微生物组核心OTU鉴定usearch otutab_core
扩增子分析神器VSEARCH 分析流程 2.8.1中文帮助文档
otutab_core
http://www.drive5.com/usearch/manual/cmd_otutab_core.html
鉴定核心微生物组——大多数样品中出现的OTUs，这也是Usearch11新增的功能。
本质上是统计每个OTUs在大量样品中出现的频率。比如在所有样本中都出现，即100%。特别大量样本时，如几千个，可能很少有OTU存在于所有样本，可选择95%，或90%的样本中出现的OTUs作为核心OTUs。
Identifies a possible “core microbiome” of OTUs which are present in more samples than others.
输入文件为经典的QIIME格式OTU表
Input is an OTU table in QIIME classic format.
在一些样品或许多样本中出现的OTU可能是假的，原因可能是串道(cross-talk)或扩增、测序错误的假OTU。为方便检查，otutab_core命令产生分析报告，标明哪些OTU可能是串道、哪些可能是测序错误。
The presence of an OTU in some or many samples can be spurious because of cross-talk or because the OTU itself is spurious. To enable manual review, the otutab_core command generates a report indicating cases where the presence of an OTU may be spurious due to cross-talk, and where an OTU may be spurious due to sequence errors.
如果使用-sintaxin参数指定物种注释，报导中也会包括物种信息。
If a sintax tabbed file is provided using the -sintaxin option, then the taxonomy of the core OTUs is included in the report.
如果使用distmxin选项提供距离矩阵，可用于鉴定主导的OTUs，例如在报告中高丰度OTUs与低丰度OTUs相似，如果存在主导OTUs，那这些低丰主OTUs是假的。
If a distance matrix is provided using the distmxin option, this is used to identify possible dominant OTUs, i.e. high-abundance OTUs which are similar to a low-abundance OTUs in the report. If there is a dominant OTU, this may indicate that the low-abundance OTU is spurious.
-tabbedout参数指定输出文件。OTUs按样品中出现频率排序，包括如下12个字段。
The -tabbedout option specifies the output file. OTUs are sorted in order of decreasing number of samples where they are present. Fields are:
OTU = name of the OTU.
Samples = number of samples where the OTU has a non-zero count.
Size = total number of reads assigned to this OTU.
DomOTU = high-abundance “dominant” OTU which is very similar to this OTU, if any.
DomSize = total number of reads assigned to the dominant OTU.
DomId = identity of the dominant OTU with this OTU.
Min = minimum count for this OTU.
LoQ = low quartile count for this OTU.
Med = median count for this OTU.
HiQ = high quartile count for this OTU.
Max = maximum count for this OTU.
Taxonomy = condensed taxonomy prediction.
If the minimum or LoQ count is much smaller than the maximum count, this suggests that the smaller counts may be due to cross-talk.
If the size of an OTU is much smaller than a neighboring “dominant” OTU, then the OTU itself may be spurious due to sequence error.
使用实例
Example
基于OTU序列计算距离矩阵
usearch -calc_distmx otus.fa -tabbedout distmx.txt \
-sparsemx_minid 0.9 -termid 0.8物种注释(己完成可跳过)
usearch -sintax otus.fa -strand both -db ref16s.txt \
-tabbedout sintax.txt鉴定核心OTUs
usearch -otutab_core otutab.txt -distmxin distmx.txt \
-sintaxin sintax.txt -tabbedout core.txt在使用中，我碰到了报错。可以把-distmxin distmx.txt去掉。可以正常获得结果。
结果文件如下：
OTUID Samples Size Freq DomOTU DomSize DomId Min LoQ Med HiQ Max Taxonomy Core
OTU_2 1000 5079019 0.131 . . . 162 1915 3270 5470 23217 d:Bacteria,p:"Proteobacteria",c:Betaproteobacteria,o:Burkholderiales,f:Burkholderiaceae,g:Ralstonia,s:Ralstonia_mannitolilytica 100
OTU_34 999 180434 0.00466 . . . 1 40 83 174 2484 d:Bacteria,p:"Proteobacteria",c:Gammaproteobacteria,o:Clostridiales,f:Chloroplast,g:Streptophyta,s:Porticoccus_litoralis 99.9154具体每列的意义见上方帮助文档。最主要的结果是Samples列，即该OTU在多少个样本中检测到。我们还需要将此数值除以总样本量，才能获得Core OTU的比例，方便筛选核心OTUs。
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