<p>I was able to get good annotation using prokka by using following commands in prokka.</p>

<pre><code>prokka --proteins reference.gb --outdir annotation  --prefix myprotein contigs.fa
</code></pre>

<p>All the reading frames were annotated. Actually the ones that are hypothetical in reference genomes are also annotated. </p>

CC BY-SA 4.0
2022-03-08T16:02:56.347Z
I definitely agree that Prokka could be repurposed and think this is a good response.
M__
CC BY-SA 4.0
2018-07-22T17:16:33.033Z
2018-07-22T17:16:33.033Z
LoveVirology
<p>If you are looking for another tool, you could try <a href="https://github.com/jhcepas/eggnog-mapper" rel="nofollow noreferrer">eggnog mapper</a>: it is an annotation tool based on orthology assignments. The EggNOG database happens to have a section for <a href="http://eggnogdb.embl.de/#/app/viruses" rel="nofollow noreferrer">viral Orthologous Groups</a> so you might obtain better results with it.</p>

<p>Note that emapper annotates coding sequences, not whole genomes. You need an extra step to predict CDS, for instance using <a href="https://github.com/hyattpd/Prodigal" rel="nofollow noreferrer">Prodigal</a>.</p>

<p>Basic usage with the optimized database for viral models:</p>

<pre><code>prodigal -i my.genome.fna -o my.genes -a my.proteins.faa
</code></pre>

<p>then</p>

<pre><code>python emapper.py -i my.proteins.faa --output polb_viruses  -d viruses
</code></pre>

CC BY-SA 4.0
2018-07-24T08:34:05.460Z
Interesting, do you have a source on this? The documentation of eggnog-mapper mentions "the annotation of novel genomes". I am currently using it to annotate MAGs, so I'd rather know if I am doing something wrong.
Nils Giordano
CC BY-SA 4.0
2018-07-26T02:15:31.157Z
Eggnog resulted in empty file. Maybe it doesn't work for individual genomes as mentioned by @EliKorvigo
LoveVirology
CC BY-SA 4.0
2018-07-26T06:48:39.633Z
eggnog-mapper takes predicted proteins as input, not whole genomes. You should not use it on raw metagenomes either. You need an extra step to predict CDS with the tool of your choice (e.g. [Prodigal](https://github.com/hyattpd/Prodigal)). `prodigal -i my.genome.fna -o my.genes -a my.proteins.faa` then feed `my.proteins.faa` to eggnog-mapper.
Nils Giordano
CC BY-SA 4.0
2018-07-26T19:49:25.737Z
My wording has been quite a bit too harsh, I guess. I have only applied eggnog to metagenomes and metatranscriptomes, because in these cases it's not practical to use group-specific homology databases and HMMs. Eggnog's authors have even emphasised this point in their [paper](https://www.ncbi.nlm.nih.gov/pubmed/28460117). On the other hand, when you do have a clean assembly, you can use a hierarchical group-specific annotator, such as Prokka. Of course, it doesn't mean one can't use eggnog for genomes, hence I find it necessary to apologise for my previous statement.
Eli Korvigo
CC BY-SA 4.0
2018-07-23T16:09:50.017Z
2018-07-26T06:56:47.623Z
2018-07-26T06:56:47.623Z
Nils Giordano
<p>Both of the annotation tools you used are designed for prokaryotic genome annotation. I would not expect them to work very well for viruses. </p>

CC BY-SA 4.0
2018-07-21T19:33:13.847Z
You can choose between Bacteria, Archea or Viruses when using prokka. So, I think it should work for viruses too.
LoveVirology
CC BY-SA 4.0
2018-07-21T19:47:52.843Z
@LRJoshi viruses (especially small ones) tend to have overlapping and nested ORFs: something a standard prokaryotic/eukaryotic gene detector (e.g. Glimmer, Genemark, Prodigal) is unlikely to account for.
Eli Korvigo
CC BY-SA 4.0
2018-12-24T10:47:23.380Z
Prokka can do viruses, but you may want to change which translation table it uses to get better blast results
Joe Healey
CC BY-SA 4.0
2018-07-21T09:13:07.217Z
2018-07-21T09:13:07.217Z
heathobrien
<p>I was experimenting Prokka and RAST annotation tools. So, I took a well-annotated swinepox virus genome from genebank <em>(NCBI Reference Sequence: NC_003389.1)</em>.</p>
<p>I ran those sequences on Prokka and RAST Seed server at the same time. I can see that only a few (may be around 1%) of the genes were annotated. Most of them were predicted as hypothetical protein. And the results were comparable between Prokka and RAST.</p>
<p>I would assume that these tools look for similar sequences in NCBI and find the best-match protein. But looks like that is not the case.</p>
<p>I am looking for prediction tool(s) that will:</p>
<ul>
<li>find a well annotated swinepox virus genome in Genbank to predict most of the proteins.</li>
<li>improve on the prediction of Prokka and Rast to assign functions to otherwise <strong>hypothetical protein</strong></li>
</ul>
<p>Would prediction tools such as Genemark or other genome annotation tools be beneficial.</p>
<p>Alternatively have I misunderstood the concept of annotation?</p>
<p>Assistance would be very welcome.</p>
<p><em>I have attached the image for comparison. Left one the Swinepox genome in .gb format and right one is the same genome annotated with Prokka.</em></p>
<p><a href="https://i.sstatic.net/eefc1.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/eefc1.png" alt="Image : sorry for the low quality screenshot" /></a></p>

CC BY-SA 4.0
2018-07-20T19:47:23.973Z
Please, provide links to the annotation servers along with all options you've specified.
Eli Korvigo
CC BY-SA 4.0
2018-07-21T19:29:54.423Z
I used Galaxy server.  I chose viruses under kingdom. All the other parameters were kept as default.
LoveVirology
CC BY-SA 4.0
2018-12-23T22:48:52.107Z
Prokka works well on bacterial genomes. I quesiton whether it will be useful here.
M__
<sequence-annotation><ngs><covid-19><virus>
Annotation with Prokka or RAST
# Annotation with Prokka or RAST

(Asked by: username_0 on 2018-07-20)

I was experimenting Prokka and RAST annotation tools. So, I took a well-annotated swinepox virus genome from genebank *(NCBI Reference Sequence: NC\_003389.1)*.

I ran those sequences on Prokka and RAST Seed server at the same time. I can see that only a few (may be around 1%) of the genes were annotated. Most of them were predicted as hypothetical protein. And the results were comparable between Prokka and RAST.

I would assume that these tools look for similar sequences in NCBI and find the best-match protein. But looks like that is not the case.

I am looking for prediction tool(s) that will:

* find a well annotated swinepox virus genome in Genbank to predict most of the proteins.
* improve on the prediction of Prokka and Rast to assign functions to otherwise **hypothetical protein**

Would prediction tools such as Genemark or other genome annotation tools be beneficial.

Alternatively have I misunderstood the concept of annotation?

Assistance would be very welcome.

*I have attached the image for comparison. Left one the Swinepox genome in .gb format and right one is the same genome annotated with Prokka.*

[![Image : sorry for the low quality screenshot](https://i.sstatic.net/eefc1.png)](https://i.sstatic.net/eefc1.png)

**Tags:** sequence-annotation, ngs, covid-19, virus
> **Comments:**
> **username_1**: Please, provide links to the annotation servers along with all options you've specified.  
> **username_0**: I used Galaxy server.  I chose viruses under kingdom. All the other parameters were kept as default.  
> **username_2**: Prokka works well on bacterial genomes. I quesiton whether it will be useful here.  

---

## 3 answers

### Answer by username_0 on 2018-07-22. Score: 1

I was able to get good annotation using prokka by using following commands in prokka.
prokka --proteins reference.gb --outdir annotation  --prefix myprotein contigs.fa

All the reading frames were annotated. Actually the ones that are hypothetical in reference genomes are also annotated.

> **Comments:**
> **username_2**: I definitely agree that Prokka could be repurposed and think this is a good response.  

---

### Answer by username_3 on 2018-07-23. Score: 1

If you are looking for another tool, you could try eggnog mapper: it is an annotation tool based on orthology assignments. The EggNOG database happens to have a section for viral Orthologous Groups so you might obtain better results with it.
Note that emapper annotates coding sequences, not whole genomes. You need an extra step to predict CDS, for instance using Prodigal.
Basic usage with the optimized database for viral models:
prodigal -i my.genome.fna -o my.genes -a my.proteins.faa

then
python emapper.py -i my.proteins.faa --output polb_viruses  -d viruses

> **Comments:**
> **username_3**: Interesting, do you have a source on this? The documentation of eggnog-mapper mentions "the annotation of novel genomes". I am currently using it to annotate MAGs, so I'd rather know if I am doing something wrong.  
> **username_0**: Eggnog resulted in empty file. Maybe it doesn't work for individual genomes as mentioned by @EliKorvigo  
> **username_3**: eggnog-mapper takes predicted proteins as input, not whole genomes. You should not use it on raw metagenomes either. You need an extra step to predict CDS with the tool of your choice (e.g. [Prodigal](https://github.com/hyattpd/Prodigal)). `prodigal -i my.genome.fna -o my.genes -a my.proteins.faa` then feed `my.proteins.faa` to eggnog-mapper.  
> **username_1**: My wording has been quite a bit too harsh, I guess. I have only applied eggnog to metagenomes and metatranscriptomes, because in these cases it's not practical to use group-specific homology databases and HMMs. Eggnog's authors have even emphasised this point in their [paper](https://www.ncbi.nlm.nih.gov/pubmed/28460117). On the other hand, when you do have a clean assembly, you can use a hierarchical group-specific annotator, such as Prokka. Of course, it doesn't mean one can't use eggnog for genomes, hence I find it necessary to apologise for my previous statement.  

---

### Answer by username_4 on 2018-07-21. Score: 1

Both of the annotation tools you used are designed for prokaryotic genome annotation. I would not expect them to work very well for viruses.

> **Comments:**
> **username_0**: You can choose between Bacteria, Archea or Viruses when using prokka. So, I think it should work for viruses too.  
> **username_1**: @LRJoshi viruses (especially small ones) tend to have overlapping and nested ORFs: something a standard prokaryotic/eukaryotic gene detector (e.g. Glimmer, Genemark, Prodigal) is unlikely to account for.  
> **username_5**: Prokka can do viruses, but you may want to change which translation table it uses to get better blast results  

---

