Using HantaNet

Analyze new hantavirus sequences using the pre-loaded reference dataset

1. Navigate to https://cdcgov.github.io/HantaNet/

2. Select the segment for the new sequences (Small, Medium or Large) in order to load the reference gene module

3. The reference 2D network is loaded

4. On the main menu, click File and Add Data (options to download the reference metadata (.csv) and alignment (.fas) files are available)

5. Download the aligned FASTA .fas file. Use a multiple sequence alignment tool like MAFFT to align the new sequences against the aligned reference FASTA (Currently, HantaNet doesn't include a multiple sequence alignment tool; new sequences must be aligned against the HantaNet's reference alignment before loading them into HantaNet for classification)

Refer to Sequence Alignment Protocols for instructions on how to use MAFFT to align your sequences to the HantaNet's reference alignment

6. We recommend to use this Metadata Template to create the node file with your new sequence metadata (if available). Save metadata as .csv file before loading to HantaNet (the identifier under the Accession_ID field in the metadata file must match the sequence identifier in the FASTA file)

7. Repeat steps 1-4 to load the new aligned FASTA .fas and metadata .csv files. In the loading page, select Node for the metadata and FASTA for the alignment files

8. Once loaded, click Launch

9. You will see the network which includes your sequences; unpin nodes, re-scale the 2D network and pin nodes again

10. On the main menu, click Settings and adjust the Filtering Threshold (genetic distance) and additional parameters as needed

11. Modify the 2D Network Settings (Nodes, Links, Network, Polygons) as needed

12. On the main menu, click File and Save to save the current view as a .microbetrace session file that can be shared and uploaded into MicrobeTrace or HantaNet

13. Save customized settings as a .microbetracestyle file that can be uploaded onto the Global Settings- Styling menu to apply them on a new HantaNet session

Visualize and analyze phylogenetic trees

You can load a phylogenetic tree in NEWICK format into HantaNet. If you would like to use the reference alignments available in HantaNet with your sequences to build a phylogenetic tree using the method of your choice, make sure the Accession IDs match in both the metadata and Newick files. The example below shows a phylogenetic tree built in MEGA using an M gene alignment of hantavirus reference and new sequences (the alignment was generated following the Sequence Alignment Protocols.)

1. Navigate to https://cdcgov.github.io/HantaNet/

2. Select the module (Small, Medium or Large) in order to load the reference gene module

3. The reference 2D network is loaded

4. On the main menu, click File and Add Data

5. Load NEWICK file, select Newick in the loading page and click Launch.

6. The 2D network default view is loaded.

7. On the main menu, click View and select Phylogenetic Tree.

8. A new tab will open with the phylogenetic tree view. Use the Tree View Settings to customize the view.

9. Continue data analysis or build a dashboard using additional views available in HantaNet (step 7).