Alpha diversity with no singletons

Hi, after picking OTUs I have removed singleton sequences from my entire dataset, but am now wondering whether it is still possible to compute richness alpha diversity metrics (eg. Chao1) after excluding singletons, since these metrics treat singletons explicitly in their calculation?

More specifically, I am interested in comparing my samples in terms of a particular metadata category (sample site), so because I removed singletons across all samples, should the output of diversity analysis still be of relevance for means of comparison? Even if the absolute values may not be accurate?

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Hi, there is a good discussion on this topic in this post. Let us know if that answers your question.

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