# Large, novel Spike deletions of unknown origin (not reproducible outside nextstrain framework)

**URL:** <https://nextstrain.discourse.group/t/large-novel-spike-deletions-of-unknown-origin-not-reproducible-outside-nextstrain-framework/845>\
**Category:** Help and Getting Started\
**Created:** [November 15, 2021, 3:18pm UTC](https://nextstrain.discourse.group/t/large-novel-spike-deletions-of-unknown-origin-not-reproducible-outside-nextstrain-framework/845 "2021-11-15T15:18:10Z")\
**Posts on this page:** 4\
**Page:** 1

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**Author:** ![guy](https://avatars.discourse-cdn.com/v4/letter/g/9e8a1a/32.png) [@guy](https://nextstrain.discourse.group/u/guy)\
**Post date:** [November 15, 2021, 3:18pm UTC](https://nextstrain.discourse.group/t/large-novel-spike-deletions-of-unknown-origin-not-reproducible-outside-nextstrain-framework/845/1 "2021-11-15T15:18:11Z")

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Dear forum,  
I was alerted by my colleagues that a newly sequenced SARS-CoV-2 sample had multiple novel deletions in the starting amino acids of the Spike protein.

Bracing for another variant of concern, I ran the sequencing data through my own pipeline (bwa-mem2+iVar) and submitted both to nextclade.

Nextclade identified large Spike deletions in both sequences:

 ![Screenshot from 2021-11-15 16-55-33](https://canada1.discourse-cdn.com/flex031/uploads/nextstrain/original/1X/8634472aa9fc15d5b0f0e25f9e126925a9ca36bf.png)

However, I could not find a single indication of these deletions outside of the nextclade results.

Both consensus sequences had the Spike start codon fully in-tact.

My variant caller (iVar) found no such deletions, save for the well known ones further down the gene (69-70 and so forth).  
Other tools, such as the GISAID CoVsurver and cov-glue, didn’t find anything out of the ordinary either.

[The consensus sequences (same sequencing data, different piplines) can be obtained here.](https://gitlab.com/-/snippets/2206864)

Do you have any idea what happened here?

Thanks in advance,  
Guy

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**Author:** ![rneher](https://yyz1.discourse-cdn.com/flex031/user_avatar/nextstrain.discourse.group/rneher/32/20_2.png) [@rneher](https://nextstrain.discourse.group/u/rneher)\
**Post date:** [November 16, 2021, 8:47am UTC](https://nextstrain.discourse.group/t/large-novel-spike-deletions-of-unknown-origin-not-reproducible-outside-nextstrain-framework/845/2 "2021-11-16T08:47:30Z")

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Thanks for flagging this. You are right, these deletions aren’t real.

the reason seems to be that the amino acid alignment is thrown off by the large numbers of `N` early in the sequence preceding the deletion around position 65 in spike. We’ll try to fix…

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**Author:** ![ivan-aksamentov](https://yyz1.discourse-cdn.com/flex031/user_avatar/nextstrain.discourse.group/ivan-aksamentov/32/80_2.png) [@ivan-aksamentov](https://nextstrain.discourse.group/u/ivan-aksamentov)\
**Post date:** [November 29, 2021, 1:12am UTC](https://nextstrain.discourse.group/t/large-novel-spike-deletions-of-unknown-origin-not-reproducible-outside-nextstrain-framework/845/3 "2021-11-29T01:12:07Z")

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Hi @guy,

We have improved alignment of peptides and now it should be a little better. It is released in the Nextclade CLI 1.5.0 and Nextclade Web 1.8.0.

Let us know if it solves the problems you observed.

See this PR for more details:

> <https://github.com/nextstrain/nextclade/pull/613>
>
> This is an implementation ~of Richard's idea here: https://github.com/nextstrain…/nextclade/issues/609#issuecomment-973891398~ Update: since then we refined the idea quite more.
> 
> The aim is to improve peptide alignment, and to hopefully solve problems like #609 #594, which might be due to inferred band width of the banded alignment being too small.
> 
> The proposed solution is to infer the band width and mean shift for peptide alignment from nucleotide alignment, instead of running another seed matching pass.
> 
> ~This branch is based on #612 and includes debug traces.~ (Not important)
> 
> Resolves #609 
> Resolves #594

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<div class="post-metadata">

**Author:** ![guy](https://avatars.discourse-cdn.com/v4/letter/g/9e8a1a/32.png) [@guy](https://nextstrain.discourse.group/u/guy)\
**Post date:** [December 23, 2021, 3:22pm UTC](https://nextstrain.discourse.group/t/large-novel-spike-deletions-of-unknown-origin-not-reproducible-outside-nextstrain-framework/845/4 "2021-12-23T15:22:47Z")

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Hi Ivan,  
Yes, this solved it. Thanks for the quick reply!