raritan, NJ, USA) [9]. All these samples were also titrated for anti-SARS-CoV-2 neutralizing antibodies using the plaque reduction neutralization test (PRNT), as previously described [10]. In addition, the chemiluminescent immunoassay (CLIA)-based technology (LIAISON SARS-CoV-2 S1/S2 IgG, DiaSorin, Vercelli, Italy) was used on these serum samples for the quantitative determination of anti-SARS-CoV-2 IgG antibodies, which were measured at basal and post-adsorption (incubation for 60 min at 37 °C) on blood group A RBCs, to remove from convalescent plasma donors’ serum anti-A IgG antibodies [10]. All these tests were performed also in a control group (group II) of 12 consecutive O blood type convalescent plasma donors, with the only difference that their serum was adsorbed on O blood type RBCs for the measurement of post-RBC adsorption anti-SARS-CoV-2 IgG antibodies.

The two groups (group I-cases and group II-controls) of convalescent plasma donors were comparable in terms of male-female ratio (5.0 vs. 3.3) and median age (46.5 vs. 46.0 years). The median anti-A IgG isoagglutinin titer and anti-SARS-CoV-2 neutralizing titer in the study group (group I) were 96 (range 32–256) and 80 (range 20–320), respectively. The median anti-A IgG isoagglutinin titer and anti-SARS-CoV-2 neutralizing titer in the control group (group II) were 80 (range 32–256) and 120 (range 20–320), respectively. No significant differences were observed in these parameters (i.e., anti-A IgG isoagglutinin titer and anti-SARS-CoV-2 neutralizing titer) between the two groups. Regarding the anti-SARS-CoV-2 IgG CLIA-based assay, the median basal and post-RBC (blood group A) serum adsorption levels in group I were 82 UA/mL (range 8–249 UA/mL) and 73 UA/mL (range 7–240 UA/mL), respectively. In group II (control group), the median basal and post-RBC (blood group O) anti-SARS-CoV-2 IgG levels were 95 UA/mL (range 33–265 UA/mL), and 81 UA/mL (range 22–254 UA/mL), respectively (Table 1). Using Student’s t test for statistical analysis, no statistically significant difference between basal and post-RBC adsorption anti-SARS-CoV-2 IgG levels were detected in both group I and group II convalescent plasma donors. No difference was also observed comparing the same parameters between the two groups. A similar difference between median anti-SARS-CoV-2 IgG levels pre- and post-RBC adsorption studies was observed in group I and group II (11 and 15%, respectively). Thus, considering that in group I the donors’ serum was adsorbed on A blood type RBCs and in group II it was adsorbed on O blood type RBCs, this slight difference was certainly not due to the presence of anti-A IgG isoagglutinins but rather to the adsorption procedure itself.

In conclusion, in this study we demonstrated the lack of cross-reactivity between anti-A IgG isoagglutinins and anti-SARS-CoV-2 IgG antibodies measured using an immunoassay, therefore excluding a possible interference of isoagglutinins on seroneutralizing tests.