When a patient is diagnosed with a bacterial infection, medical staff need to identify the species attacking the body before it can be treated. The problem is, this process can take a few days and may require highly-specialized equipment that's not available in many hospitals. Now, microparticles and biomolecules are teaming up to form a new dynamic duo, which could help speed up and simplify the process.
Developed by researchers from the Institute of Physical Chemistry of the Polish Academy of Sciences (IPC PAS), the technique uses bioconjugates – complexes created by combining microparticles and biomolecules. In this case, the biomolecules are made up of the T4 bacteriophage, a virus that attacks the E. coli bacteria. The microparticles, meanwhile, are both fluorescent and magnetic – properties that are very important for the job at hand.
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"We started by searching for inexpensive, commercially available microparticles that met our requirements," says Marta Janczuk-Richter, one of the study's authors. "It turned out that appropriate particles were already available on the market - and exactly the ones we were looking for! Their surface was covered with just those chemical functional groups we needed to place virtually any type of bacteriophage on them."
The bioconjugates used are made up of magnetic, fluorescent microparticles (red) with two or three bacteriophages (green) attached to each of them
When the bacteriophages and microparticles are bound together, the resulting bioconjugate is claimed to be the first to have three distinct functions: they glow, they're magnetic, and they bind to just one species of bacteria. Together, those functions allow the bioconjugate to streamline the task of testing a sample for the presence of a specific bacteria.
First, a small amount of the bioconjugates are added to a diluted fluid sample from a patient, or even a food product, like fruit juice. Due to the bacteriophages, the bioconjugates will seek out the E. coli bacteria. After letting the mixture sit for a short time, the bioconjugates can be retrieved with a magnet – thanks to the magnetic microparticles – and separated from the rest of the sample.
That bioconjugate precipitate is then re-diluted and run through a flow cytometer, a relatively common and inexpensive piece of equipment that's often used to conduct blood tests. In this device, the solution is forced to flow through a nozzle so narrow that the larger particles and cells move through it single-file. The resulting stream can be analyzed with lasers and detectors that measure how the light reflects from the individual particles.
"Measurement in the cytometer typically takes about a minute," says Lukasz Richter, one of the study's authors. "The result is a graph on which we see how all the bioconjugates scatter the incident light and emit the fluorescence. Since we know the signal we should obtain from pure bioconjugates, i.e. those without bacteria, we can easily determine whether the sample contains the bacteria we are looking for, and if so, in what concentration."
While the T4 bacteriophage will hunt down E. coli, the researchers say that their technique could be used to detect other types of bacteria, by simply switching which bacteriophage is used to make the bioconjugate. Even if a hospital laboratory needs to sift through a list of possible suspects that could be causing an infection – sepsis, for example, can be the result of a dozen different bacteria – the relative ease and speed of the process means it can be repeated several times as needed, with results still coming in quicker than testing for a single type of bacteria through existing methods, which sometimes involve days-long cultures.
Before the new technique can be used in hospitals though, the team is putting it through real-world tests, that target different bacteria. Then, they plan to develop a kit that different institutions could use to develop their own bioconjugates, according to what they need to test for.
"We are attempting to detect some really dangerous bacteria, like Pseudomonas aeruginosa, in biological samples from rats at various stages of infection," Jan Paczesny, lead researcher on the project, tells New Atlas. "Only when such targeted tests will be validated might the method be introduced. We make the concept really open for people to adjust it to their needs, because we probably don't know of all possible cases and we are not able to prepare all of the needed probes. So we are giving away a toolbox, which might be used to prepare desired bioconjugates depending on specific tasks."
The research was published in the journal Bioconjugate Chemistry.
Source: IPC PAS via EurekalertView gallery - 3 images