India has been hit hard by COVID-19, with at least 66,000 deaths and almost 4 million cases. The need to understand the infection, including the various antibody isotypes, and whether neutralizing antibodies and memory B cells are being produced, has driven much research into the way immunity is developing in Indian patients. A recent study published in the preprint server bioRxiv* in August 2020 reports significant variation in these parameters among COVID-19 convalescents.
This news article was a review of a preliminary scientific report that had not undergone peer-review at the time of publication. Since its initial publication, the scientific report has now been peer reviewed and accepted for publication in a Scientific Journal. Links to the preliminary and peer-reviewed reports are available in the Sources section at the bottom of this article. View Sources
Anti-RBD Antibodies
Since many of the neutralizing epitopes are on the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) receptor binding domain (RBD), the researchers measured IgG, IgA, and IgM antibodies to the RBD, as well as the neutralization activity and memory B cells, in healthy and convalescent COVID-19 subjects in India. They aimed to find out if anti-RBD antibodies reflected neutralizing antibodies.
Patient Characteristics
They evaluated antibodies in the blood samples taken from recovered COVID-19 patients at three hospital centers in North India. The mean age of the participants was 39 years, and all were tested positive for the virus by polymerase chain reaction (PCR) at first diagnosis but had reverted to PCR-negative at the time of the study. All patients were between 3.6 and 12 weeks from diagnosis at this point.
The researchers also measured these parameters in blood samples drawn from adult blood donors in 2018 as pre-pandemic controls.
High Anti-RBD Titers in Only Half
They found that antibodies targeting the RBD were highly elevated in convalescent patients relative to the controls, but all isotypes showed substantial difference in titer between individual patients. IgG, IgA, and IgM titers varied from undetectable to above 24,000, over 5,600, and almost 3,000, respectively.
Undetectable titers of IgG and IgA were found in four patients, and one of them had undetectable IgM as well. The antibody titer failed to show any obvious association with age or to the time since the initial diagnosis.
Neutralization Tests Show Variation Among Patients
The researchers then carried out a neutralization assay, using plasma at different dilutions. They found that neutralization of 50% of the virus occurred at a dilution as high as 1:20 in only half the tested patients, while none of the controls showed detectable activity. The neutralizing activity also appeared to be highly variable among individuals.
Anti-RBD IgG Correlates with Neutralization Titer
They then attempted to make a connection between the titer of any specific isotype and the neutralizing activity. They found that only anti-RBD IgG titers were linked to SARS-CoV-2 neutralizing activity.
Plasma therapy is appearing on the scene as a form of treatment for COVID-19 in India but requires that individuals with high plasma neutralizing activity be identified accurately in a cost-effective manner. At present, commercial ELISA tests are used to test for anti-SARS-CoV-2 antibodies in plasma. The researchers used one such test detecting IgG against the viral antigens to compare the IgG titer with neutralization titer. They found that 33 of 42 recovered patients tested positive for IgG. Of the remaining 9 individuals, 4 were negative for anti-RBD IgG by ELISA as well. The measurement of IgG antibodies targeting the whole virus failed to show a good correlation with neutralizing antibody titer, in contrast to those IgGs that were directed against the RBD.
Anti-RBD IgG Correlates with Memory B Cells
The researchers also counted memory B cells targeting the RBD since these are essential for rapid recognition and prevention of reinfection by the speedy large-scale production of antibodies. They found that here, too, there was significant variability among individuals in the memory B cell frequency. However, their frequency was moderately correlated with the titer of anti-RBD IgGs.
Implications for Plasma Therapy
The researchers say, “Our correlative analysis of RBD-specific IgG binding titers with neutralizing antibody titers and memory B cells has important implications for not only identifying potential donors for plasma therapy but also for understanding humoral and cellular memory post COVID-19.”
The United States Food and Drug Administration (FDA) guidelines advise using plasma with a neutralizing antibody titer of 1:160 or 1:80 for treating COVID-19 patients. However, India does not have such an advisory at present. In light of this, it is notable that this study provides a less expensive surrogate for neutralizing activity, which requires specific neutralization assays, in the form of anti-RBD-IgG titers. The strong correlation between the latter and the neutralization titer allows the identification of plasma with high neutralization titers.
When the anti-RBD IgG titer is ~3,700, it corresponds to a neutralizing titer of 1: 160, while at ~ 1,900, it correlates with neutralizing titers of 1: 80. These observations require validation in more extensive studies to test their strength but may be necessary for directing further research.
Implications and Future Directions
The researchers point out the large number of individuals who recovered from COVID-19 but failed to demonstrate neutralizing activity at a dilution of 1: 20. The reasons may include different levels of exposure and viral loads, genetic factors, and the severity of the clinical illness.
In fact, earlier studies suggest that individuals with more severe disease also had higher levels of neutralizing antibodies. The current study included mostly individuals who had mild to moderate symptoms, which may underlie the poor development of neutralizing activity and adaptive immunity.
The question now is whether such low neutralizing titers in over half the cohort are reflected in poor immunological memory by immune cells also.
The study suggests that, indeed, those patients who failed to develop neutralizing antibodies also had lower frequencies of memory B cells. Of course, further research is mandatory since T cells are equally or more important in establishing immunity against this virus. Moreover, it must be established whether memory T cells also show the same degree of inter-individual variability.
And of course, the all-important question remains: will recovered patients with low or undetectable neutralizing activity or IgG titers, or memory B cells, be protected against reinfection, with either SARS-CoV-2 or a similar virus? Only more research into this phenotype and the associated immune response can afford an answer.
This news article was a review of a preliminary scientific report that had not undergone peer-review at the time of publication. Since its initial publication, the scientific report has now been peer reviewed and accepted for publication in a Scientific Journal. Links to the preliminary and peer-reviewed reports are available in the Sources section at the bottom of this article. View Sources
Article Revisions
- May 18 2023 - The preprint preliminary research paper that this article was based upon was accepted for publication in a peer-reviewed Scientific Journal. This article was edited accordingly to include a link to the final peer-reviewed paper, now shown in the sources section.
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