>> Welcome to module 2.6, "Detecting
and Prioritizing SARS-CoV-2 Variants."
This presentation is part
of the COVID-19 Genomic Epidemiology Toolkit
from CDC's Office of Advanced Molecular Detection.
My name is Dr. Shannon Matzinger,
and I'm the deputy program manager
with the Colorado Department of Public Health and Environment.
This module describes the detection and genome sequencing
of SARS-CoV-2 variants
to prioritize COVID-19 investigation
and contact tracing activities.
Be sure to check out the toolkit's other modules
which include a combination of case studies
and training material to help you get started supplementing
epidemiology with genome sequencing data.
As a PulseNet regional laboratory,
the Colorado Department of Public Health
and Environment entered the COVID-19 pandemic
with established genome sequencing capacity.
We had one Oxford Nanopore Minion
and two Illumina MiSeq instruments.
We also had funding from CDC to serve as a regional resource
for cloud computing capacity and analysis
of genome sequencing data.
Our laboratory director, Dr. Emily Travanty,
understood that whole genome sequencing would be important
to responding to COVID-19 and prepared from the beginning
by requesting additional instrumentation, reagents,
and staff in the pandemic's response budget.
So it seemed we were set up for success.
However, we had recently lost our single CDC funded
bioinformatician and our sequencing staff was pulled
into the diagnostic testing effort.
This left us with some rebuilding to do,
but we were able to hire a new bioinformatician as well
as two additional term limited bioinformaticians
to focus on SARS-CoV-2.
Once our diagnostic/testing laboratory was operating
at full capacity, we were able
to begin rebuilding a wet-bench staff for SARS-CoV-2 sequencing.
We completed our first sequencing run
in late August 2020.
Initially we focused on outbreak detection and cases
of suspected re-infection within congregate living facilities.
Between August and December 2020,
these sequencing efforts were made in an attempt to support
over 100 investigations.
The phylogenetic tree in this slide is based
on our CDPHE Nextstrain build,
but you can explore a similar one calculated
by CDC's SPHERES Consortium
for public sequencing data via the link
at the bottom of the slide.
Importantly, sequencing
from this time period was not representative
of infections statewide because our selection criteria were
biased towards congregate living facilities.
At the time, we were using sequencing data
to investigate outbreaks at these facilities,
addressing questions such as, "Was the virus introduced
to this facility once or several times?"
Or to confirm suspected re-infection cases
with questions like, "Was the resident re-infected
with a distinct SARS-CoV-2 viral type, or is the virus shedding
from a previous infection?"
We used information gained from these data
to inform infection control recommendations
such as cohorting and contact tracing efforts
to improve the response to subsequent outbreaks
in similar types of facilities.
Then in mid-December 2020,
Public Health England reported the spread of an emerging,
more transmissible variant of SARS-CoV-2,
referred to by its PANGO lineage designation as B.1.1.7.
It was soon classified as a variant of concern, or VOC,
by the World Health Organization and CDC.
One of the defining mutations to this variant,
which WHO was now calling Alpha, is a small deletion
in the gene encoding the spike protein.
This deletion happens to interfere
with the ThermoFisher TaqPath RT-PCR diagnostic assay's
ability to detect the spike gene.
This ends up producing a characteristic S gene target
failure, also known as SGTF.
This plot, taken from the earlier report on B.1.1.7
in the UK, suggested that SGTF could function
as a diagnostic screening tool for B.1.1.7.
For additional information on this and other emerging variants
of SARS-CoV-2, see module 1.4 of the toolkit.
CDC, along with clinical and commercial laboratories
in Colorado, was using the ThermoFisher TaqPath assay
for diagnostic testing.
Beginning in December 2020,
we started routinely routing all diagnostic samples with SGTF
to CDPHE to confirm the sequencing.
With these efforts,
we identified the first US infection with B.1.1.7
in Colorado later in the same month.
From December 2020 to January 2021,
our sequencing data confirmed that 100% of SGTF specimens
that were sequenceable were B.1.1.7.
This suggests that, in contrast to other parts of the country,
no other viral variants with the same signature were circulating
widely in Colorado.
We were careful to communicate that SGTF serves
as a screening tool and that sequencing was needed
to confirm the identification of B.1.1.7.
However, given that other lineages --
there were no other lineages identified with the signature
in Colorado, we felt comfortable reporting the confirmatory
RT-PCR results
to our epidemiology partners while sequencing was
still pending.
We found that early communication
of SGTF results was very useful in contact tracing
and investigation of potential instances
of B.1.1.7 transmission.
Colorado's detection and investigation
of the first US case of B.1.1.7 infection was an example
of epidemiology and laboratory coming together
to respond quickly to a public health concern.
Identification of the index case
and containment were swift and effective.
This success helped to increase statewide engagement
for broader SARS-CoV-2 surveillance.
We set up a daily meeting
to discuss SARS-CoV-2 variants sequencing results,
allowing our epidemiology colleagues to start each day
with the most up-to-date laboratory data
to support their contact tracing efforts.
Here's an example of a B.1.1.7 cluster investigation
that was informed by genome sequencing.
A commercial laboratory identified a single person
with an SGTF test result.
Although the specimen was not available for sequencing,
case investigation and contact tracing identified additional
B.1.1.7-positive cases confirmed by sequencing both
within this person's family and among co-workers.
CDPHE analyzes SARS-CoV-2 Illumina
or Oxford Nanapore sequencing data
on the GCP/Terra platform using the Cecret network pipelines
adapted by our bioinformatician Dr Laura Bankers.
The phylogenetic tree here displays genetic relatedness
highlighted in a cluster of B.1.1.7 sequences recovered
from the 10 secondary cases identified
through this investigation.
The PANGO lineage and Nextstrain clade assignments are also
indicated to the right of the tree.
Looking specifically at the spike gene sequences,
we could see that all of them share the characteristic
mutations that define B.1.1.7,
including the deletion responsible for SGTF.
In addition to prioritizing SGTFs for sequencing,
we implemented two other approaches to aid detection
of emerging variants
by improving our surveillance sampling.
First, each week we sequenced a random sampling
of at least 30 positive samples
from each participating clinical laboratory
within our state sentinel surveillance network.
Secondly, we now sequence all positive specimens identified
by diagnostic testing performed
at CDPHE laboratory the previous week.
SGTF specimens submitted by other laboratories
for confirmation and sequencing are flagged at submission
and excluded from surveillance sampling.
By April 2021, these sampling strategies allowed for us
to increase our sequencing throughput at CDPHE to more
than 1,000 samples a week.
As mentioned earlier, the CDPHE laboratory still perform
SARS-CoV-2 diagnostic testing mainly
for congregate living facilities,
from which all positive specimens then get sequenced.
These activities led to the identification
of the first cluster in Colorado of another variant of concern,
or VOC, B.1.351, which WHO has now named Beta.
This first cluster was suspected in the first week of March 2021
when multiple positive specimens collected
from the same county were linked to the same facility
and were identified as B.1.351.
Acting on this information, CDPHE epidemiologists were able
to identify additional related B.1.351 cases
from the outbreak at that facility.
The following week, a second cluster was suspected
from sequenced samples received from a different county,
this time among a family.
This phylogenetic tree
from Nextstrain displays those B.1.351 cases identified
by all surveillance samples within the CDPHE laboratory.
The first cluster, associated
with the congregate living facility, is labeled at the top
and includes a number of similar specimens
with few mutations apart.
The longer branches within this cluster also suggest
that cases may have been undersampled from this facility.
The sequence data also identified a potential linked
case from the community with no known connection
to this facility.
The second cluster was identified among family members
in a separate county.
The sequence data suggested that it was not related
to the outbreak at that congregate living facility.
This tree also includes four other epidemiologically
unrelated cases with different B.1.351 sequences showing the
difference within this lineage.
Together, this evidence suggests
that the VOC was already circulating throughout Colorado
beyond the clusters that have been identified.
In addition, the facility associated
with cluster one responded to detection of B.1.351
by increasing efforts to quickly vaccinate facility staff
and residents.
This facility continues to monitor and no additional cases
of B.1.351 have been identified since March 2021.
In the case of cluster two,
no secondary cases were identified outside
of the family unit.
We continue to monitor the relative proportions
of known variants of concern and variants
under investigation identified
through sentinel surveillance in Colorado.
As explained on previous slides,
these frequency calculations exclude specimens selected
for sequencing due to (1) signatures such as SGTF,
(2) identification during investigation
of known VOC/VOI investigations, or (3) special studies conducted
by other partners such as CDC.
CDC's national genomics surveillance program estimates
biweekly proportions of most variants for SARS-CoV-2 lineages
in the United States, accessible from the COVID-19
or COVID Data Tracker via the link at the bottom of the slide.
Sequencing at the CDPHE laboratories supports monitoring
of SARS-CoV-2 transmission and disease severity.
These data improve our situational awareness of efforts
to control the spread of emerging variants
such as the VOC B.1.427/429 which we highlighted
in the May 2021 MMWR article.
Looking forward, we hope these data will help us study certain
-- whether certain variants of concern may be associated
with either re-infections or vaccine breakthrough events,
a topic that is covered in toolkit module 2.5.
Finally, as more Coloradoans become vaccinated
and test positivity rates decrease,
we expect that SARS-CoV-2 detection and sequencing
for municipal wastewater to help focus testing efforts
and inform public health response.
As an example of this, see the case study in module 2.7
to learn how wastewater testing has been used
to guide sequencing efforts in Vermont.
In summary, sequencing efforts support both national
and statewide characterization of SARS-CoV-2 transmission
and COVID-19 severity.
Here in Colorado, sequencing adds valuable information
for case investigation, contact tracing, and outbreak response.
Most cases involving SARS-CoV-2 variants of concern identified
by sequencing have led to detection and quarantine
of additional secondary cases.
We've taken a layered approach
to prioritizing suspected variances based on SGTF
but also monitoring the contribution of targeted
and random genomics surveillance in our state.
This concludes module 2.6.
Part two of the toolkit contains case studies that review the use
of genomics in response to COVID-19.
Please visit the COVID-19 Genomic Epidemiology Toolkit
page where you can find more information on this topic
as well as a short survey
to provide feedback about this module.
On the toolkit page, you can also subscribe
to our mailing list to receive announcements as new modules
and material are released.
Thank you.