Up next for MALDI-TOF mass spec: AFB, molds

Speaking at the ASM Microbe 2016 conference in June, John Branda, MD, cautioned that databases’ up-to-date taxonomy means “they’re going to report identifications that may be unfamiliar—unfamiliar to you in the laboratory or unfamiliar to the end users.”

Labs need to figure out how to address that. Otherwise, “You can create confusion,” says Dr. Branda, associate director of clinical microbiology, Massachusetts General Hospital, and assistant professor of pathology, Harvard Medical School.

He suggests using the newest taxon first (for example, Peptoniphilus asaccharolyticus) followed by the older, more familiar taxon in parentheses (in this case, Peptostreptococcus).

He also suggests creating a translation table. If, for example, a MALDI result identifies Streptococcus gallolyticus, subspecies gallolyticus, it’s critical to report to the subspecies level. “We know that the various subspecies within the Strep bovis group have different clinical implications in terms of their likelihood to be related to colonic neoplasms,” Dr. Branda says. But in addition to alerting clinicians to the subspecies, “You also don’t want the clinician to miss the fact that this is a bovis.” Since they may be unfamiliar with the new name, Dr. Branda recommends putting “Strep bovis” in parentheses. Making note of that in a translation table—“If the MALDI says this, this is how we report it”—can provide uniformity in reporting.

In other cases, however, a confusing name may be nothing more than confusing. In the case of an Enterobacter kobei, for instance, it may be reasonable to assume clinicians will not know that the result points to an Enterobacter cloacae complex organism. In such a case, providing both the unfamiliar species name and the parenthetical information may be overkill, if members of the broader group carry the same clinical implications. “You may decide that if any of these species is identified, you’re going to translate that as E. cloacae complex.”

A translation table may also help the laboratory deal with pitfalls. In some situations, the MALDI-TOF may not reliably distinguish between two closely related species—say, Achromobacter xylosoxidans and denitrificans. Given that uncertainty, “We’ll just report Achromobacter species in most cases,” Dr. Branda says. In cases where a more specific ID is critical, such as a patient with cystic fibrosis, the lab will do supplemental testing to distinguish them.—Karen Titus

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Molds are tricky, agrees Anna Lau, PhD. MALDI has been live for molds at the National Institutes of Health since 2012; its research-use-only database was published in 2013, says Dr. Lau, co-director of bacteriology, parasitology, and molecular epidemiology, Department of Laboratory Medicine, Clinical Center.

For laboratories that plan to use MALDI-TOF for molds, a little education is in order, says Dr. Lau. “In mycology, we’re used to seeing common organisms like Aspergillus and Penicillium,” she says. With MALDI-TOF identification, “Now we’re seeing a lot of the strange, or rare, organisms.” An added complication is the constantly changing taxonomy and nomenclature brought about by advances in sequencing technology. Laboratories adopting species-specific identification should include comments, where relevant, in the clinical report, Dr. Lau suggests. For example, after the new organism name is listed, note, in brackets, “previously known as” (or words to that effect) so clinicians will know what to refer to in the literature in terms of therapy guidance. “Or add a comment such as: ‘This organism is related to this particular complex, with demonstrated resistance to a particular drug,’” she says.

Dr. Lau says any laboratory with a mass spectrometry instrument should be able to adopt MALDI-TOF for mold identification. Whether smaller labs will is, for now, up in the air. “I would love for this to be universal,” Dr. Lau says. But not every laboratory is the NIH. Those without large resources will find it much easier to use an FDA-approved database—a step that hasn’t yet happened—than develop their own. “Mold databases from manufacturers are certainly available for purchase as a research-use-only tool,” she says. Alternatively, in-house–developed databases such as the NIH mold database are freely available through transfer agreements.

With molds, fears of abandonment—of old ways as well as employees—might be less acute than they’ve been with yeast or AFB, at least at the NIH. Dr. Lau and her colleagues still correlate MALDI results with the morphological or phenotypical findings. “Mass spec isn’t always perfect,” she says. But it has major advantages in being able to provide identification for sterile molds; that is, those that refuse to produce any morphologically identifiable structures.

Because mass spec provides results faster, “Our staff is freed up to explore other avenues of testing including research and expansion of the test menu. It depends on the institution,” she says.

What has to happen for Dr. Lau’s dream of universality to come to pass?

Clinical validation, for starters. Only one such study has looked at the Vitek system comprehensively for molds. “So there has to be a lot more performance reviews in the clinical setting.”

Even though the Biotyper has been the subject of more clinical validation studies, Dr. Lau notes the paucity of data on following the manufacturer’s guidelines and instrument performance. “On the Biotyper, most of the data has been based on in-house–developed databases. As beautiful as that is for clinical mycology in terms of moving diagnostics forward,” that doesn’t address two problems, she says: lack of standardization and lack of availability of those research databases.

And with two systems available, a comparison study might be in order. “There’s only been one study to date that looked at a large number of organisms, but very few of these test organisms were molds. And so a rigorous comparison in the clinical setting of both systems” would be helpful, she says.

Mold identification, moreover, has some technical challenges. Extraction methods are particularly complicated, Dr. Lau says, and can vary according to media (liquid or solid), sample type (entire mold versus spores), and so on. All give different spectra, Dr. Lau says, and therefore different levels of performance, which in turn are dependent on the database and its coverage.

Dr. Lau is finishing up a large study comparing the NIH and Bruker mold databases across eight U.S. academic medical centers. The results are somewhat disorienting, she says, with wide variability in performance, which has not been the case with bacteria and yeast.

In trying to pluck answers from the data, Dr. Lau hints at several possibilities.

Instrument settings and variability certainly have something to do with it, she says. But the fact that other organisms haven’t been affected by these differences means there’s likely even more going on. Dr. Lau suspects that the answer has something to do with the concentration of protein in fungi. “It’s harder to extract proteins for them,” she says. “The extraction process is so much more complex than with yeasts.”

Despite these mysteries, Dr. Lau remains optimistic about the potential for using MALDI-TOF for mold identification. The NIH database has been shared with about 82 laboratories worldwide so far. “The feedback that we’re getting from some of the labs that are now using it routinely, in a day-to-day setting, has been great, and we continue to expand the database with new MSPs.”

“I’m hoping that more labs will soon have the capacity to bring this in-house and make it a real-time clinical test,” she says.
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Karen Titus is CAP TODAY contributing editor and co-managing editor.