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Network-based visualization of ARG-microbe associations in ResistoXplorer

Network-based visualization of ARG-microbe associations in ResistoXplorer
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Hi, and welcome back. In this video, we’ll see how we can explore the known ARG microbial host associations through a network based approach in ResistoXplorer. So let’s quickly open a browser and go to the Home page of ResistoXplorer. Now click on the first module section, named ARG List. This brings us to the gene list upload page where I can enter or paste the list of ARGs of interest with their optional fold change or abundance value. This list can be of significant ARGs that are detected in differential abundance testing or ARGs identified through high throughput PCR. Next, we have to select a target database that we want our genes to be searched against.
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The first three primary reference databases are dedicated to antibiotic resistance genes, while BacMet have reference database for bio sites and metals resistance genes and their functional annotations. Lastly, we have a database of antimicrobial peptide resistance genes too. Now, just to make things clear, I’m going to use an example antibiotic resistance gene list. As you can see, we have a list of 14 genes and their corresponding abundance values. Let’s upload this example list to search it against ResFinder database by clicking the Submit button.
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If everything is OK, we will see the interaction table page. Here, we can see, on the top, the stats about how many ARGs uploaded and how many mapped to the selected database, and also how many total number of associations and number of unique microbial hosts found for the uploaded ARGs. While in the bottom, we have the results that are represented as an association table which each row corresponding to a particular reference ARG and its potential microbial host. When available, the table also provides other association information along with hyperlinks to the corresponding gene bank accession number and PubMed literature.
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We can also directly remove each row by clicking the Delete icon in the last column to keep only high-quality associations supported by literature or experimental evidence. That, this ARG host associations, are used to build the networks. Now let’s click on the Proceed button to go to the network builder page. As you can see here on the top, we have the statistics of notes and edges to have an overview of the size and complexity of the generated network. Since not all the nodes will be connected, this approach may lead to generation of multiple networks.
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As we can see here in the bottom, that we have three networks, one big with most of the queries for ARG, and two small networks with one query each, along with statistics of nodes and edges for each. These networks are small. But in case of big networks, one can also philtre the nodes based on their topological measures, such as the degree and betweenness, for better interpretation. Now let’s click on the Proceed button to visualise this generated networks. So this, now, is how the network visualisation system in ResistoXplorer looks like.
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As you can see, it is comprised of three main components, the central network visualisation area, the network customization and functional analysis explorer panel on the left, and node table on the right. In network, the circular node represents the ARG, while the rectangular node represents the microbe, while the association between the ARG and microbe is shown by match. We can intuitively visualise and manipulate the network using a mouse with a scroll wheel. For example, we can scroll the wheel to zoom in and out of the network, or hover the mouse over any node to view its name, or click a node to display its details on the bottom right corner, or even double-click a node to select it.
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The toolbar, to the top, exhibits basic functions to manipulate the network. The first is the colour picker. That enables us to choose a highlight colour for the next selection, followed by some basic functions such as zoom in, zoom out, reset, move up and down, or to the right and left. By using the dashed square icon in the toolbar, we can also select and drag multiple nodes. Next, the network customization panel provides us various options to configure the general visualisation features of the default network or to specify the range of mouse operation. The layout option enables us to perform automatic network layout using different algorithms. The background option enables us to select between a white or black background.
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We can also vary the range of mouse operation during highlighting and dragging and dropping, using the scope option. For example, in single node mode, only the node which has been clicked or dragged, will be highlighted or affected. Whereas all the neighbour nodes, along with the selected node, will be affected in the node neighbours mode. Additionally, we can save the current network in different formats or download the network file in GraphML format for visualisation in other software. Also, we can perform enrichment analysis of the ARGs present within the current network at different functional category or level, using Hypergeometric tests from the function explorer panel.
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By clicking on a row of the result table, we can highlight all nodes related to an enriched function within the network. Lastly, the node table on the right panel displays all the ARGs and microbial hosts, along with corresponding network topological measures such as degree and betweenness. Nodes with high degree and betweenness value are important nodes in the network, as nodes with the highest degree of centrality will act as a hub of the network, while a node with the highest betweenness centrality controls the flow of information in that network. The optional abundance values, if we have provided while uploading, will be presented in the last column.
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We can directly click on any row of interest, select, and the network field will automatically zoom to the related node. The panel in the bottom provides detailed information related to the nodes being highlighted or currently selected on the network. There are also several other options and functionalities present in the system which you can explore on your own by uploading your own list of ARGs or use example lists present in ResistoXplorer. This is it for this video. Bye-bye. Take care, and happy exploration.

In this video, you will learn how to intuitively explore, manipulate and customize the ARG-microbial host association networks in ResistoXplorer. We recommend you follow along on your own computer and try to answer some of the questions below in the exercise section.

Hands-on Exercise:

Go to the home page of ResistoXplorer and click on the “ARG List” module. Try exploring the ARG-microbial hosts hosts’ associations with your own list of genes or example dataset. Then, based on it, answer or discuss the following points:

  • What important biological insights can you gain from such network-based exploration of ARG-host associations?
  • What is its relevance in the context of antimicrobial resistance?
  • What do you think are the limitations of such an approach?
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Exploring the Landscape of Antibiotic Resistance in Microbiomes

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