Durability and extent of protection of SARS-CoV-2 antibodies among patients with COVID-19 in Metro Manila, Philippines

Predictive models of immune protection from COVID-19 are urgently needed to identify correlates of protection to assist in the future deployment of vaccines. To address this, we analyzed the relationship between in vitro neutralization levels and the observed protection from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection using data from seven current vaccines and from convalescent cohorts. We estimated the neutralization level for 50% protection against detectable SARS-CoV-2 infection to be 20.2% of the mean convalescent level (95% confidence interval (CI) = 14.4-28.4%). The estimated neutralization level required for 50% protection from severe infection was significantly lower (3% of the mean convalescent level; 95% CI = 0.7-13%, P = 0.0004). Modeling of the decay of the neutralization titer over the first 250 d after immunization predicts that a significant loss in protection from SARS-CoV-2 infection will occur, although protection from severe disease should be largely retained. Neutralization titers against some SARS-CoV-2 variants of concern are reduced compared with the vaccine strain, and our model predicts the relationship between neutralization and efficacy against viral variants. Here, we show that neutralization level is highly predictive of immune protection, and provide an evidence-based model of SARS-CoV-2 immune protection that will assist in developing vaccine strategies to control the future trajectory of the pandemic.

The global supply of COVID-19 vaccines remains limited. An understanding of the immune response that is predictive of protection could facilitate rapid licensure of new vaccines. Data from a randomized efficacy trial of the ChAdOx1 nCoV-19 (AZD1222) vaccine in the United Kingdom was analyzed to determine the antibody levels associated with protection against SARS-CoV-2. Binding and neutralizing antibodies at 28 days after the second dose were measured in infected and noninfected vaccine recipients. Higher levels of all immune markers were correlated with a reduced risk of symptomatic infection. A vaccine efficacy of 80% against symptomatic infection with majority Alpha (B.1.1.7) variant of SARS-CoV-2 was achieved with 264 (95% CI: 108, 806) binding antibody units (BAU)/ml: and 506 (95% CI: 135, not computed (beyond data range) (NC)) BAU/ml for anti-spike and anti-RBD antibodies, and 26 (95% CI: NC, NC) international unit (IU)/ml and 247 (95% CI: 101, NC) normalized neutralization titers (NF50) for pseudovirus and live-virus neutralization, respectively. Immune markers were not correlated with asymptomatic infections at the 5% significance level. These data can be used to bridge to new populations using validated assays, and allow extrapolation of efficacy estimates to new COVID-19 vaccines.

To the editor Coronavirus disease 2019 (COVID-19), which emerged in Wuhan, China in December 2019, is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has become a major global public health concern [1]. Positive detection of SARS-CoV-2 RNA in nasopharyngeal swab samples, sputum samples or bronchoalveolar lavage samples by reverse transcriptase polymerase chain reaction (RT–PCR) has been used to confirm SARS-CoV-2 infection [2]. Recently, positive detection of IgM and IgG antibodies specific to SARS-CoV-2 has also been recognized as deterministic evidence for confirmed SARS-CoV-2 infection [3,4]. However, the antibody response to SARS-CoV-2 currently remains inadequately understood in COVID-19 patients. In the present study, we investigated the patterns of antibody response to SARS-CoV-2 in patients with COVID-19, aiming to better clarify the humoral immunological response during SARS-CoV-2 infection. Patient characteristics A total of 32 patients with a confirmed diagnosis of COVID-19 were included in our cohort. All were positive for SARS-CoV-2 according to nucleic acid testing by RT–PCR of nasopharyngeal swab, sputum, or bronchoalveolar lavage specimens. Patients exhibiting one or more of the following conditions were classified as having severe COVID-19: (a) respiratory distress (≥30 breaths/min); (b) oxygen saturation ≤93% at rest; (c) arterial partial pressure of oxygen (PaO2)/fraction of inspiration O2 (FiO2) ≤300 mmHg (1 mmHg = 0.133 kPa); (d) respiratory failure requiring mechanical ventilation; (e) septic shock development; or (f) critical organ failure requiring ICU care. Patients not meeting the above criteria were classified as having mild COVID-19. The median age of the 32 patients was 55 years old, and 66.7% of them were male. Among the 32 patients, 18 (56.3%) were severe cases, and 14 (43.7%) were mild cases. The most common symptoms at onset of illness were fever, cough, fatigue, dyspnoea and headache. The demographic details and clinical disease severity of all patients in our cohort are shown in the supplementary materials (Table 1). Our study was approved by the Ethics Committee of Chongqing Public Health Medical Center (No. 2020-006-01), and informed consent was obtained from all subjects prior to blood sample collection. Table 1. Demographic characteristics and antibody titre of 32 COVID-19 patients. Patient number Age Sex severe case Sample number ID day IgG (RU/ml) IgM (RU/ml) 1 73 male YES 2001278733 74827 4 0.94 15.57         2001312952 74827 8 177.44 176.69         2002059960 74827 13 664.2 282.74         2002093544 74827 17 731.94 314.94         2002127130 74827 20 665.58 236.87 2 52 male YES 2001267752 74833 7 39.92 129.6         2001290545 74833 10 189.55 194.99         2002048959 74833 16 202.85 460.93         2002093481 74833 21 246.72 632.64         2002127090 74833 24 233.22 464 3 70 male NO 2001299883 74839 10 98.27 11.88         2001313114 74839 12 874.23 44.57         2002059503 74839 17 1572.32 67.32         2002093351 74839 21 1377.84 66.91         2002140361 74839 26 1213.54 58.79 4 70 female YES 2001267391 74851 7 0.97 2.1         2001288808 74851 9 1.17 6.75         2001312862 74851 12 66.3 25.02         2002036073 74851 15 238.5 43.74         2002060585 74851 18 275.85 94.55         2002093998 74851 21 406.59 114.09 5 65 female YES 2001267790 74862 7 2.43 13.9         2001278746 74862 8 43.98 33.59         2001299874 74862 10 74.37 109.79         2001301290 74862 11 167.51 81.5         2001313232 74862 12 299.69 124.98         2002036823 74862 15 599.44 181.03         2002072533 74862 19 535.16 132.12         2002094013 74862 21 612.32 106.51         2002116878 74862 23 596.47 104.57         2002127664 74862 24 517.07 88.47         2002139300 74862 25 674.04 115.42         2002140505 74862 26 655.19 99.52 6 38 male YES 2001267654 74863 9 21.43 50.74         2001290540 74863 12 375.96 216.74         2002013601 74863 15 585.37 223.08         2002048134 74863 18 784.3 195.61         2002072220 74863 21 692.54 188.64         2002127169 74863 26 672.38 143.25 7 60 female NO 2001278322 74877 3 0.93 7.14         2001301110 74877 6 0.99 11.73         2002013575 74877 8 2.74 27.07         2002059329 74877 12 548.55 164.95         2002093476 74877 16 1067.37 255.72 8 38 male YES 2001289774 74912 9 83.75 114.35         2001290543 74912 10 141.4 145.23         2001301919 74912 11 190.77 184.79         2002025775 74912 14 213.92 259.38         2002048717 74912 16 187.13 344.38         2002071492 74912 19 199.88 408.83         2002127144 74912 24 221.51 491.04 9 56 female YES 2001290419 74921 4 0.93 5.06         2001313538 74921 6 2.78 8.81         2002014606 74921 7 2.63 18.31         2002061270 74921 12 21.88 329.72         2002082793 74921 14 68.72 373.78         2002104121 74921 16 127.14 436 10 55 female NO 2001290328 74929 9 194.35 232.74         2002013579 74929 12 266.34 233.96         2002047843 74929 15 324.93 259.5         2002071490 74929 18 350.07 283.65         2002105698 74929 21 330.38 250.95 11 67 male NO 2001290929 74935 6 1.03 17.41         2002014497 74935 9 0.93 49.75         2002025405 74935 10 4.37 115.26         2002047701 74935 12 6.34 113.27         2002072412 74935 15 199.16 193.81 12 38 male YES 2001290938 74940 3 0.93 0.93         2002014667 74940 6 0.93 3.91         2002048327 74940 9 1.52 32.12         2002083247 74940 13 380.16 144.28         2002105333 74940 15 345.5 167.62         2002116806 74940 16 438.22 134.65         2002138679 74940 18 458.95 159.62 13 50 male YES 2001301499 74948 10 197.05 186.68         2002013584 74948 12 342.23 282.93         2002047837 74948 15 437.01 354.36         2002071486 74948 18 392.84 440.22         2002105692 74948 21 352.06 380.02         2002140348 74948 25 408.94 379.38 14 69 female YES 2001301775 74952 2 0.93 0.93         2001313338 74952 3 0.93 1.9         2002037655 74952 6 2.37 17.23         2002050036 74952 8 38.18 53.33         2002072543 74952 10 105.89 49.7         2002083134 74952 11 95.69 65.68         2002093971 74952 12 87.47 61.8         2002116847 74952 14 203.35 63.8         2002127670 74952 15 217.68 68.85         2002140610 74952 17 292.36 73.84 15 46 female NO 2001301957 74959 3 1.25 2.66         2002025001 74959 6 0.93 4.72         2002037387 74959 7 1.59 9.16         2002071617 74959 11 2.49 36.24         2002116038 74959 15 40.35 64.59         2002173413 74959 21 150.88 44.96         2002239965 74959 27 194.04 32.56         2002247400 74959 28 193.16 38.1 16 47 male NO 2001302006 74965 10 17.94 51.54         2002025482 74965 13 171.21 127.42         2002059579 74965 16 333.47 238.26         2002083197 74965 19 386.43 266.42         2002140397 74965 25 401.4 275.64 17 29 male NO 2001302051 74966 2 0.93 0.93         2002037412 74966 6 0.93 3.9         2002083093 74966 11 2.17 26.96         2002128004 74966 15 2.35 59.39 18 59 male YES 2001302104 74972 6 0.93 6.72         2002025853 74972 9 0.93 3.06         2002048964 74972 11 0.93 5.94         2002071517 74972 14 118.25 24.53         2002105336 74972 17 640.9 32.06         2002138710 74972 20 940.68 32.43         2002161517 74972 23 938.8 9.9264         2002172344 74972 24 840.87 15.136         2002206637 74972 27 936.13 15.8576         2002212014 74972 28 704.64 15.5848 19 29 male NO 2001312176 74976 7 0.96 1.36         2002024985 74976 9 0.93 10.36         2002059778 74976 12 1.54 60.06         2002083023 74976 15 5.75 173.2         2002127421 74976 19 14.51 264.95 20 64 male NO 2001312184 74977 3 0.93 1.98         2002024934 74977 5 0.93 29.85         2002048795 74977 7 44.59 143.65         2002060346 74977 9 357 283.46         2002093320 74977 12 920.51 356.17         2002127291 74977 15 914.47 307.65         2002173315 74977 20 1172.36 107.26         2002206672 74977 23 999.36 90.26         2002247458 74977 27 641.88 86.3 21 55 male NO 2002014486 75021 7 0.96 7.76         2002059168 75021 11 1.46 10.33         2002083123 75021 14 1.99 14.39         2002138386 75021 19 136.11 107.79         2002148386 75021 20 151.39 88.49         2002183964 75021 24 168.56 71.72 22 57 female NO 2002048799 75024 5 0.94 0.93         2002071864 75024 8 1.46 9.03         2002115816 75024 12 4.87 23.02         2002128125 75024 13 0.93 8.05         2002140291 75024 15 7.5 23.9         2002150771 75024 16 6.32 9.6448         2002162072 75024 17 20.44 11.1848         2002172124 75024 18 46.43 12.5752         2002185056 75024 19 64.48 13.6136         2002196373 75024 20 95.51 11.6952         2002206593 75024 21 112.44 15.8928         2002218177 75024 22 98.31 13.0592         2002229264 75024 23 110.88 12.496         2002252541 75024 26 101.86 12.0296         2002276138 75024 28 92.73 9.7768 23 43 female YES 2002014480 75025 3 0.93 4.83         2002061352 75025 8 1.03 4.36         2002093995 75025 11 0.93 4.86         2002127609 75025 14 21.46 46.51         2002149871 75025 16 216.1 165.02         2002183780 75025 20 623.16 98.78 24 36 male YES 2002025485 75054 4 0.93 9.72         2002037670 75054 5 0.93 26.65         2002059680 75054 7 32.55 125.39         2002072539 75054 9 179.46 280.55         2002116856 75054 13 316.22 292.07         2002140612 75054 16 569.92 184.53 25 42 male YES 2002025637 75062 6 0.93 4.86         2002059116 75062 9 0.93 4.6         2002072520 75062 11 0.93 13.72         2002093817 75062 13 10.87 56.42         2002127491 75062 16 173.85 186.24 26 64 male NO 2002025704 75063 3 0.93 2.42         2002036781 75063 4 0.93 0.93         2002059307 75063 6 0.93 7.35         2002093337 75063 10 0.93 15.56         2002149720 75063 15 1 9.37 27 51 male YES 2002025573 75064 5 1.18 5.86         2002050025 75064 8 1 6.89         2002083137 75064 11 0.94 40.17         2002127652 75064 15 151.37 214.94         2002151216 75064 18 235.79 117.63         2002207684 75064 23 200.61 98.39         2002256162 75064 28 268.33 64.29 28 51 male NO 2001313503 75067 7 3.95 23.07         2002025616 75067 9 1.57 29.21         2002059132 75067 12 64.59 85.86         2002104233 75067 17 489.38 112.11         2002149377 75067 21 158.84 155.96 29 63 female NO 2002025748 75070 5 0.93 0.93         2002059563 75070 8 0.93 4.74         2002071702 75070 10 1.69 39.62         2002093416 75070 12 25.16 100.78         2002140375 75070 17 653.13 282.11 30 77 male YES 2002036834 75091 7 1.74 6.88         2002059179 75091 9 0.93 13.38         2002061322 75091 10 1.07 14.8         2002082791 75091 12 2.52 32.63         2002104575 75091 14 4.07 47.88         2002138576 75091 17 23.38 95.77 31 57 male YES 2002048509 75131 4 0.93 2.46         2002072082 75131 7 2.45 15.9         2002093308 75131 9 1.41 8.07         2002105687 75131 10 2.48 21.97         2002116802 75131 11 3.23 29.59         2002127853 75131 12 9.63 26.04         2002138971 75131 13 15.94 35.01         2002150964 75131 15 22.64 21.24         2002184149 75131 18 50.6 30.84         2002229495 75131 22 167.98 57.87         2002240640 75131 24 213.22 53.94 32 68 male YES 2002048842 75138 4 2.41 4.26         2002061329 75138 6 0.93 10.52         2002083162 75138 8 3.28 23.33         2002093836 75138 9 4.19 34.83         2002127457 75138 12 39.23 69.07         2002161407 75138 16 126.36 43.64         2002195400 75138 19 209.24 39.59         2002230157 75138 23 157.06 28.29         2002288414 75138 28 229.09 8.66 Detection of antibodies against SARS-CoV-2 In total, 217 blood specimens were obtained from 32 patients (6.8 blood specimens per patient on average; supplementary materials). A quantum dot immunofluorescence assay was used to semi-quantitatively detect IgM and IgG antibodies. The anti-SARS-CoV-2 IgG and IgM kits were manufactured by Chongqing Xinsaiya Biotechnology Company in Chongqing, China. Assays were performed according to the manufacturer’s detailed instructions. Briefly, serum collected from patients was incubated at 56°C for 30 min, and then, 80 μl of the diluted serum was added to the well dented on the test chip and was incubated at room temperature for 10 min. During the process, IgM or IgG antibodies in the serum sample reacted with quantum dot nanocrystal-conjugated secondary antibodies and purified recombinant SARS-CoV-2 spike (S) protein, respectively, which were both coated on a cellulose nitrate membrane. Subsequently, the immunofluorescence signal strength of the sample was analysed by a quantum dot fluorescence detector, which emitted a wavelength of 610 nm and excited a wavelength of 365 nm. The quantitative results were expressed in relative vitality units (RU/ml) according to the calibration curve. A value ≥10 RU/mL was considered to be a positive result. All serum samples were tested in triplicate, and the average of all three relative vitality units was used as the final test result. Patterns of anti-SARS-CoV-2 IgG and IgM antibodies As shown in Figure 1, anti-SARS-CoV-2 S-specific IgG and IgM antibodies were not detectable in the very early days of infection (from day 0 to day 3). Anti-SARS-CoV-2 S-specific IgM antibodies were detectable from day 4 onward; the IgM antibody titres increased over time, peaking at approximately day 20, and then began to decline. The positivity rate of IgM antibody was only 60%, with a marked reduction in antibody levels 4 weeks after onset of illness. Anti-SARS-CoV-2 S-specific IgG antibodies were identifiable from day 7 onwards, peaking at approximately day 25, as shown in Figure 1(A). Serum IgG antibodies were still maintained at a high level after 4 weeks of infection. Figure 1(B) shows a typical IgG and IgM antibody response in a 65-year-old woman with COVID-19 (supplementary materials, Table 1). It is widely accepted that the IgM antibody response provides early-stage defence during viral infections prior to the development of the class-switched, high-affinity IgG response for long-term immunity and immunological memory [5]. Figure 1. Patterns of anti-SARS-CoV-2 S antibody response in patients with COVID-19. (A) IgG and IgM antibody response patterns in serum samples of all 32 patients with confirmed COVID-19; s, a significant difference by non-parametric repeated measures ANOVA; (B) IgG and IgM antibody response in a 65-year-old woman with severe COVID-19; (C) The difference in anti-SARS-CoV-2 IgG antibody response between severe cases and mild cases; (D) The difference in anti-SARS-CoV-2 IgM antibody response between severe cases and mild cases. (E) Numbers of blood samples collected during the study period. For statistical analyses, the Mann-Whitney U test was performed for continuous variables. Statistical testing could not be performed on days 22, 24, 25 and 27 due to the very small number of available samples. *p < 0.05 was considered statistically significant. Comparison of antibody response between mild cases and severe cases We further compared the difference in antibody detectability between mild cases and severe cases of COVID-19. As shown in Figure 1(C), serum IgG antibody levels were not significantly correlated with clinical severity in the early stage of infection. However, the difference in IgG antibody levels between mild cases and severe cases from day 15 onward was found to be statistically significant (day 15 (N = 17), day 20 (N = 6) and day 21 (N = 11), all p < 0.05). Severe cases of COVID-19 tended to have a more vigorous IgG response against SARS-CoV-2 compared with mild cases. Notably, some patients with mild disease had a robust IgG antibody response from 9 days after symptom onset, while a few mild cases did not generate adequate IgG antibodies (approximately 21.43%). Our results also showed that mild cases tended to develop faster peak anti-SARS-CoV-2 S-specific IgM responses (approximately 17 days after symptom onset) compared with severe cases (approximately 21 days after symptom onset). It is also worth noting that IgM antibodies disappeared 4 weeks after symptom onset both in mild cases and severe cases (Figure 1(D)). In summary, we observed that (1) the IgM antibody response to SARS-CoV-2 occurred earlier and peaked earlier than the IgG antibody response; (2) the IgM antibody response began to decline at week 3 of the illness, while the IgG antibody response persisted and was maintained in patients with COVID-19; and (3) severe cases of COVID-19 tended to have a more vigorous response in both IgG and IgM antibodies to COVID-19 illness. Our findings may be of significance in interpreting anti-SARS-CoV-2 antibody test results and in understanding humoral immune response patterns for SARS-CoV-2 infection in current and potential future COVID-19 outbreak scenarios. Importantly, the timing of IgM and IgG antibody occurrence in patients varies greatly, and this variation in timing may be associated with age as well as comorbidity [6]. More care needs to be taken when using levels of anti-SARS-CoV-2 antibodies to make a clinical diagnosis of COVID-19 or determine discharge criteria.