RISK FACTORS and

IMMUNE RESPONSES OF
SARS COV-2
REINFECTION
Lisyani Budipradigda-Suromo
Bagian Patologi Klinik Fakultas Kedokteran Undip
 VIRAL INFECTIONS
Some viral infections result in lifelong immunity

Other viruses can result in repeated reinfections

throughout life
e.g. influenza, seasonal coronaviruses:
- reinfections are very common
- serum antibody (Ab). persist for only months - a
few years
Source : Cohen JI, Burbelo PD. Clin Infec Dis. 2020 .
Available from : https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7799323/
A study of 4 reinfection patients in Qatar :
* there were distinct viral variants between the first &
second infections (using genetic sequencing data)

A study (Roberto Bertollini) among 133,266 confirmed

SARS-CoV-2 cases:
Two out of those cases had identical viral genomes
on the first & second positive test !
► suggesting a lingering infection that never cleared

in the weeks between the two positive tests

Source : Yeager A. Repeat COVID-19 cases could offer clues about people’s immunity to the novel
coronavirus and how to vaccinate against it. 2020. Available from : https://www.the-scientist.com/news-
opinion/more-sars-cov-2-reinfections-reported-but-still-a-rare-event-68089
Chronic infection has been linked to the reemergence
of infection or relapse state

Relapse is a recurrent infection with the same type of

pathogen

Reinfection is defined as the emergence of infection

with a different species or serologic strain of pathogen

Relapse / reinfection (?) may occur in COVID-19 !

 SARS CoV-2 reinfection
The duration of protective immunity to the SARS–CoV-2
infection is unknown

Cases of reinfection with the SARS-C0V-2 have been

reported throughout the world ► the frequency ↑

After SARS CoV-2 infections :

* How common reinfection with SARS-CoV-2 is ?
* How long serum Ab & virus-specific T cells persist ?

UNCLEAR
Source : Cohen JI, Burbelo PD. Clin Infec Dis. 2020 .
Available from : https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7799323/
 A CASE REPORT OF REINFECTION
(from Hongkong)

Note : TMA = transcription-mediated amplification test

Source :Tillet RL. Genomic evidence for reinfection with SARS-CoV-2: a case study. The Lancet 2021;
21(1):52-8 Available from : https://www.thelancet.com/journals/laninf/article/PIIS1473-3099(20)30764-7/fulltext
In all four seasonal coronaviruses :
* Reinfection are common & frequently occur after about
a year
* Antibody levels spiked & dropped at regular intervals

SARS-CoV-2 infection would be the same

Other researcher :
* In other coronavirus reinfections may happen, because
the infection is only mild ► a strong immune response

was’nt elicit (with lots of long-lasting antibodies)

less convinced that SARS-CoV-2 infection will

have the same action
Source : Yeager A. Repeat COVID-19 cases could offer clues about people’s immunity to the novel coronavirus and
how to vaccinate against it. 2020 . Available from : https://www.the-scientist.com/news-opinion/more-sars-cov-2-
 Several reports of
RISK FACTORS / CAUSES OF SARS-CoV-2 REINFECTION

- Reinfection can be proved, if the viral variants from

1st & 2nd positive swabs are different
- “ it’s very unlikely that an individu will be infected with
the exact same variants a second time.”
Source : Angela Rasmussen in Yeager A, 2020

Reinfection suggests that the immune response to the

first infection was not adequate to provide protection
against reinfection !!!
►their immunity were immature
 Waning immunity ► may cause reinfection

Prolonged immunity after a first infection may not be

uniform ► possible to cause reinfection

In a healthy person on a second exposure, NK cells &

memory cells (B & T cells) may be reactivated for a
quicker respons

Drake J. Covid-19 and the science of reinfection. 2021. Available from :

https://www.forbes.com/sites/johndrake/2021/03/25/covid-19-and-the-science-of-reinfection/?sh=
Another analysis :

* Patients with reinfection were :

- relatively younger than without reinfection
- elderly with comorbidities **

* Patients with reinfection had experienced two or

fewer symptoms during the first episode of COVID-19
Symptoms of COVID-19 :
dry cough, fever, breathlessness, fatigue, diarrhea,
loss of smell and/or taste
Source : Dutta SS. Researchers identify risk factors for SARS-CoV-2 reinfection.2021. Available from :
https://www.news-medical.net/news/20210416/Researchers-identify-risk-factors-for-SARS-CoV-2-reinfection.aspx
** Alinaghi SAS, Oliaei S, Kianzad S, Afsahi AM, MohsseniPour M, Barzegary A, et al. Reinfection risk of novel
coronavirus (COVID-19): A systematic review of current evidence. World J Virol 2020; 9(5): 79-90 [**
 * Other clinical characteristics of reinfected patients
during the initial COVID-19 episode were :
- the presence of a secondary infection
- leukocyte count : higher than normal
- lymphocyte count : lower than normal (<1500/µL)
- increased blood circulation rate

* Convalescent individuals with lymphopenia history

* or having two / fewer symptoms during the initial
COVID-19 episode
at higher risk for developing SARS-CoV-2 reinfection

are independent risk predictors of SARS - CoV-2

reinfection
Source :Dutta among
SS. Researchers identify COVID-19
risk factors recovered
for SARS-CoV-2 reinfection.2021. individuals
Available from :
https://www.news-medical.net/news/20210416/Researchers-identify-risk-factors-for-SARS-CoV-2-reinfection.aspx
Patients with severe ill at the first infection,
► will be asymptomatic at the second time
 The life cycle of any virus involves :
►attachment, penetration, uncoating, replication,
assembly, release

When SARS-CoV-2 replicate ► it will relatively

mutate slowly

Available from : https://www.osfhealthcare.org/blog/how-covid-19-mutates-and-how-it-affects-vaccines /

 VIRAL MUTATION

All viruses – including SARS-CoV-2, evolve over time

A normal virus replicates / makes copies of itself,

►sometimes changes a little bit / mutated (antigenic drift)
The more opportunities a virus has to spread ► the more it
replicates ► the > opportunities it has to undergo changes

When a virus is widely circulating in a population and

causing many infections ► mutation ↑
A virus with one or more new mutations = a “variant” of
the original virus
Source :
https://www.who.int/news-room/feature-stories/detail/the-effects-of-virus-variants-on-covid-19-vaccines?gclid=Cj0KC
Qjws-OEBhCkARIsAPhOkIZ9WVpanwcNEzKTXW7kB-xvlLey4QIjf-sVS
 MUTATION OF SARS- CoV-2

- SARS-CoV-2 has an RNA genome (ssRNA), which has

a higher mutation rate than a DNA viral genome

- the genetic changes that SARS-CoV-2 has so far

accumulated, seem to be functionally inconsequential

- within an individual host who’s infected ► when the

virus replicates and makes changes / mistakes
► multiple variants will emerge !

- in the population: there are many millions of different

coronavirus cases ► different variants have emerged !
Korber et al. present evidence that there are now more SARS-CoV-2 viruses circulating in the human population
globally that have the G614 form of theSpike protein versus the D614 form that was originally identified from the first
human cases in Wuhan, China. Follow-up studies show that patients infected withG614 shed more viral nucleic acid
compared with those with D614, and G614-bearing viruses show significantly higher infectious titers in vitro than
their D614 counterparts. Korber et al., 2020, Cell182, 812–827August 20, 2020 Published by Elsevier
Inc.https://doi.org/10.1016/j.cell.2020.06.043ll
SARS-CoV-2 uses RNA dependent RNA polymerase
(RdRp) in its replication :
► may produce progenies with slightly mutated
genes

The presence of mutations in SARS-CoV-2 genome :

► particularly in ORF1ab, ORF8, and N genes

Mutation at primer and /or probe sites

► cause false-negative result of RT-PCR method
CORONA VIRUS GENOME AND MUTATION

VARIANTS of
SARS-CoV-2 e.g :

B.1.1.7 (Brazil)
B.1.3.5.1 (South
Afrika)
P.1 (Japan)

Available from :
https://www.ingenetix.com/en/new-sars-cov2-mutations-variant-of-concer
PREVENTIONS TO NEW VARIANTS OF THE SARS –CoV-
2:

► stopping the spread of the virus sources, includes :

* frequent hand washing

* wearing a mask
* physical distancing,
* good ventilation
* avoiding crowded places or closed settings

► reducing the amount of viral transmission ► also

reducing the opportunities of viral mutation
 IMMUNITY TO SARS-CoV-2 INFECTION

Functional immunity to SARS-CoV-2 is quite

individualistic
not everyone who is infected with SARS-CoV-2 will
develop antibodies ►vulnerable to a second exposure

Prolonged immunity after a first infection may not be

uniform
Attention should be paid at the longevity of
antibodies
after infection
Immune durability should be the focus of reinfection
 Antibodies to SARS-CoV-2 can target many
of its encoded proteins, including structural
and nonstructural antigens

The SARS-CoV-2 coronavirus molecule.

Available from : new-coronavirus-variant-what-is-the-spike-protein-and-why-are-mutations-on-it-important-152463
https://theconversation.com/new-coronavirus-variant-what-is-the-spike-protein-and-why-are-mutations-
on-it-important-152463
Two structural proteins have been used as target antigens
for serological assays :
1. the abundant nucleoprotein (NP), which is found inside
the virus/ inside infected cells
►because of : the biological function of NP & it is
shielded from antibodies by virus cellular membranes
► NP Abs can’t directly neutralize SARS-CoV-2

2. the spike protein = a large trimeric glycoprotein

that contains the receptor binding domain (RBD)
► used by the virus to dock to its receptor ACE-2
& for cellular membranes fusion
► the main / the only potentially neutralizing antibody,
relatively stable for at least a period of ± 5 months
Source : Wajnberg F, Amanat F, Firpo A , Altman DR, Baily MJ, MansourM. Robust neutralizing antibodies to SARS-CoV-2
infection persist for months. Science 2020; 370(6521):1227-1230. doi: 10.1126/science.abd7728. Epub 2020 Oct 28.
 Fig. 7 Representation of possible SARS-CoV-2 antibody binding sites

https://www.imd-berlin.de/en/subject-information/diagnostics-information/indication-and-interpretation-of-
sars-cov-2-antibody-diagnostics.html
The S protein consists of two subunits, S1 which
contains the receptor binding domain (RBD) & S2

Neutralizing Abs. reached a plateau 2 weeks post-

symptom onset & then declined in the majority of
inpatients (does not induce a prolonged neutralizing Ab
response),
► undetectable in 56% of asymptomatic patients
Brochot E, Demey B, Tauze A, Belouzard S, Dubuisson J, Schmit JL et al . Anti-spike, anti-nucleocapsid and neutralizing
antibodies in SARS-CoV-2 inpatients and asymptomatic individuals. Front Microbiol 2020 https://doi.org/10 3389/fmicb 2020
584251. Available from : https://www.frontiersin.org/articles/10.3389/fmicb.2020.584251/full
 Most patients infected with SARS-CoV-2 showed
detectable antibody response between 10 -14 days
after symptom onset

In some mild cases the level of Ab can be low (or

require a long time to response) or undetectable

Long et al. have reported :

•The IgG specific levels of SARS-CoV-2-infected
individuals had an ~70% reduction during the early
convalescent phase
• 40% of asymptomatic & 12.9% of symptomatic patients
became IgG seronegative
Source : Wang z, Yang X , Zhong J, Zhou Y, Tang Z, Zhou H et al. Exposure to SARS-CoV-2 generate T cell memory in
the absence of detectable viral infection. Nature Communications 2021. https://dot.org/10.1038/s41467-021-22036-z
Fig. 1. A schematic representation of the SARS-CoV-2 immune response following infection. Seroconversion
occurs from approximately 10 days after symptom onset with the exact timing of IgM (green line) and IgG
(red line; high titre, solid line; low titre, dashed line) appearance presently unclear, but with a suggestion
that the IgM occurs at the time of, and overlapping with, the IgG response. The IgG antibody titres rise from
day 10 onwards to reach a peak whose height is likely to be influenced, on a case by case basis, by disease
severity and virus load. Seropositive status for those that seroconvert is detectable from 3 to 4 weeks from
symptom onset. The level of antibody protection from reinfection (black dotted line), the duration of the total
humoral immune response above this level, and the rate of decline from mild or severe infection induced
antibodies is not known for SARS-CoV-2. Similarly, the proportion of infected individuals that do not mount
a protective immune response (blue line) is not known.
Available from : https://pubmed.ncbi.nlm.nih.gov/32430094/#&gid=article-figures&pid=fig-1-uid-0
Another study showed that :

Neutralizing titers reached a peak at 23 days after onset

of symptoms and then declined (using ELISA method)

Persons with more severe disease had higher levels of

peak neutralizing titers & still be detected ± 2 to 3 months
after onset of symptoms

Those who were asymptomatic / had mild symptoms had

lower levels of peak antibody titers and & some fell below
the level of detection at 2 months after infection
Source : Source : Cohen JI, Burbelo PD. Clin Infec Dis. 2020 .
Available from : https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7799323/
Fig 3. Schematic showing the scale of IgG/IgM/IgA/Neutralising Ab response over time from
disease onset.
Note that the y-axis is illustrative only and therefore no scale is given: this figure gives an indicative overview of findings
from all included studies with relative peaks and decline indicate

Source : Post N, Eddy D, Huntley C, van Schalkwyk MCI, Shrotri M, Leeman D.et.al Antibody response to SARS-CoV-2
infection in humans: A systematic review..Plos One 2020. https://doi.org/10.1371/journal.pone.0244126
 Reports of other study
Antibodies against spike & RBD declined
moderately over 8 month

Spike IgA was still present in the large majority

of subjects at 6 - 8 months after infection
Source : Dan JM, Mateus J, Kato Y, Hastie KM, Yu ED, Faliti CE. Immunological memory to SARS-CoV-2
assessed for up to 8 months after infection. Science 05 Feb 2021: Vol. 371, Issue 6529, eabf4063 DOI:
10.1126/science.abf4063

IgA serum : may be detected in 3- 6 days & reach the peak

level at 3 weeks after symptom onset :
- anti-RBD IgA is detected earlier than anti-RBD IgG
- was > potent than IgG in neutralizing SARS-CoV-2
- ►highlight the potential role of IgA during early
SARS- CoV-2 infection

IgA saliva : the concentrations were higher than IgG &

persisted for several more weeks
Source: Sterlin D, Mathian A, Miyara M, Mohr A, Anna F, Laetitia.Sci Transl Med 20 Jan 2021: Vol. 13, Issue 577,
eabd2223 DOI: 10.1126/scitranslmed.abd2223
 NOTES

* In certain mutations in the RNA genome of SARS-CoV-2 :

► changes in epitope structure and / or characteristics

► production of progenies with new epitopes

* In reinfection :

► the antibodies produced in the primary infection may

not be able to recognize the epitope of the new virus
► host defense towards infection will start from the
beginning (from innate to adaptive immune activation)
► eventually will result in either a recovery or reinfection
 IMMUNOLOGICAL MEMORY

* is the basis for durable protective immunity after

infections (or vaccinations )
* helps to determine protection against reinfection,
disease risk & vaccine efficacy

* The duration of immune memory after SARS-CoV-2

infection / COVID-19 is unclear !

Result of several studies : ≥ 90% of subjects retained

immune memory for > than 5 months – 8 months
after infection (average 6 months)
IN GENERAL:
The presence of memory T & B cells :
► provides steadfast response & a high level of
protection to the host
► leads to continuous /prolong protective immunity
(may up to several years)

The roles of memory lymphocytes

► provide prolonged antiviral protection
► the decreased quantity & quality of these
cells will ↓ the host immune responses against
reintroduction of previously encountered
pathogenic microbes
Nainu F et al. SARS-CoV-2 reinfection and implications for vaccine development. HUMAN VACCINES &
IMMUNOTHERAPEUTICS 2020; 16(12) :3061–73 . Available
from:https://doi.org/10.1080/21645515.2020.1830683
 MEMORY LYMPHOCYTES IN SARS-CoV-2
INFECTION
► memory B cells, CD4 +T cells & CD 8 + cells
Memory B cells against SARS-CoV-2 spike ↑ between
1 - 8 months after infection

70% of individuals possessed detectable CD8+ T cell

memory at 1 month after infection, but ↓ to 50% by 6 - 8
months after infection

93 % of subjects had CD4+ T cell memory at 1 month

after infection, & 92% remained high at 6 - 8 months
SARS-CoV-2 spike-specific memory CD4+ T cells with the
specialized capacity to help B cells (T follicular helper / TFH)
cells were also maintained
Source : Dan JM, Mateus J, Kato Y, Hastie KM, Yu ED, Faliti CE. Immunological memory to SARS-CoV-2
assessed for up to 8 months after infection. Science 05 Feb 2021:
 Immunological memory to SARS-CoV-2
assessed for up to 8 months after infection
Dan JM, Mateus J, Kato Y, Hastie KM, Yu ED, et al. Science 2021:;Vol. 371, Issue 6529, eabf4063 DOI:
10.1126/science.abf4063

Fig. 1 | T cells and B cells in immunity to SARS-CoV-2 a. Infection with SARS-CoV-2 leads to activation of innate
immunity and dendritic cells (DCs), which will drive the induction of virus-specific T cell and B cell responses. Little
is currently known concerning the memory response to SARS-CoV-2, but this will be important for developing an
effective vaccine. b | A predicted time-course of adaptive immunity to SARS-CoV-2. CTL, cytotoxic T lymphocyte;
TFH, T follicular helper cell; TH, T helper cell; Treg, regulatory T cell.
Available from : https://www.nature.com/articles/s41577-020-00436-4
In COVID-19 :
* lymphopenia may lead to suboptimal production
of an anti-SARS-CoV-2 nAbs and or reduced
activities of CD4 Th cells & CD8 T CTL cells

* it is possible that recovered COVID-19 patients

with lymphopenia history :
► increased ↑ vulnerability to SARS-CoV-2
reinfection
Nainu F et al. SARS-CoV-2 reinfection and implications for vaccine development. HUMAN VACCINES &
IMMUNOTHERAPEUTICS 2020; 16 (12): 3061–73
Avaialable from : https://doi.org/10.1080/21645515.2020.1830683

Virus specific CD4+ T cell numbers were associated

with the production of RBD IgG of SARS-CoV-2
MEMORY B CELLS RESPONSES
* IgG was the dominant isotype
* Ig A in a minor population
* IgM appeared to be short-lived

Sokolowska M, Lukasik ZM, Agache I, Akdis CA, Akdis D, Akdis M et.al. EAACI
2020. Available from : https://onlinelibrary.wiley.com/doi/full/10.1111/all.14462
B cells play an important role in clearance of SARS-
CoV-2 :
► mostly through the production of nAbs
► but, the duration of this protection remains
unknown

? a questioned whether prolonged immune

protection against SARS-CoV-2 truly prevails
Nainu F et al. SARS-CoV-2 reinfection and implications for vaccine development. HUMAN VACCINES &
IMMUNOTHERAPEUTICS 2020;16(12):3061–73
Available from : https://doi.org/10.1080/21645515.2020.1830683
 THE KINETICS OF IMMUNE MEMORY CELLS

Each of different types of immune memory had distinct

kinetics :
► a complex interrelationships between the
abundance of T cell, B cell, and Ab immune
memory over time

Spike IgG, RBD IgG & neutralizing antibody titers

exhibited similar kinetics

! heterogeneity in memory to SARS-CoV-2 was observed

Circulating Ab titers could not be used as a predictive of T cell

memory
Figure 1. Antibody
response in SARS-CoV-2
infected inpatients. (A)
Kinetics of anti-S1, anti-
S2, anti-RBD, anti-N and
NAb detection in 30
COVID-19 inpatients post-
symptom onset. (B)
Evolution of the anti-S1,
anti-S2, anti-RBD, and
anti-N antibody levels
during the first month
post-symptom onset.

Brochot E et.al 2020

https://www.frontiersin.org/articles/10.3
389/fmicb.2020.584251/full
SUMMARY
Risk factors of SARS CoV-2 infections :
- the mutant virus
- inadequate / mild immune responses(primary mild infection)
- lower lymphocyte count
- few symptoms during the first infection
- younger age patients / elderly with comorbidities
- higher leukocyte count
- secondary infection
- increased blood circulation rate
Immunity to SARS-CoV-2 infection is not just antibodies!
► the duration of host protection by nAbs remains unknown
► a prolonged neutralizing antibody response is individualistic

The immunological memory (T & B cells) will protect

against severe disease if reinfection occur
THANK
YOU