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Data analysis and follow-up
Data analysis and storage
The analytical phase of NGS testing occurs in two distinct stages, referred to as primary analysis and
secondary analysis. During primary analysis, raw data is generated by a sequencing instrument. Secondary
analysis takes this raw data as input and, through comparison with a reference genome, identifies genetic
variants present in the specimen. Following quality control assessment of the results of primary and
secondary analysis, the post-analytical phase, referred to as tertiary analysis, begins. Tertiary analysis
[29]
includes annotation, interpretation, and reporting . For secondary and tertiary analysis of NGS-based NBS
pilot data, more than half of the respondents (n = 8) will be using a hybrid solution including a mix of in-
house and commercially available analytical tools. Four initiatives have selected commercial software, while
one will be using in-house developed bioinformatic tools. Another initiative has yet to be decided regarding
this part of the project. Although three are undecided and others may change strategy during the course of
their studies, six initiatives have chosen to store data on premises, three will be using cloud-based solutions,
while five others will be using both on-premise storage and cloud-based solutions. The type of files to be
stored includes, for most, variant call format (VCF) and FASTQ, with a minority also looking at keeping
compressed reference-oriented alignment map (cram), special callers, annotated/prioritized variant outputs,
and files on quality control. The duration of data storage is not standardized across the initiatives; four
initiatives will keep these files for three to four years, while the others intend to store them for longer, with
two respondents specifying that they will store data for 10 years to support long-term clinical follow-up.

Return of results
The desired or estimated time from sample collection to results varies widely among respondents, from four
days to four months, although a third of initiatives have yet to define this aspect. Apart from three initiatives
not seeking to return any results to study participants, seven initiatives aim to return results to families with
positive and negative genetic screening results while four will only inform parents of babies with a positive
screening result.

Post-service evaluation, data linkage and clinical follow-up
Ten of 14 initiatives plan to recontact the parents of the newborns for post-service evaluation of their
participation in the pilot studies at one or more of the following time points: at the end of the study, 3 or 12
months after the end of the initiative, or even within three years after study conclusion.

When asked whether genetic screening results will be linked to clinical datasets in the long term, five
respondents who answered positively were largely undecided as to how this linkage will or should happen.
Only one initiative has a specific plan to store de-identified genomic sequence data together with ongoing
health data in a national repository. This practice will continue until the participant withdraws, i.e., parents
withdraw on behalf of the newborn, or at age 16 when the young person will be asked to consent for their
data to remain as part of the study.

Half of the initiatives will follow up on clinical outcomes, although how this will be done is not yet fully
defined. For one pilot specifically, follow-up will be done with clinicians and families of babies who
screened positive to assess clinical outcomes.

Of the seven respondents who answered positively to follow-up on clinical outcomes, four indicated that
these clinical outcomes will not be linked to electronic health records (EHRs) or other data sources.
However, one initiative has indicated that some outcome data will be ascertained through de-identified
health records and included in a national repository. Two initiatives would like to link clinical outcomes to