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IS-diagnostics Publishes Validation of PCR-Based Clinical Metagenomic MDx

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NEW YORK (GenomeWeb) – Researchers in the Netherlands have published a validation of an automated PCR-based bacterial metagenomic molecular diagnostic method from IS-diagnostics.

An analysis of 66 clinical samples published last week in the Journal of Clinical Microbiology showed the open molecular approach was able to find more bacterial species than routine culture.

Specifically, in samples from sites without an endogenous microbiota, IS-pro had identical results to culture in 30 percent of samples. The group at VU University Medical Center in Amsterdam and Maastricht University Medical Center showed the method also detected more species than culture in 47 percent of samples, and five of the 10 culture-negative samples in the cohort were positive using IS-pro.

Founded in 2011 by microbiologists at the VU University Medical Center — including two of the authors of the JCM study — IS-diagnostics' core technology is a eubacterial PCR assay that detects a region between 16S and 23S ribosomal DNA.

But the group has been working on the IS-pro technique for more than six years, CEO Dries Budding, who is also a clinical microbiologist at VU, told GenomeWeb. It has also published eight peer-reviewed studies on the method.  

"Our aim was to make an assay in which clinicians do not need to decide in advance what bacteria they want to look for," said Budding.

"I'm also a clinician myself, and a lot of what we do in regular clinical routine is qPCR," he explained. With that method a laboratorian needs to have targets in mind, and he or she is limited by the number of PCRs that can be done in a given amount of time and by what reagents are available in qPCR format.

However, the length of the 16S to 23S region is species-specific, Budding said, so, "you don't need to sequence the region, all you need to do is sort by fragment length," to identify specific bacteria.

The length is typically between 200 and 1,000 nucleotides — a good range for PCR — and most bacteria have multiple alleles of ribosomal DNA with lengths that tend to vary within chromosomes of the same species, Budding said.

The IS-pro method involves a two-step process of PCR followed by capillary gel electrophoresis.

The method also uses color labels for phyla important in humans — for example Proteobacteria like Salmonella, Campylobacter, and Escherichia coli, as well as Firmicutes, Actinobacteria, Fusobacteria, Verrucomicrobia, and Bacteroidetes. "So, you can make the distinction by color, and then by length you can really zoom in to the species level," Budding said.

IS-diagnostics has also built a software pipeline so that the profiles are automatically uploaded and peak profiles are translated to a list of bacterial species present in the sample.

In all, the technique takes five hours to run, and can accommodate any PCR system and CE instrument. Runs per day would be limited by a lab's existing instrumentation, but Budding said about 100 per day with a 24-sample CE system is reasonable.

The firm is in the process of obtaining CE-IVD marking, and expects that to be completed in about one month. In the meantime, production of the kits will begin next week, Budding said.

IS-diagnostics is also on a trajectory to seek US Food and Drug Administration clearance of IS-pro, a process it will probably initiate next year, Budding said.

The first kit is called the molecular culture kit, and it is for use on clinical samples such as abscesses, cerebrospinal fluid, or any kind of material derived from a site that doesn't have an endogenous microbiota.

In continued use of the kit in their own work as researchers, Budding and his team have found that about 30 percent of samples that go out as culture-negative actually contain bacteria.

"So, there's actually a big gain in doing PCR analysis on these samples," Budding said.

The price has not been finalized yet, and it is early stages of production, but currently the cost is €150 ($162) per patient sample.

While this is likely more expensive than routine culture, it may be less expensive than 16S Sanger sequencing in terms of the cost of technician time, Budding said. That method is also less sensitive, and it only works well for samples containing a single species of bacteria. Next-generation sequencing would also work, but it is not currently economically viable for most clinical labs.

Another competing technology, the CE-marked Iridica platform from Abbott, takes a similar metagenomic approach, and can also identify viruses and fungi. However, Iridica requires special and expensive instrumentation to do extraction and mass spectrometry-based detection, and cannot process high numbers of samples per day.

IS-diagnostics will now continue clinical studies of specific sample types, Budding said. Furthermore, the technique also works on samples with an endogenous microbiota."The whole microbiota diagnostic field is something that is very promising, [so] one of our big efforts is in finding clinical applications to use the technique there," he said.

The firm is also looking into industry partnerships, particularly for distribution, but Budding said it is wary of limiting the method to a single platform.

"One of the nice things about our technique is that it is really applicable in different settings [and] that makes the whole approach very versatile."

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