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Nelly-Wambui edited this page Jan 31, 2022 · 7 revisions

nf-core/ampliseq

Test on Yosef's dataset

Pipeline exited with an error on metadata:

Functional analysis - picrust

Run command

#16S rRNA gene amplicon analysis of Illumina paired-end data
nextflow run nf-core/ampliseq -profile <docker/singularity/podman/shifter/charliecloud/conda/institute> --input "data" --FW_primer "GTGYCAGCMGCCGCGGTAA" --RV_primer "GGACTACNVGGGTWTCTAAT" --metadata "data/Metadata.tsv --picrust"

output

EC prediction

function description sampleID_1 sampleID_1a sampleID_2 sampleID_2a EC:1.1.1.1 Alcohol dehydrogenase 944.32 687.8 1402.0000000000002 763.12
EC:1.1.1.100 3-oxoacyl-[acyl-carrier-protein] reductase 2057.99 1762.1999999999996 2723.83 2094.82
EC:1.1.1.103 L-threonine 3-dehydrogenase 0.0 3.2 0.0 1.5
EC:1.1.1.108 Carnitine 3-dehydrogenase 46.0 12.0 28.0 34.4
EC:1.1.1.11 D-arabinitol 4-dehydrogenase 66.0 0.0 92.0 0.0
EC:1.1.1.122 D-threo-aldose 1-dehydrogenase 10.0 0.0 0.0 10.0
EC:1.1.1.125 2-deoxy-D-gluconate 3-dehydrogenase 46.0 15.2 28.0 28.5
EC:1.1.1.130 3-dehydro-L-gulonate 2-dehydrogenase 0.0 3.2 0.0 1.5
EC:1.1.1.132 GDP-mannose 6-dehydrogenase 46.0 12.0 28.0 34.4
EC:1.1.1.133 dTDP-4-dehydrorhamnose reductase 603.0 648.0 881.0 730.2
EC:1.1.1.136 UDP-N-acetylglucosamine 6-dehydrogenase 22.0 0.0 21.0 0.0
EC:1.1.1.14 L-iditol 2-dehydrogenase 151.0 259.0 230.0 342.0
EC:1.1.1.140 Sorbitol-6-phosphate 2-dehydrogenase 0.0 3.2 0.0 1.5
EC:1.1.1.157 3-hydroxybutyryl-CoA dehydrogenase 518.75 61.2 749.0 104.9
EC:1.1.1.159 7-alpha-hydroxysteroid dehydrogenase 0.0 3.2 0.0 1.5
EC:1.1.1.169 2-dehydropantoate 2-reductase 386.75 156.2 379.0 214.7
EC:1.1.1.17 Mannitol-1-phosphate 5-dehydrogenase 22.0 3.2 21.0 1.5
EC:1.1.1.18 Inositol 2-dehydrogenase 224.0 12.0 290.0 44.4
EC:1.1.1.193 5-amino-6-(5-phosphoribosylamino)uracil reductase 489.08 434.2000000000001 723.5 494.23\

KO prediction

function description sampleID_1 sampleID_1a sampleID_2 sampleID_2a
K00001 E1.1.1.1, adh; alcohol dehydrogenase [EC:1.1.1.1] 144.16 4.0 153.0 8.66
K00003 E1.1.1.3; homoserine dehydrogenase [EC:1.1.1.3] 598.08 699.0 889.4999999999999 759.13
K00004 BDH, butB; (R,R)-butanediol dehydrogenase / meso-butanediol dehydrogenase / diacetyl reductase [EC:1.1.1.4 1.1.1.- 1.1.1.303] 55.75 2.0\ 28.0 14.8
K00005 gldA; glycerol dehydrogenase [EC:1.1.1.6] 0.0 3.2 0.0 1.5
K00007 dalD; D-arabinitol 4-dehydrogenase [EC:1.1.1.11] 66.0 0.0 92.0 0.0
K00008 SORD, gutB; L-iditol 2-dehydrogenase [EC:1.1.1.14] 151.0 259.0 230.0 342.0
K00009 mtlD; mannitol-1-phosphate 5-dehydrogenase [EC:1.1.1.17] 22.0 3.2 21.0 1.5
K00010 iolG; myo-inositol 2-dehydrogenase / D-chiro-inositol 1-dehydrogenase [EC:1.1.1.18 1.1.1.369] 224.0 12.0 290.0 44.4
K00012 UGDH, ugd; UDPglucose 6-dehydrogenase [EC:1.1.1.22] 536.16 667.0 752.6700000000002 734.13
K00013 hisD; histidinol dehydrogenase [EC:1.1.1.23] 561.08 702.2 861.4999999999999 753.2299999999998
K00014 aroE; shikimate dehydrogenase [EC:1.1.1.25] 834.08 722.2 1210.5 805.0299999999999
K00015 gyaR, GOR1; glyoxylate reductase [EC:1.1.1.26] 59.0 6.0 123.0 17.0
K00016 LDH, ldh; L-lactate dehydrogenase [EC:1.1.1.27] 22.0 0.0 21.0 0.0
K00018 hprA; glycerate dehydrogenase [EC:1.1.1.29] 234.5 16.0 383.33 45.73
K00019 E1.1.1.30, bdh; 3-hydroxybutyrate dehydrogenase [EC:1.1.1.30] 356.75 14.0 516.0 58.4
K00020 mmsB, HIBADH; 3-hydroxyisobutyrate dehydrogenase [EC:1.1.1.31] 615.66 101.0 770.33 174.58999999999997
K00021 HMGCR; hydroxymethylglutaryl-CoA reductase (NADPH) [EC:1.1.1.34] 13.0 193.0 10.0 188.0
K00023 phbB; acetoacetyl-CoA reductase [EC:1.1.1.36] 439.0 27.0 814.0 45.0\

Metacyclic pathway

pathway description sampleID_1 sampleID_1a sampleID_2 sampleID_2a 1CMET2-PWY N10-formyl-tetra\hydrofolate biosynthesis 526.527664084303 347.08079355769274 780.8916021044117 405.84973943743387 3-HYDROXYPHENYLACETATE-DEGRADATION-PWY 4-hydroxyphenylacetate degradation 109.92910599645849 18.811789371846768 85.06884911544789 50.2414031080214
ALL-CHORISMATE-PWY superpathway of chorismate metabolism 0.0 11.575132219341493 0.0 5.985983227364799
ANAEROFRUCAT-PWY homolactic fermentation 413.63447082617796 155.0766572263989 554.5721127129322 219.3084247425869
ANAGLYCOLYSIS-PWY glycolysis III (from glucose) 524.3632145962823 620.6473219841798 772.1713957533265 674.9066443207296
ARG+POLYAMINE-SYN superpathway of arginine and polyamine biosynthesis 155.21945884030328 268.78221307848816 199.2504180408352 332.4801580646289
ARGDEG-PWY superpathway of L-arginine, putrescine, and 4-aminobutanoate degradation 43.086934492011224 9.809319229777303 21.321620172510354 17.52629059463373
ARGORNPROST-PWY arginine, ornithine and proline interconversion 83.35873429655051 27.49242053505412 61.386195824271326 70.30705400767395
ARGSYN-PWY L-arginine biosynthesis I (via L-ornithine) 434.8448212544232 657.3522014687397 587.5562795083328 725.0708265416972
ARGSYNBSUB-PWY L-arginine biosynthesis II (acetyl cycle) 625.4323962283961 692.5956382599318 930.9378469363913 770.3689225138959
ARO-PWY chorismate biosynthesis I 633.1710876328017 619.4667172424088 935.962798904578 695.8269243720179

MBBU Accreditation dada2

Output

Raw plots

raw plots

count tracking

dada2_input	filtered	dada_f	dada_r	merged	nonchim	final_perc_reads_retained\

1 2500 1751 1429 1420 1265 1265 50.6
1a 2500 2237 1539 1600 1315 1315 52.6
2 2500 2310 1857 1857 1630 1630 65.2
2a 2500 2239 1567 1646 1362 1362 54.5\

Taxonomic assignment

	Kingdom	Phylum	Class	Order	Family	Genus
GGGTGCGAGCGTTAATCGGAATTACTGGGCGTAAAGCGTGCGCAGGCGGTTTTGTAAGACAGACGTGAAATCCCCGGGCTTAACCTGGGAACTGCGTTTGTGACTGCAAGGCTAGAGTACGGCAGAGGGGGGTAGAATTCCACGTGTAGCAGTGAAATGCGTAGATATGTGGAGGAATACCGATGGCGAAGGCAGCCCCCTGGGTCGATACTGACGCTCATGCACGAAAGCGTGGGGAGCAA	Bacteria	Proteobacteria	Betaproteobacteria	Nitrosomonadales	Gallionellaceae	Sideroxydans
AGCCACAAGTGGCAACCACCTTTATTGGGCCTAAAGCGTCCGTAGCCGGTTTTGTAAATTTTCTGTGAAATCGTTGCGCTTAACGTAACGGCGTGCAGAAAACACTGCAAGACTAGGGACCGGGAGGAGTCAGGGGTATTTTGGGGGGAGCGGTAAAATGCTATAATCCCCGAAGGACCACCTGTGGCGAAGGCGCCTGACTAGAACGGATCCGACGGTGAGGGACGAAAGCTAGGGGAGCAA	Archaea	Woesearchaeota	NA	NA	NA	NA
CACCTCAAGTGGTCAGGATGATTATTGGGCCTAAAGCATCCGTAGCTCGTTTTGTAAGTTTTCGGTTAAATCCATGCGCTTAACGTATGGGCTGCCGGGAATACTGCATAACTAGGAAGTGGGAGAGGTAGACGGTACTCGGTAGGAAGGGGTAAAATCCTTTGATCTATTGATGACCACCTGTGGCGAAGGCGGTCTACCAGAACACGTTCGACGGTGAGGGATGAAAGCTGGGGGAGCAA	Archaea	Thaumarchaeota	Nitrosopumilales	Nitrosopumilaceae	Nitrosopumilus	NA
GGGTGCGAGCGTTAATCGGATTTACTGGGCGTAAAGCGTGCGTAGGCGGCTTTTTAAGTCGGATGTGAAATCCCTGAGCTTAACTTAGGAATTGCATTCGATACTGGGAAGCTAGAGTATGGGAGAGGATGGTAGAATTCCAGGTGTAGCGGTGAAATGCGTAGAGATCTGGAGGAATACCGATGGCGAAGGCAGCCATCTGGCCTAATACTGACGCTGAGGTACGAAAGCATGGGGAGCAA	Bacteria	Proteobacteria	Gammaproteobacteria	Pseudomonadales	Moraxellaceae	Acinetobacter
GGGTGCGAGCGTTAATCGGAATTACTGGGCGTAAAGCGTGCGCAGGCGGTTCTATAAGACAGATGTGAAATCCCCGGGCTCAACCTGGGAATGGCATTTGTGACTGTAGAGCTAGAGTACGGTAGAGGGGGATGGAATTCCGCGTGTAGCAGTGAAATGCGTAGATATGCGGAGGAACACCGATGGCGAAGGCAATCCCCTGGACCTGTACTGACGCTCATGCACGAAAGCGTGGGGAGCAA	Bacteria	Proteobacteria	Betaproteobacteria	Burkholderiales	Comamonadaceae	Polaromonas

Error

  • Creating a phyloseq object

Yosef's Pipeline

Output

Quality Plot

quality plot

Assigning Taxonomy

Kingdom Phylum Class Order Family
ASV_1 "Bacteria" "Proteobacteria" "Gammaproteobacteria" "Burkholderiales" "Gallionellaceae"
ASV_2 "Archaea" "Nanoarchaeota" "Nanoarchaeia" "Woesearchaeales" NA
ASV_3 "Bacteria" "Proteobacteria" "Gammaproteobacteria" "Pseudomonadales" "Pseudomonadaceae"
ASV_4 "Bacteria" "Proteobacteria" "Gammaproteobacteria" "Pseudomonadales" "Moraxellaceae"
ASV_5 "Bacteria" "Proteobacteria" "Gammaproteobacteria" "Pseudomonadales" "Moraxellaceae"
ASV_6 "Bacteria" "Nitrospirota" "Thermodesulfovibrionia" NA NA
Genus Species
ASV_1 "Sideroxydans" NA
ASV_2 NA NA
ASV_3 "Pseudomonas" NA
ASV_4 "Acinetobacter" NA
ASV_5 "Acinetobacter" NA
ASV_6 NA NA \

Error

Error in validObject(.Object) : invalid class “phyloseq” object:
Component sample names do not match.
Try sample_names()\

TO DO

  • Nextflow script for automating MBBU 16S Accreditation dada2
  • Run 18S, ITS on nf-core pipeline
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