COVID-19 prevalence surveys might be unreliable given how quickly these infection-fighting proteins fade

COVID-19 prevalence surveys that detect levels of antibodies to determine whether people were infected in the past might be unreliable given how quickly these infection-fighting proteins fade, a US government study said Wednesday.

Whether the decline in antibodies increases the risk of reinfection remains uncertain.

It is thought likely that if re-exposed to the virus, most people’s immune systems will produce new antibodies, as well as immune cells that target and destroy cells that have been infected.

But the new study highlights an important limitation of “seroprevalence studies” that are being carried out around the world in an attempt to better understand the true number of people who have been infected by a virus that often causes no symptoms.

The US Centers for Disease Control and Prevention collected blood serum samples from frontline health care personnel at 13 hospitals between April 3- June 19 2020, and followed up approximately two months later.

Overall, 194 of 3,248 participants (six percent) had detectable antibodies to SARS-CoV-2 the first time they were tested.

Among the participants who tested positive, 156 agreed to re-test, between 50-91 days later.

In this group, 94 percent saw a decline in their antibody levels, with 28 percent not producing enough COVID-19 antibodies to meet a clear threshold that would suggest prior infection.

“These results suggest that serology testing at a single time point is likely to underestimate the number of persons with previous SARS-CoV-2 infection, and a negative serologic test result might not reliably exclude prior infection,” wrote the authors.

Participants who had reported symptoms of disease had higher levels of antibodies initially than those who had asymptomatic infection, and were less likely to fall below the “positive” threshold.

Most people infected with the virus develop antibodies over the next several weeks, but people with milder or asymptomatic infection develop fewer antibodies.

Apart from helping to quantify how many infections seroprevalence studies may be missing, the results “challenge the notion of using serologic testing results at an individual level to designate previous SARS-CoV-2 infection,” said the authors.

Finally, the study also shows that the window is limited for collecting potentially effective “convalescent plasma” from the blood of patients who have fought off COVID-19, they added.

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Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-related coronavirus disease (COVID-19) is a respiratory transmissible disease that may cause critical illness to death [1,2]. Various control measures against SARS-CoV-2 were implemented in Wuhan, the first city hit by the coronavirus. After months of endeavour, viral transmission was largely contained [3].

However, sporadically infected cases and asymptomatic carriers were still detected. Hence, Wuhan launched a population-scale, massive SARS-CoV-2 testing campaign for detecting viral nucleic acid and antibodies in residents to further prevent viral transmission, screen out infected patients who were in the incubation period or were asymptomatic virus carriers, and map the epidemiological serodistribution of this infectious disease in the epicentre.

Wuchang District, one of the 13 administrative divisions in Wuhan, is located in the central urban area and is adjacent to the Yangtze River.

According to the Wuhan Statistical Yearbook 2018, the total population of Wuchang District is 1.28 million, accounting for 11.7% of the Wuhan population. The present study describes the screening results of 61 437 community members in Wuchang District, Wuhan.

Results

Among 61 437 community members included in the study, 30 032 (48.88%) were male. The age range is from several months to 101 years old and the median age was 48 (interquartile range (IQR) 32–64) years. The sex and age distribution is depicted in Fig. 1.

As shown in Tables 1 and 1470 people tested positive for at least one SARS-CoV-2 antibody, accounting for 2.39% (95% CI 2.27–2.52) of the whole tested population. Among these seropositive people, 616 (41.88%) were male.

The subgroup data showed that 324 (0.53%, 95% CI 0.47–0.59) and 1200 (1.95%, 95% CI 1.85–2.07) residents were positive for IgM and IgG antibodies, respectively. A 3.7-fold positivity rate was observed in IgG groups compared with IgM groups.Table 1Information of antibodies positive populations

Further, a discrepancy in seroprevalence was observed in age and sex subgroups (Table 1 and Fig. 2). The positivity rates of IgM, IgG or their combination gradually increased with the increase in age, with the peak in the subset of people aged 70–79 years (Fig. 2).

The lowest positivity rates (IgM, 0.12%; IgG, 0.75%; and IgM and IgG, 0.86%) were observed in groups of people aged 20–29 years. When considering subgroups by sex, the MFR, was 0.75 in IgM- and IgG-seropositive populations (Table 1 and Fig. 2), suggesting a significantly higher frequency in women than in men.

In age subgroups, a trend of the female positivity rate higher than the male positivity rate (MFR <1) was captured in most age groups, except for the ≤19 subgroups. Additionally, the MFR values decreased when age increased, and the lowest MFR (0.18) was observed in 90+ age subgroup, indicating an increased probability of the female antibody carrier with the increase in age.

The real-time RT-PCR identified eight (0.013%, 95% CI 0.006–0.026) asymptomatic virus carriers. Five of them were male and three were female. The age range was 45–77 years. Among them, four were negative for both IgM and IgG antibodies, and four were positive for IgG antibody only.

Discussion

A total of 1470 individuals tested positive for at least one SARS-CoV-2 IgM or IgG antibody, with an overall seropositivity rate of 2.39%. Only healthy members were included in the campaign, while confirmed patients with COVID-19, people under quarantine and hospitalized inpatients were excluded to reduce possible cross-infection.

This group of people was more susceptible to SARS-CoV-2 infection than other citizens, thus decreasing the seropositivity rates. In addition, most non-local workers who left Wuhan before city shutdown because of the Chinese New Year in January returned to Wuhan by the date of sampling. However, the universities still remained closed, and hence a majority of college students were not included in the study.

Wuchang District had many colleges; therefore, exclusion of this population had a significant effect on the positive rates. The lack of college students, who were mainly around 20 years old, might have led to elevated seropositivity rates because the 10–19 and 20–29 age groups had the lowest antibody positivity rates (Fig. 2 and Table 1).

Furthermore, the participants were voluntarily recruited, and hence it was reasonable to speculate that a small proportion of mobility-impaired individuals were reluctant to participate, although they performed fewer activities during the outbreak and had fewer chances to be infected.

The present study was conducted in Wuchang District, while data in different districts in Wuhan hit by COVID-19 might vary. However, the observed rate was comparable with that reported in other similar studies. A recent study detected 17 368 individuals from different geographic regions in China, including 1993 residents from different Wuhan sub-cohorts.

It suggested a seropositivity rate of 3.8%, 3.2% and 3.8% in healthcare workers, their family members and their staff members, respectively, from hotels designated for the accommodation of healthcare workers during city lockdown in Wuhan between 30 March and 10 April 2020 [5].

As the exposure of healthcare workers and their close contacts to SARS-CoV-2 was relatively higher than that of most other citizens, it was reasonable that the seropositivity from these subgroups was higher than that from populations with massive testing. Another study testing 452 asymptomatic Hong Kong residents evacuated from Hubei province in early March 2020 indicated a 3.76% (17/452) seropositivity rate [6]. All these seropositivity rates indicated that the prevalence of the population carrying the antibody in Wuhan was low.

Among 61437 community members included in the study, eight (0.013%), 324 (0.53%) and 1200 (1.95%) individuals were detected positive for nucleic acid, IgM and IgG, respectively. As IgM is regarded as the first class of immunoglobulins in response to initial exposure, the presence of IgM antibody represents an early exposure to the antigen [7].

The anti-SARS-CoV-2 IgM antibody could be detected in patients after 4 days of onset, peaking at 2–3 weeks after the onset of symptoms before its level started to decline [8,9].

However, IgM positivity alone may not be a good diagnostic indicator because not all of the people develop a detectable IgM antibody [8]. In addition, the IgM antibody may still be detectable after several months, although it is considered as an ‘early infection’.

Most of the recent studies showed detectable SARS-CoV-2 anti-IgM antibodies after 1 to 2 months [10,11]. IgG represents the most robust and long-duration antibody against the virus [12]. The positivity of the IgG antibody indicates a ‘past infection’ of the virus. Wuhan reported approximately 50 000 patients with confirmed symptomatic COVID-19.

The result suggested that Wuhan, a city with 11 million people, had proportionally 269 200 (2.39%) asymptomatic individuals infected with SARS-CoV-2 in the last few months, besides confirmed patients. A more accurate seropositivity rate in Wuhan may need further analysis, including multidistrict testing results.

Remarkably, the present study detected more female carriers of asymptomatic antibodies compared with male carriers in most age subgroups and a reverse correlation trend of MFR with the increase in age. Another study in Wuhan also reported similar findings [13].

Indeed, it was suggested that SARS-CoV-2 affected women less than men due to different innate immunity, steroid hormones and factors related to sex chromosomes [14]. However, this female over male trend in asymptomatic carriers was not captured in other areas and countries, such as South Korea, Thailand, Iran, Spain and California in the USA [15, 16, 17, 18].

Whether this trend is observed only in Wuhan or can be observed in other parts of China or other countries too needs further investigation.

This study had several limitations. First, several groups of people were not included in the study, which might have had different impacts on detecting real seropositivity. Second, the rates were affected by the quality of the kit. As mentioned in the kit specifications, the sensitivity and specificity of this commercially available IgM/G detection kit were 91.54% (86.87–94.65) and 97.02% (94.74–98.33), respectively.

The sensitivities were comparable with those of other brands [19]. These intrinsic shortcomings of the rapid immunochromatographic kit might inevitably cause false-positive and false-negative results. Third, massive tests were conducted within 12 days, and hence the possibility of higher false-positive or false-negative rate due to the labour-intensive work was unavoidable.

Fourth, a rapid and ready-to-use method, the immune colloidal gold technique, was adopted for the screening test because a large number of samples were needed for handling and the technicians had some limitations. The test provided only a qualitative positive or negative result.

A more quantitative result may be obtained using the chemiluminescence enzyme immunoassay.
In summary, the majority of the residents in Wuhan are still immunologically naive to SARS-CoV-2, far from herd immunity. Although Wuhan re-opened on 8 April 2020, proper control measures, such as frequent hand hygiene, wearing face masks and keeping proper social distance, are necessary before effective vaccines or antiviral drugs are available.

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