# Question in lefse

**URL:** <https://forum.mothur.org/t/question-in-lefse/21173>\
**Category:** Commands in mothur\
**Created:** [November 1, 2021, 6:40am UTC](https://forum.mothur.org/t/question-in-lefse/21173 "2021-11-01T06:40:27Z")\
**Posts on this page:** 4\
**Page:** 1

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**Author:** ![uihyun](https://avatars.discourse-cdn.com/v4/letter/u/dc4da7/32.png) [@uihyun](https://forum.mothur.org/u/uihyun)\
**Post date:** [November 1, 2021, 6:40am UTC](https://forum.mothur.org/t/question-in-lefse/21173/1 "2021-11-01T06:40:27Z")

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Hi. i have a question while i’m using the lefse command.  
I analyzed my samples from 2 time points(0 time and 1 month).  
I got the p-value 0.013 using amova.  
As i wanted to know which differences were between 2 time points, i used metastats then lefse.  
I knew which OTUs were significantly different between 2 time points form metastats.  
But during using lefse, it was said “You did not provide a class”.  
I wonder the OTUs significantly different are in which time point and i want to know how i can use the class option and whether i should add the class in which file (design file or metadata file)

Thanks  
uihyun

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**Author:** ![pschloss](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/pschloss/32/4_2.png) [@pschloss](https://forum.mothur.org/u/pschloss)\
**Post date:** [November 2, 2021, 6:07pm UTC](https://forum.mothur.org/t/question-in-lefse/21173/2 "2021-11-02T18:07:45Z")

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Hi,

You need to provide infomration about the time point in a design file

That being said, you shouldn’t be doing metastats and then lefse. The two test are not independent. You should pick one and use that. Normally, I’d suggest lefse. However, if you have paired data where you are analyzing the same entity at two time points, you really need to perform a paired analysis. You could do that with a wilcoxon test in R to see if there are taxa that have a change that is significantly different from 0.

Pat

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**Author:** ![uihyun](https://avatars.discourse-cdn.com/v4/letter/u/dc4da7/32.png) [@uihyun](https://forum.mothur.org/u/uihyun)\
**Post date:** [November 4, 2021, 4:54pm UTC](https://forum.mothur.org/t/question-in-lefse/21173/3 "2021-11-04T16:54:26Z")

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Hi!  
Your feedback was so helpful to me. I’m studying about which test will be optimal for my data.  
I’m gonna do the wilcoxon test in R!

Thanks  
uihyun

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**Author:** ![system](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/system/32/2_2.png) [@system](https://forum.mothur.org/u/system)\
**Post date:** [November 14, 2021, 4:54pm UTC](https://forum.mothur.org/t/question-in-lefse/21173/4 "2021-11-14T16:54:36Z")

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