SARSLIVA: utility of saliva in COVID-19 diagnosis - a household study

In mitigating the SARS-CoV-2 pandemic, governments have to weigh the public
health benefit of interventions such as closure of schools, restaurants and
theatres against the significant societal and economic disruption they impose.
After opening up schools and public life, and restarting the economy, close
monitoring of spread of the virus is a crucial tool for success of tracing the
virus and SARS-CoV-2 spreading in the population. Recently, tracking the virus
in saliva by molecular diagnostics was shown to be at least as sensitive as the
most broadly used method of viral detection in nasopharyngeal (NP) swabs.
Saliva sampling would be a very attractive way for large scale monitoring of
SARS-CoV-2 spreading, mostly because it gives the possibility of self-sampling
in the home situation. However, saliva sampling has not yet been validated for
asymptomatic and pre-symptomatic SARS-CoV-2 infected individuals. If we can use
saliva for early detection, containment of viral spread is made easier. Saliva
is also not yet validated at lower viral density in the track of SARS-CoV-2
infection. This would allow us to see whether persons, who we now know may
remain symptomatic for a prolonged period of time (weeks to months), continue
to shed the virus. In addition to SARS-CoV-2 detection, saliva may also be a
good specimen for detecting emerging mucosal IgA, IgM and IgG antibodies
against SARS-CoV-2.

Much is still unknown about host and virus factors influencing (long-term)
immunity after mild or asymptomatic SARS-CoV-2 infection.
Data on long-term persistence of antibodies after COVID-19 is limited and in
most cases follow-up is restricted to 6 months after infection. There is a need
for in-depth studies investigating factors dictating long-term dynamics of
SARS-CoV-2 antibodies after mild or asymptomatic COVID-19.
Data regarding susceptibility for re-infection after COVID-19 are limited,
especially in mild or asymptomatic cases. In the light of emerging new
SARS-CoV-2 variants, antibody cross-reactivity to multiple new SARS-CoV-2
variants is an important topic for further studies.
In January 2021, the COVID-19 vaccination campaign has started in the
Netherlands. In-depth data regarding topics such as antibody response in
individuals with previous mild or asymptomatic COVID-19, long-term persistence
of vaccine antibody response or cross-reactivity of vaccine induced antibodies
to emerging new SARS-CoV-2 variants is essential to address these issues. .
Data on detection of long-term antibody response in saliva after mild or
asymptomatic SARS-CoV-2 infection or vaccination is lacking. Since collection
of saliva is easy and non-invasive, when proven a reliable method for antibody
detection it will be a very useful tool for population screening for
seroprevalence of SARS-CoV-2 infection or measuring vaccination response.
Long-term impact of SARS-CoV-2 infection on presence and abundances of common
respiratory bacterial pathogens such as Streptococcus pneumoniae, Haemophilus
influenzae, Neisseria. meningitidis and Streptococcus pyogenes in the upper
airways. H. influenzae is of particular interest due to the increased incidence
of invasive disease in the Netherlands during the COVID-19 pandemic
Finally, data on long-term sequela of COVID-19 (*Long COVID*) are limited,
especially after mild or asymptomatic COVID-19.

Substudy on reinfections
Reinfections with SARS-CoV-2 occur frequently due to declining immunity after
previous infection and/or vaccination and the emergence of new virus variants
that partially circumvent existing immunity. Therefore, there remains a risk of
new corona waves in the current phase of the Covid-19 pandemic, especially
during the winter season. A better understanding of the longevity of protective
immunity and the risk of reinfections, including their impact on individual
health, allows improved preparedness and can inform optimally targeted measures
to limit the impact on individuals and society. For this reason, this project
mainly focuses on research into identifying risk factors for reinfections,
including the role of waning immunity, and on understanding how these
reinfections affect our health and daily functioning in the short and long
term.

Primary objective
1. The sensitivity of saliva collection for SARS-CoV-2 tracing, with home
self-sampling, storage and transport to the laboratory.
2. The sensitivity of saliva collection by self-sampling for SARS-CoV-2 tracing
in a- and pre-symptomatic household members of COVID-19 patients
3. The emergence of IgA, IgM and IgG anti-SARS-CoV-2 specific antibodies in
saliva and serum

FUP:
4. Long-term dynamics of neutralizing and non-neutralizing antibodies after
mild or asymptomatic SARS-CoV-2 infection in both serum and saliva.
5. Dynamics of SARS-CoV-2 neutralizing and non-neutralizing antibodies in
saliva after SARS-CoV-2 vaccination in participants with and without previous
SARS-CoV-2 infection.
6. Determine the sensitivity and specificity of saliva as a specimen as
compared to serum for detection of the analyses mentioned above.
7. Long-term sequelae of SARS-CoV-2 infection, such as quality of life and
clinical symptoms (*long COVID*).

Reinfection substudy:
8. to determine the incidence rate of reinfections with SARS-CoV-2.
9. To investigate risk factors (demographic, clinical, virological,
immunological) for SARS-CoV-2 reinfections.
10. To understand which factors determine the longevity and breadth of
protective SARS-CoV-2-specific humoral immunity.
11. To determine the health impact of SARS-CoV-2 reinfections or their sequelae.

Secondary objective
1. Evaluation of tracing other respiratory viruses in saliva.
2. Evaluation of co-infection with respiratory bacterial pathogens in NP, OP
swabs and in saliva by conventional (culture-based) and by molecular
diagnostics (PCR).

FUP:
3. Dynamics of SARS-CoV-2 neutralizing and non-neutralizing antibodies in both
serum and saliva in study participants with SARS-CoV-2 re-infection.
4. Cross-reactivity of antibodies to multiple SARS-CoV-2 strains with suspected
altered sensitivity, like variant 20I/501Y.V1 and 20H/501Y.V2 (often referred
to as the UK and SA variant).
5. The effect of age on SARS-CoV-2 neutralizing and non-neutralizing antibody
response after SARS-CoV-2 infection or vaccination.
6. Long-term impact of SARS-CoV-2 infection on presence and abundances of
common respiratory bacterial pathogens


Reinfection study:
7. To determine the incidence rate of (co-)infections with influenza viruses
and RSV

exploratory objective
Host (respiratory microbiome, co-infections) and virus (viral load and
genotype) determinants of long-lasting immunity after SARS-CoV-2 infection.

This will be a prospective cohort study of patients with COVID-19 and their
household contacts

The burden of participating in this study is minimal. Blood collection will
take place only once and will be performed by a finger prick, which may cause
transient discomfort and only rarely infection. NP and OP swabs are also
collected only once. NP swabbing gives mild discomfort, may induce sneezing,
and may give a nosebleed in rare cases. OP swabbing also gives mild discomfort
and may induce a gagging reflex. Saliva collection holds no risk. Saliva
sampling will be repeated in total 10 times for SARS-CoV-2 positive subjects
and household members, but represents no burden also when repeated.
Participants will also be required to complete a total of 10 questionnaires,
which is expected to take 10x20 minutes maximum per household. No additional
visits to the hospital are required. Study staff will drop off required
sampling material and picked up saliva samples at participant homes.
It is important to note that study participants should still follow the advice
from the municipal health services regarding testing for SARS-CoV-2

The study physician will also inform participants about the results on
SARS-CoV-2 testing from the NP and OP swab obtained on day 7 by phone as soon
as results are available. Testing of these swabs will be performed immediately.
At the end of the study, the study physician will also inform participants on
the results from the saliva samples, which are not analysed directly.

FUP:
The burden of participating in the SARSLIVA-FUP study is minimal. Blood
collection will be performed by a finger prick, which may cause transient mild
discomfort and only rarely infection. Saliva collection holds no risk. The
questionnaires are expected to take 2x 5 minutes per person. The home visits
are estimated to take 10 minutes per person.

Substudy SARS-CoV-2 reinfections
The burden of participating in this study is minimal. Blood collection will
take place four times. On M3 and M9, blood will be self-collected at home by
finger prick. At M0 and M6, blood will be collected by venepuncture in adults
and children >= 16 years old. In children <1 years old, blood will be collected
by finger prick. In children 12-15 years old, blood will preferably be
collected by venepuncture (with EMLA), or by finger prick in case of
participants preference. This may cause transient mild discomfort and only
rarely infection. Oropharyngeal swab collection holds no risk. Oropharyngeal
sampling can cause mild discomfort. Participants will also be asked to complete
at least 4 questionnaires, which is expected to take 20 minutes maximum per
person per questionnaire. In case of symptoms, participants will fill in an
additional questionnaire, which is expected to take 20 minutes maximum per
person. The 2 home visits (Month0 and Month6) are estimated to take 5 minutes
per person in the household. Study staff will drop off required sampling
material and perform the venous blood collections.
Participants participating in the extension of the substudy, will be followed
up for additional 6 months. During these months, participants will continue
collecting NP/OP swabs. One additional blood collection and questionnaire is
implemeted at april 2024.