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New Paper-Based Platform can Rapidly Detect Antibiotic-Resistant Bacteria

Image created by Dr. Michael J. Miller

Researchers have developed a paper-based platform that could help quickly detect the presence of antibiotic-resistant, disease-causing bacteria.

One of greatest challenges facing the world is the rise of disease-causing bacteria that are resistant to antibiotics. Their emergence has been fuelled by the misuse and overuse of antibiotics, the researchers said.

According to the World Health Organisation (WHO), a handful of such bacteria -- including E. coli and Staphylococcus aureus -- have caused over a million deaths, and these numbers are projected to rise in the coming years.

Timely diagnosis can improve the efficiency of treatment, the researchers said.

"Generally, the doctor diagnoses the patient and gives them medicines. The patient then takes it for 2-3 days before realising that the medicine is not working and goes back to the doctor," said Uday Maitra, a professor at the Indian Institute of Science (IISc) Bangalore.

"Even diagnosing that the bacteria is antibiotic-resistant from blood or urine tests takes time. We wanted to reduce that time-to-diagnosis," Maitra said in a statement.

The latest research, published in the journal ACS Sensors, addressed this challenge by developing a rapid diagnosis protocol that uses a luminescent paper-based platform to detect the presence of antibiotic-resistant bacteria.

There are different ways by which a bacterium becomes resistant to antibiotics. In one, the bacterium evolves, and can recognise and eject the medicine out of its cell.

In another, the bacterium produces an enzyme called beta-lactamase, which hydrolyses or breaks down the beta-lactam ring -- a key structural component of common antibiotics like penicillin and carbapenem -- rendering the medication ineffective.

The approach developed by researchers at the IISc and Jawaharlal Nehru Centre for Advanced Scientific Research (JNCASR) in Karnataka involves incorporating biphenyl-4-carboxylic acid (BCA) within a supramolecular hydrogel matrix containing terbium cholate (TbCh).

This hydrogel normally emits green fluorescence when ultraviolet (UV) light is shined on it.

"In the lab, we synthesised an enzyme-substrate by tethering BCA to the cyclic (beta-lactam) ring that is a part of the antibiotic. When you mix this with TbCh hydrogel, there is no green emission as the sensitiser is ‘masked,’" said Arnab Dutta, a PhD student at IISc, and lead author of the research paper.

“In the presence of beta-lactamase enzyme, the gel will produce green emission. Beta-lactamase enzyme in the bacteria is the one that cuts open the drug, destroys, and unmasks the sensitiser BCA. So, the presence of beta-lactamase is signalled by green emission," Dutta said.

The luminescence signals the presence of antibiotic-resistant bacteria, and the intensity of the luminescence indicates the bacterial load, the researchers said.

For non-resistant bacteria, the green intensity was found to be extremely low, making it easier to distinguish them from resistant bacteria, they said.

The next step was to find a way to make the technology inexpensive. Currently used diagnostics instruments are costly, which drives up the price for testing.

The team collaborated with a Tamil Nadu-based company called Adiuvo Diagnostics to design a customised, portable and miniature imaging device, named Illuminate Fluorescence Reader.

Infusing the hydrogel in a sheet of paper as the medium reduced the cost significantly. The instrument is fitted with different LEDs that shine UV radiation as required, the researchers said.

Green fluorescence from the enzyme is captured by a built-in camera, and a dedicated software app measures the intensity, which can help quantify the bacterial load, they said.

The team then analysed their approach on urine samples.

“We used samples from healthy volunteers and added pathogenic bacteria to mimic urinary tract infections. It successfully produced the outcome within two hours," Maitra added.

The researchers now plan to tie up with hospitals to test this technology with samples from patients. 

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