Scientists have made a major breakthrough in the accuracy and speed at which often deadly pathogen infections can be identified and treated. This is time that, in many cases, is critical to saving a patient’s life.
Scientists at the Ulsan National Institute of Science & Technology (UNIST) in South Korea have unveiled their novel diagnostic technique known as fluorescence in situ hybridization (FISH), using artificial polymers – peptide nucleic acid (PNA) – that act as probes to bind to different genetic sequences within bacteria. When the two probe molecules bind to the target, fluorescent signals are emitted, which essentially reveal the fingerprint of different pathogens.
“The fluorescence in situ hybridization (FISH) technique allows the rapid detection and identification of microbes based on their variation in genomic sequence without time-consuming culturing or sequencing,” the scientists noted. “However, the recent explosion of microbial genomic data has made it challenging to design an appropriate set of probes for microbial mixtures. We developed a novel set of peptide nucleic acid (PNA)-based FISH probes with optimal target specificity by analyzing the variations in 16S ribosomal RNA sequence across all bacterial species.”
UNIST
In the lab, FISH detected seven species of bacteria that commonly infect humans – Klebsiella pneumoniae, Proteus mirabilis, Bacillus subtilis, Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus. It proved to be more than 99% accurate for all but S. aureus – a pathogen that infects skin tissue – which had a still-high success rate of 96.3%. Some detections were as high as 99.9%.
“Detection [is] based on Förster resonance energy transfer (FRET) between pairs of adjacent binding PNA probes eliminated crosstalk between species,” the scientists said. “Rapid sequential species identification was implemented, using chemically cleavable fluorophores, without compromising detection accuracy. Owing to their outstanding accuracy and enhanced speed, this set of techniques shows great potential for clinical use.”
While it may sound (kind of) simple, the effectiveness of the system belies the work that has gone in to develop it. The team analyzed 19,885 known genetic sequences of 14,614 species of bacteria, then designed species-specific PNA sequences to target the bug’s ribosomal RNA. PNA probes have distinct advantages over traditional DNA probes and can travel through the pathogen’s cell walls. Having the two probes bond to the same pathogen improves accuracy, particularly in cases where illness has been caused by several pathogens.
And FISH can produce these results in less than three hours. Conventional diagnostic tools – blood tests and polymerase chain reaction (PCR) analyses can take days to weeks. For patients battling time-critical, life-threatening infections such as sepsis, it’s critically important to diagnose and treat as soon as possible.
The quick turnaround with results can also help avoid patients being administered incorrect antibiotics.
“This method will aid in the diagnosis of infections requiring immediate antibiotic treatment, such as sepsis, urinary tract infections, and pneumonia, while also helping to reduce unnecessary antibiotic usage,” said first author Dr. Sungho Kim, from the Department of Biomedical Engineering at UNIST.
Now, the team will conduct more testing on blood samples taken from sick patients to assess how well FISH can perform in a clinical setting.
The research was published in the journal Biosensors and Bioelectronics.
Source: UNIST