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WGS Better Defines Staph Transmission in ICU; TB First Candidate for Routine Clinical Use of NGS

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A team led by researchers at the Royal Sussex County Hospital in Brighton, England, has shown that whole-genome sequencing of Staphylococcus aureus isolates from patients in the intensive care unit detects patient-to-patient transmissions more accurately than conventional typing methods.

The study, conducted mostly in 2010 and published online last month in Clinical Infectious Diseases, served as a pilot project for sequencing-based typing of S. aureus in a hospital environment. As part of a UK-wide consortium, the Brighton team is currently exploring ways to implement microbial whole-genome sequencing in routine clinical practice, starting with tuberculosis.

Over the last few years, hospital infections with methicillin-resistant S. aureus, or MRSA, have decreased significantly in the UK as a result of screening and other control measures. However, infections with methicillin-sensitive strains, or MSSA, remains high, and patient-to-patient transmissions were assumed to be the cause.

"We were not convinced that was necessarily what was happening, given that everyone was trying very hard to break chains of infections," Martin Llewelyn, a researcher at the Brighton and Sussex Medical School and one of the senior authors of the study, told In Sequence, prompting him and his team to take a closer look.

"The problem was that the existing typing schemes made that very difficult to do," he said, because MRSA strains in particular often look very similar, so their origin cannot be traced precisely.

For their study, the researchers used both conventional approaches — spa-typing, which is a targeted sequencing method, and patient stay data — and whole-genome sequencing.

Over a 14-month period, they screened more than 1,100 patients admitted to the intensive care unit of their hospital for S. aureus, and again every week after admission.

They cultured S. aureus from more than 300 patient samples and analyzed a total of 275 isolates by both spa-typing and whole-genome sequencing, the latter conducted by an off-site laboratory in Oxford on the Illumina HiSeq.

Forty-four patients acquired S. aureus while in the ICU. For the 37 isolates analyzed from these patients, whole-genome sequencing revealed that about a fifth were transmissions from other patients.

Conventional methods falsely identified three transmissions as being from one patient to another when whole-genome sequencing showed they were not, and they failed to detect two disease acquisitions and four transmissions that sequencing did discover.

"The resolution and the granularity that you get from typing down to base pair differences gives us much better insights into who is getting Staph aureus from whom," said John Paul, a consultant microbiologist at the Royal Sussex County Hospital, and the other senior author of the study. "And you just can't do that with conventional methods."

Overall, sequencing provided a more clear-cut picture than conventional typing. "If you look at the similarities of or differences between the isolates [using sequencing], they're either very similar, in which case you can really say they are related to one another, or they're very far apart," Llewelyn said. "There is very little ambiguity. There are very few isolates that are somewhere in the middle."

Having established that only a fifth of Staph infections result from transmission between patients, the researchers are now in the midst of a study exploring other possible transmission routes, for example, from staff members, visitors, or the food chain. Alternatively, patients may have already been colonized with S. aureus when they were admitted but this was not picked up by the initial screening test.

Last year, the hospital acquired its own MiSeq instrument and plans to do a number of near real-time studies of S. aureusand other infectious agents for surveillance and outbreak investigations, Paul said, in collaboration with other groups.

Whether whole-genome sequencing will be used more routinely in the future for typing S. aureus isolates will depend on decreases in cost and turnaround time and on better automation. "At some point, when it's as cheap to get the bug from the culture plate and sequence it as it is to identify it and do the antibiotic susceptibility testing and typing by conventional methods, it's kind of a no-brainer that you get much superior information through whole-genome sequencing," Paul said.

Right now, sequencing consumables per isolate cost on the order of $50, he said, about 10 times as much as a plate-based susceptibility test for S. aureus, and about three times as much as spa-typing.

For Mycobacterium tuberculosis, which causes tuberculosis, on the other hand, "you can already argue that there is a business case for changing over to sequencing," he said. That organism is very slow-growing and needs to be sent out to one of a small number of reference laboratories in the UK for reliable susceptibility testing and typing, which is expensive.

"TB stands out as something where you can beat conventional methods, because with such a slow-growing organism, to do the susceptibility testing by conventional methods takes many weeks, whereas by whole-genome sequencing, as soon as you got a culture-positive specimen, you can do the extraction and library prep and sequencing and analysis within a couple of days and get back to the clinician," Paul said.

A project called the UK CRC Modernizing Medical Microbiology Consortium that involves researchers at the University of Oxford, the Wellcome Trust Centre for Human Genetics in Oxford, and groups affiliated with the UK's Health Protection Agency in Brighton, Birmingham, Leeds, and London is currently investigating the routine use of whole-genome sequencing for tuberculosis.

According to Derrick Crook, the leader of the consortium and a consultant microbiologist for infectious diseases at the Oxford University Hospitals NHS Trust, the group has been developing both software and workflows for a prototype to implement whole-genome sequencing in TB diagnostics.

It is currently testing this prototype in a feasibility study, results of which are expected in the spring and will allow the consortium to improve the software, automate the test further, develop reporting systems, and work out data models for storing the genomic information and linking it to the clinical information.

By the end of this year, the consortium plans to start a one-year parallel study where whole-genome sequencing will be offered as a routine service for TB diagnosis, along with conventional typing.

That study, which may also involve other laboratories, including in other countries, will provide information on turnaround times and costs, which Crook said are likely going to be "substantially less" than for current TB testing.

"If that goes well, you switch off the old system and you're left with mostly a new system," Crook told In Sequence.

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