lefse analysis error

[KOMALSKK]

Hello, Can you help me with the following error. I created a phyloseq object physeq and performed lefse.

library(microbiomeMarker) norm_tss(physeq) normalize(physeq, method = "TSS")

data(physeq) mm_test <- normalize(physeq, method = "CPM") %>% run_lefse( wilcoxon_cutoff = 0.01, norm = "none", # must be "none" since the input has been normalized group = "type", kw_cutoff = 0.01, multigrp_strat = TRUE, lda_cutoff = 2

But I get the following output; mm_lefse <- run_lefse(

• physeq,
• wilcoxon_cutoff = 0.05,
• group = "type",
• kw_cutoff = 0.05,
• multigrp_strat = TRUE,
• lda_cutoff = 2
• ) Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Found more than one class "phylo" in cache; using the first, from namespace 'phyloseq' Also defined by ‘tidytree’ Error: Too small samples in each class or the variance of feature abundances within a class too small (zero or near zero).

Can you help me finding the error, here.

Thank you.