A issue in validate_and_count.py

[TequilaSunrise84]

https://github.com/FredHutch/Galeano-Nino-Bullman-Intratumoral-Microbiota_2022/blob/d72ffa3efbc5dd67d8697ce84611020fe316fa8b/10X_Visium_samples/validate_and_count.py#L13 I just run the code in validate_and_count.py I'm not sure which file is "metadata_file", I notice that the path of "metadata_file" is metadata_file = 'data_processing/selected_samples_for_counting/sample.visium_D1_metadata.csv', but the path and the file didn't appear in the previous analysis... I would like to know how to find this file. I would appreciate your help.

[hanruiw]

Hi, metadata file is the metadata of a Seurat object. Please let me know if you have any questions!

[TequilaSunrise84]

Hello, Thanks for your reply. I tried to use the metadata file extracted from sample.filtered_matrix@meta.data in Visium.R. But I still meet problem in process of extracting the Bacteria Positive cells. It seems that the code int(each_line_list[position]) has some problems. So I would like to check with you whether the "metadata_file" is correct and whether subsequent changes to this file are required. Thanks you very much !

[hanruiw]

Hi, sorry for the late response, the "metadata_file" is written using write.csv(sample.visium_metadata,sample.visium_metadata_filename, quote = FALSE,row.names = TRUE) and looks like this:

make sure it is annotated and the 'Total' is calculated. Please let me know if you have any questions!

[TequilaSunrise84]

Thanks you for your reply ! I will try it again !

[hanruiw]

Hi, sorry for confusion, I updated the pipeline code and sample names in visium output, could you please try again using the updated output files?

[TequilaSunrise84]

Hello, I have tried the pipeline code again with new output file and successfully run the code, thanks you for your help !