On the trail of viruses on their “criminal mission”: Using a new microscopy method, researchers can now track individual pathogen particles live as they search for “possibilities of breaking into” cells. This could provide insights into how certain factors influence the spread of viruses in cell tissues and thus their infection success, the scientists say.
Coronavirus, HIV, flu pathogens and Co: Many different viral pathogens plague mankind and are therefore the focus of science. Researching them is, however, much more tricky than in the case of bacteria. In contrast to these unicellular organisms, viruses are extremely tiny without their own metabolism: virus particles only consist of a protein-covered shell in which genetic material is packed. Infection begins when a virus attaches itself to a cell and enters it. It then causes the cellular machinery to replicate its viral genome and make more virus particles. These offspring then travel again to hijack other cells.
What is the route to the crime scene?
In order to trigger an infection, a virus particle must first diffuse through the protective layers of mucus and outer cells that line our airways and intestines. Although viruses can be easily visualized in and on cells using microscopy techniques, they have not been clearly observed along this important route prior to infection. This is partly because the virus particles, which are tiny compared to the cells, move much faster in free space than inside the cell due to the dynamic processes. "With microscopic examinations, it's like trying to take pictures of a person moving in front of a skyscraper: you can't capture the whole building and the details of the person in front of it in a single image," explains senior author Kevin Welsher from Duke University in Durham. But he and his team have now succeeded in developing a method that enables real-time recordings of viruses as they approach their cellular targets.
They call their process 3D Tracking and Imaging Microscopy (3D-TriIm). It is essentially based on the combination of two microscopes. The first detects a virus marked by fluorescent substances during its rapid movements in the vicinity of living cells. A laser also slews around the glowing virus at high speed to calculate and update its position. While the fine dynamic processes in the interstices and on the cell surfaces move the virus, the slide is continuously and subtly tracked to keep it in focus. While tracking the glowing virus, the second microscope takes three-dimensional images of the surrounding cells. The combination of both data sources then produces a result that is comparable to the impression when navigating with Google Maps, the researchers explain: In addition to the current location during the movement, the terrain, sights and the general situation of the area are also displayed.
Promising visualization
In their presentation video, the researchers show how a specially prepared research virus moves between densely packed human intestinal cells that are located on a microscope slide. They compare the video with the recordings of a surveillance camera: it is like watching a burglar trying to find entry points into a house. The virus comes into contact with a cell for a moment, but then glides along its surface without stopping. "If this were a planned burglary, this would be the stage before the burglar has broken the window," says lead author Courtney Johnson of Duke University. "When I present our research work, I am sometimes asked whether it is a simulation," says the scientist. But then she can proudly explain that these are real microscopic images.
As the team points out, their work demonstrates the potential of their concept. They hope that further developments of the method will allow it to be used in ever more realistic - tissue-like environments in which infections occur. “That is the real aim of this method. We believe that we have now created the fundamental possibilities to do this,” said Welsher. The team is currently working primarily on extending the possible observation times. So far, they can only track a virus for a few minutes before the fluorescent label goes out. "The biggest challenge for us now is to make brighter experimental viruses," says co-author Jack Exell of Duke University.
Source: Duke University, professional article: Nature Methods, doi: 10.1038/s41592-022-01672-3