Mayo Clinic Laboratories is one of the few laboratories in the country to offer two special sequencing-based tests for bacteria in its catalog. The first is Bacterial Typing by Whole Genome Sequencing (BTWGS), a test designed to help protect patients by investigating potential outbreaks of a single bacterial species or by identifying recurrent infections in an individual patient. The second is the Broad Range Bacterial PCR and Sequencing (BRBPS) test, which has the ability to detect DNA of any bacteria to help identify the cause of an infection by testing a specimen type such as tissue or fluid.
Discriminating between two or more bacterial isolates
Nicholas Chia, Ph.D., biophysicist and Co-Director of Mayo Clinic’s Center for Individualized Medicine (CIM) Microbiome Program, is a visionary in applying microbial whole-genome sequencing (WGS) in clinical microbiology. Because infectious outbreaks are an increasing threat in health care facilities and beyond, he urged CIM leadership to support development of bacterial WGS a few years ago, in order to accurately and quickly identify the spread of germs in health care facilities.
"All biology really only makes sense in the light of evolution,” says Dr. Chia. “To investigate [using the bacterial lab test] if a microbe that infects a patient today is related to a microbe that infected someone else two days ago, or if it’s the same microbe but with a few small differences, you want to track those differences. What you’re really tracking is the history of that microbe. And if you find in that history that, yes, all of these microbes are related, now you’ve very quickly identified an outbreak."
With the BTWGS test, the genomic sequences of individual microbes are identified and compared to the genomic sequences of the other co-submitted bacterial isolates. Clients then receive a diagram illustrating the results. “It’s a graphic showing how closely related bacteria from different patients are,” says Robin Patel, M.D., Chair of Mayo Clinic’s Division of Clinical Microbiology, Co-Director of the Bacteriology Laboratory, and Director of the Infectious Diseases Research Laboratory.
Not so long ago, if you sent in your bacterial strain to the Centers for Disease Control and Prevention (CDC), “you’d have to wait around two months for them to figure out what was going on,” says Dr. Chia. This is no longer the case, now that the CDC, like Mayo, is using the much faster WGS technology.
Dr. Chia continues: “Let’s say you get infected with Staphylococcus aureus today, and let’s say a week later you’re better but then you get infected again. And that leaves your doctors wondering, ‘Okay, is this person really unlucky? Or is there an abscess somewhere that keeps coming back? Is there a strain he or she is picking up in-house?’
"Before [WGS] technology came along, even with just those two infections, we wouldn’t really be able to know the answer. But now, we can take the genome sequences from both of those and compare them and know if they’ve had passage of the same bug. This is because we’re basically looking at the fossil record of those organisms."
Other uses in the future
BTWGS has great potential to mitigate threats of infectious diseases before they become mass outbreaks. The integration of WGS data with epidemiological analysis in nosocomial (hospital-borne) infections identifies common pathways of transmission onwards, such as communal areas, bays, shared patient equipment, health care staff, or visitors interacting with each other and other patients.
For example, in a collaborative study, Mayo used BTWGS as a molecular typing tool in its Neonatal Intensive Care Unit (NICU) to investigate a number of cases of methicillin-resistant Staphylococcus aureus (MRSA).
“NICU babies are a vulnerable population because they’re all together in one room, then you have health care workers who come in and pick them up, and it’s just an ideal place for an infection to spread,” says Dr. Chia. “So we used this technology to monitor both the babies and the health care workers over three years [2014–2017], and we actually identified some health care workers who were passive carriers of MRSA.”
Given, there are sensitive hurdles yet to be overcome for such monitoring practices to become commonplace in hospitals and other facilities, like nursing homes, that also have vulnerable populations.
“If we start monitoring people, workers might wonder, ‘What does this mean? Does it mean I can’t come to work if I’m infected? Will I have to change my job?’ and so on,” says Dr. Chia. “In our study, the health care workers could be decolonized.”
By detecting and decolonizing the health workers who carried Staphylococcus aureus, the monitoring study, in turn, helped protect babies in the NICU from being infected. Again, while this sort of monitoring practice might strike people as controversial today, Dr. Chia believes this is the way of tomorrow.
“Because of the way the dynamics of infectious diseases work, we are at any moment close to some really big outbreaks,” he says. “And we’ve seen this happen across the last few years, where we’ve had a number of infectious disease outbreaks around the world. I think, once we use the technology this way, the world is going to change for the better. We demonstrated the ability to implement this, to be proactive. The question is, how do we do it universally?”
Going beyond traditional microbiology to identify infections
The second bacterial test in Mayo Clinic Laboratories’ catalog, BRBPS, brings a new standard to microorganism identification. The test can help detect and identify bacteria from normally sterile sites when traditional microbiology testing methods may fail to yield a diagnosis.
“There are certain bacteria that are difficult or impossible to grow by traditional culture, and so they need a molecular method for detection and identification,” says Audrey Schuetz, M.D., Director of the Initial Processing Laboratory and Co-Director of the Bacteriology Laboratory. “Or, if a bacterium is present in a patient but has been suppressed to low levels by an antibiotic, then the bacterium might not be able to proliferate in high enough numbers to be grown on culture plates.”
If a bacterium fails to show up in a culture, but the health care provider still suspects an infection, an affected specimen may be tested with BRBPS. Or, the test can be done directly from the patient specimen, without first trying to grow the organism in culture. By cutting out the time-consuming step of culturing, results can be provided faster than culture in many instances. First, the polymerase chain reaction (PCR) assay looks for a gene that’s common across bacteria and amplifies it by generating more copies to establish a good, strong signal. If a PCR assay on a patient sample comes up negative, the result is available in a matter of hours. If the PCR assay is positive, then the specimen is sequenced to identify exactly what kind of bacterium it has, which takes a bit longer. In most cases, results are available the next day.
“Once the health care provider knows what kind of bacterium it is, he or she can use appropriate antibiotics to treat that specific infection,” says Dr. Schuetz.
The BRBPS is a popular test at both Mayo Clinic and Mayo Clinic Laboratories because it can detect any kind of bacteria and can do so quickly. “We understand the ‘need for speed,’ and we’re now working on making this test even faster than it is today,” says Dr. Patel.
Diversity of uses
BRBPS can be used on many different types of specimens, such as fresh tissue or biopsies from a lymph node, joint, heart valve, brain, viscera, organ, lung, or bone. Other specimen types might include normally sterile body fluids aspirated from such areas as a patient’s joint or around the spinal cord, heart or lung, abdomen, or an abscess.
“It’s an especially useful assay because we can apply it to so many different kinds of diseases, and we can run it on so many different specimen types,” says Dr. Patel. “A lot of our molecular tests in the past could only look for one type of bacteria at a time, like Staphylococcus aureus, for example. But if that test didn’t find this specific microorganism, it wasn’t all that helpful. Time and money were wasted and no diagnosis was established. In contrast, [BRBPS] can look for any kind of bacteria. It’s what we describe as ‘agnostic’ or ‘non-biased’ in terms of bacteria.”
Speaking of agnostic, the assay can also be used on specimens from a paraffin-embedded tissue block, taken from normally sterile sites in the body like the brain, heart valves, organs, and joints.
“At Mayo, we keep tissue block samples indefinitely,” says Dr. Patel. “What that means is, if there’s ever a question after a procedure, about whether there might be a bacterium causing a patient’s disease, we can go back into those tissue blocks and look for bacteria.”
In other words, a tissue block can be tested in lieu of putting the patient through the discomfort, time, and cost of another surgery or procedure to obtain a biopsy.
Dr. Patel continues: “Being able to go back to a patient’s [paraffin-embedded] biopsy can be helpful when entertaining diagnoses with unusual bacteria, even when those tissue blocks aren’t current, and a mystery disease wasn’t solved in the past,” she says. “This scenario is especially helpful if there isn’t any remaining fresh tissue or fluid from a patient to use for this test. We don’t want to have to take a patient back to surgery just to get a new specimen.”
Helping patients from all corners of the globe
Both the BRBPS and BTWGS tests demand a high level of expertise and are labor intensive. And while these tests are reasonably priced to have done, the infrastructure (e.g., setup, instrumentation, staff) required to offer them is a significant investment. For these reasons, Mayo Clinic Laboratories is currently one of the few diagnostic laboratories in the country to offer both tests, according to Dr. Patel.
“I think that’s why there are a lot of health care facilities sending us their specimens for testing,” she says. “It takes a substantial amount of work to be able to offer these kinds of tests. And so it’s probably easier for a smaller laboratory to send us their small numbers of specimens rather than trying to implement these tests themselves. At the end of the day, we want to help clinicians investigate and diagnose the many bacterial infections out there, so they can appropriately treat and heal their patients.”