Feature-table rarefy command dropping samples

Hello!

I am trying to create a rarefied feature table and am hoping for some clarification on my output. I created my feature table and started filtering it based on a number of parameters: filtering out mitochondrial sequences; filtering out features that could not be identified to the order level or lower; filtering out low-frequency features. Once I had my filtered table, I double-checked the total number of sequence reads per sample and found that my smallest sample contained 11,039 sequences. I decided to rarefy to 11,000 sequences using the following command:

qiime feature-table rarefy
–i-table filtered-table.qza
–p-sampling-depth 11000
–o-rarefied-table filtered-table-rarefied.qza
–verbose

However, when I went to download the .csv of the output table, I noticed my sample containing 11,039 samples had been dropped. I then re-ran this command rarefying to 10,000 sequences and then to 9,000 sequences, and it wasn’t until I rarefied to 9,000 sequences that this one sample was retained in the final table.

I am wondering if there’s something I did wrong that is resulting in this sample being dropped, or whether there is another explanation for this? I would rather not have to rarefy to 9,000 sequences if I don’t have to, but I’m not sure how to remedy this on my own.

Thanks!

Sydney

Hi @Sydney_Morgan!

Did you run feature-table summarize on this filtered table? Are you 100% sure that the lowest feature frequency in this table is in fact 11,039 sequences? When you look at the “Interactive Sample Detail” tab of the aforementioned visualization, and set the sampling depth to the same values you are reporting here, do you see the same results?

If that doesn’t reveal anything on your end, can you please send us a link to download the filtered feature table you are rarefying? Thanks, we will figure this out! :t_rex:

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Hi @thermokarst,

I’m so sorry to have bothered you! @Mehrbod_Estaki helped me realize I was accidentally looking at a pre-filtered version of the table when I was double-checking my sequence counts.

I’ll be more careful in the future!

Sydney

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