Study gauges impact of genotyping on gonorrhea treatment

His researcher’s review of more than 1,000 isolates and about 10 different studies found that the summary of the wild-type gyrA to predict fluoroquinolone susceptibility was 98 percent sensitive and nearly 99 percent specific.

Dr. Klausner’s group and Mark Pandori, PhD, formerly of the San Francisco Public Health Laboratory, developed in 2006 a reverse transcription PCR assay to study fluoroquinolone susceptibility. Using different probes and melting points generated from that process, “we were able to clearly differentiate a resistant isolate from a susceptible isolate.”

Their early data looked at the MIC of about 100 different N. gonorrhoeae isolates to ciprofloxacin, and the excellent differentiation by their gyrA assay, Dr. Klausner said, showed a very high range of MICs in the wild-type and mutant isolates (Siedner MJ, et al. J Clin Microbiol. 2007;45[4]:1250–1254). “We had one missed call where the organism was determined to be wild type but actually was resistant,” he said.

Phase one of the study Dr. Klausner reported on at the AMP meeting was assay verification. They used urine triplicates seeded with wild-type or mutant gyrA isolates at a concentration of 1,000 CFU/mL, from the CDC and the University of Washington. “We found no difference in the performance across the different triplicates,” Dr. Klausner said. This phase included cross-reactivity studies because of a primer concern that there could be cross-reaction with other common Neisseria species in the throat. “We did not see that. The primers were developed to be targeted toward Neisseria gonorrhoeae, not other Neisseria species” (Hemarajata P, et al. J Clin Microbiol. 2016;54[3]:​805–808).

In phase two of the study, Dr. Klausner and colleagues introduced the assay for routine clinical use at UCLA Health, which serves more than 500,000 patients at two hospitals and in more than 150 primary care clinics and other outpatient settings. “In November 2015, we began to do reflex gyrA genotyping on all Neisseria gonorrhoeae nucleic-acid–positive clinical specimens at the UCLA Health microbiology laboratory.” GyrA results were reported to providers in the electronic health record within 24 to 48 hours of receipt of the specimen. The report showed the positive N. gonorrhoeae result and the presence or absence of the wild-type gyrA gene. A standard disclaimer said the gyrA assay was not FDA approved but had good performance results in accordance with CLIA requirements.

The study’s objectives were to examine the impact of gyrA genotyping on N. gonorrhoeae treatment at UCLA Health and to evaluate patient outcomes among those with a wild-type gyrA genotype. In a retrospective review portion of the study, Dr. Klausner’s team examined records for all laboratory-confirmed cases of N. gonorrhoeae between Jan. 1, 2015 and Sept. 8, 2017. They also collected test-of-cure data among patients with wild-type N. gonorrhoeae infections treated with ciprofloxacin.

“During this 32-month study, there were more than 500 Neisseria gonorrhoeae infected patients,” Dr. Klausner said. Some patients had multiple anatomic site-specific infections or repeat infections. Forty percent of infected patients were treated empirically on about the same day. “That is one limitation to the assay,” he said, since standard practice is to treat gonorrhea immediately before obtaining test results if a patient is symptomatic.

For patients who were not empirically treated, the average time from specimen collection to treatment was up to five days, which provides a good window for performing reflex testing and sending results to the provider to inform treatment choices, he said.

“When we looked at 655 infections that underwent gyrA genotyping, we had results for 43 percent that were wild type, 27 percent were mutant, and about 30 percent were indeterminate,” Dr. Klausner said. The reason for the high number of indeterminate infections is complicated, he added, and may have to do with the site of specimen source, either the rectum or pharynx. “We still see that difficulty,” he tells CAP TODAY. “Newer assays are somewhat better, but because of lower bacterial burden in the pharynx, or competition from other Neisseria species in the pharynx, the ability of the assay to characterize the gyrA gene is decreased.” Research is underway on the molecular assays’ performance to predict susceptibility so that they are equally successful, independent of anatomic site, “because we know that Neisseria gonorrhoeae infections in the throat can be spread to sex partners, and we also know that Neisseria gonorrhoeae infections in the throat serve as an important reservoir where drug resistance is acquired.”

One of the study’s primary outcomes showed a decline in ceftriaxone use for treatment of N. gonorrhoeae infections from 94 percent before gyrA genotyping to 76 percent after gyrA genotyping. “What replaced that ceftriaxone treatment, surprisingly, was ciprofloxacin,” Dr. Klausner said. “Over time, we had a nice increase in the use of ciprofloxacin among those who were nonempirically treated.”

A retrospective test-of-cure study of 25 patients with gyrA wild-type infections treated with ciprofloxacin showed a 100 percent cure rate, regardless of anatomic site. “The sample size is small but quite encouraging.”

To sum up, the study found that routine gyrA genotyping can be implemented in a large health system and that gyrA results can have an impact on the treatment of patients with N. gonorrhoeae. Ceftriaxone use declined with a rise in ciprofloxacin use, and the test of cure was 100 percent (Allan-Blitz LT, et al. Clin Infect Dis. 2017;64[9]:​1268–1270). It was a single-center study, further implementation and replicaton of the gyrA assay are needed, and there was no measure of the ecologic impact of the reduction in ceftriaxone use on Neisseria gonorrhoeae in the population, Dr. Klausner said, citing the study’s limitations.

“For me, it was important to show that this could be done,” he tells CAP TODAY. He isn’t certain his team’s assay is the best. “But it is a laboratory-developed method that is widely available, and any laboratory can follow the published protocol and replicate it.”

Final clinical validation is underway as phase three. It’s an NIH-funded clinical study of the assay with 240 N. gonorrhoeae gyrA wild-type culture-positive patients treated with 500 mg of ciprofloxacin. “We completed enrollment at the end of December. The results look promising,” Dr. Klausner says. “We will be able to produce a precise estimate of the efficacy of ciprofloxacin in the treatment of Neisseria gonorrhoeae infections, with predicted susceptibility as determined by the gyrA molecular assay.”

He expects genotypic testing to be a topic of discussion among groups updating gonorrhea treatment guidelines. “This is an important topic for which now there is new evidence. Molecular susceptibility testing can enable physicians to select specific antibiotics,” he says.

SpeeDx (Sydney, Australia) received the CE mark and the Australian Therapeutic Goods Administration approval for its assay to predict ciprofloxacin susceptibility in N. gonorrhoeae, Dr. Klausner says. Shield Diagnostics (San José, Calif.) announced in March the launch of its Target-NG, a rapid molecular test for antibiotic susceptibility in N. gonorrhoeae. The company says Target-NG can determine if a given gonorrhea infection is susceptible to ciprofloxacin with the same turnaround time as gonorrhea screening tests. (Dr. Klausner has advised various test manufacturers, among them Shield Diagnostics, SpeeDx, Cepheid, Hologic, and Click Diagnostics.)

Advances in molecular testing are key to stemming the growth of antimicrobial resistance, Dr. Klausner said. “New genotypic Neisseria gonorrhoeae diagnostics are here—highly accurate and predictive of susceptibility and treatment outcomes. And better, multi-targeted tests integrated with Neisseria gonorrhoeae detection and additional antibiotics are forthcoming.” 

Amy Carpenter Aquino is CAP TODAY senior editor.