Reflex Detection of Ciprofloxacin Resistance in Neisseria gonorrhoeae by Use of the SpeeDx ResistancePlus GC Assay

Lee, Darren Y. J.; Ashcroft, M; Chow, Eric P F; Sait, Michelle; Petra, Vesna De; Tschaepe, Marlene; Lange, Sigrid; Taiaroa, George; Bradshaw, Catriona S; Whiley, David M.; Fairley, Christopher K; Howden, Benjamin P; Chen, Marcus Y; Pasricha, Shivani; Williamson, Deborah A

doi:10.1128/jcm.00089-21

Cited by 15 publications

(13 citation statements)

References 12 publications

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“…These findings suggest that we should test the effect of gonococcal tolerance on the acquisition of resistance to other antimicrobials before we conclude that tolerance does not play a role in the genesis of AMR. It would be particularly interesting to test antibiotics such as CIP for which resistance mutations occur more easily and rapidly in vitro than CRO [ 45 ]. This could be achieved either with crossover E-tests or via cyclic antibiotic exposure experiments.…”

Section: Discussionmentioning

confidence: 99%

Tolerance to Ceftriaxone in Neisseria gonorrhoeae: Rapid Induction in WHO P Reference Strain and Detection in Clinical Isolates

et al. 2022

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In addition to antimicrobial resistance, bacteria contain other mechanisms to survive antibiotic exposure such as tolerance, defined as the ability to slow metabolism by the extension of the lag phase without altering antimicrobial susceptibility. In a number of bacterial species, tolerance has been associated with treatment failure and infection chronicity and is found to precede and facilitate antimicrobial resistance. It is unknown if tolerance can be induced in Neisseria gonorrhoeae. In this study, we determined if tolerance to ceftriaxone (CRO) can be induced in N. gonorrhoeae and detected in clinical isolates. To induce tolerance, WHO P N. gonorrhoeae reference strain samples were grown under daily 3 h intermittent CRO exposure (10× the MIC), partitioned by overnight growth in GC broth. This cyclic exposure was performed for 7 consecutive days in sextuplicate, with two control cultures to which GC medium without antibiotics was added. To detect tolerance and assess CRO susceptibility, modified Tolerance Disc (TD) and Epsilometer tests were performed on isolates after each CRO exposure cycle. Additionally, this experiment was carried out on 18 clinical N. gonorrhoeae isolates. Tolerance was first detected after two CRO exposure cycles in five out of six samples. The phenotype differed per cycle with no clear pattern. No tolerance was found in control samples but was detected in 10 out of 18 clinical isolates. The present study is the first to demonstrate the induction of tolerance to CRO in N. gonorrhoeae through antibiotic exposure. In addition, tolerance to CRO was found in clinical samples.

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Section: Discussionmentioning

confidence: 99%

Tolerance to Ceftriaxone in Neisseria gonorrhoeae: Rapid Induction in WHO P Reference Strain and Detection in Clinical Isolates

et al. 2022

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“…The main mutations conferring fluoroquinolone resistance are on the subunits of the DNA gyrase A, GyrA, and topoisomerase IV, ParC [ 33 ]. At the molecular level mutations conferring resistance are mainly at nucleotides C272 (amino acid S91) and A284 (amino acid D95) positions of gyrA and at the A259 (amino acid S87) location of the parC [ 28 , 32 – 34 ].…”

Section: Discussionmentioning

confidence: 99%

“…This notion is especially facilitated by the fact that the ciprofloxacin resistance mechanisms utilized by N . gonorrhoeae are well defined and there are clear targets to be selected for developing rapid molecular tests [ 28 , 29 ]. To this end, to conserve the usage of current first line antibiotics and in considering re-use of previously favored first line antibiotics, ciprofloxacin is an ideal candidate.…”

Section: Discussionmentioning

confidence: 99%

“…In conclusion, this ciprofloxacin isolate panel has been extensively characterized based on its ciprofloxacin AST profile and the gyrA and parC genes sequences. This panel should be useful for developing ciprofloxacin susceptibility related tests or for quality assurance purposes [ 27 , 28 ]. Availability of such tests for detecting ciprofloxacin susceptibility can provide clinicians with tools to improve antibiotic stewardship, potentially reducing costs, and save first line drugs for individuals that truly need it.…”

Section: Discussionmentioning

confidence: 99%

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Characterization of a Neisseria gonorrhoeae Ciprofloxacin panel for an antimicrobial resistant Isolate Bank

et al. 2022

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Objectives Neisseria gonorrhoeae (gonococcus) infection is one of the most commonly reported nationally notifiable conditions in the United States. Gonococcus has developed antimicrobial resistance to each previously used antibiotic for gonorrhea therapy. However, some isolates may be still susceptible to no longer recommended, yet still effective antibiotics. This in turn suggests that targeted therapy could slow resistance development to currently recommended empirical treatments. We curated a gonococcal Ciprofloxacin Antibiotic Resistance Isolate Bank panel (Cipro-panel) as a tool for validating or developing new tests to determine ciprofloxacin susceptibility. Method The Cipro-panel was selected using whole genome sequencing, bioinformatic tools, and antimicrobial susceptibility testing (AST) data. Isolates were further selected based on nucleotide variations in gyrA and parC genes. Results We selected 14 unique N. gonorrhoeae isolates from the 2006–2012 Gonococcal Isolate Surveillance Project (GISP) collection. They represented a wide range of antimicrobial susceptibility to ciprofloxacin and commonly observed nucleotide variations of gyrA and parC genes. This Cipro-panel consists of 5 isolates with resistant phenotypes (MIC > = 1 μg/mL), 8 isolates with susceptible phenotypes (MIC < = 0.06 μg/mL), and 1 isolate falling in the Clinical and Laboratory Standards Institute defined intermediate range. Among the gyrA variations we observed a total of 18 SNPs. Four positions had nonsynonymous changes (nucleotide positions 272, 284, 1093, and 1783). The first two positions (272 and 284) have been linked previously with resistance to ciprofloxacin (i.e. amino acid positions 91 and 95). For the parC gene, we observed a total of 21 possible SNPs. Eight of those SNPs resulted in non-synonymous amino acid changes. One location (amino acid 87) has been previously reported to be associated with ciprofloxacin resistance. Conclusions This Cipro-Panel is useful for researchers interested in developing clinical tests related to ciprofloxacin. It could also provide additional choices for validation, quality assurance purposes and improve antibiotic usage.

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“…The ResistancePlus GC test consists of a multiplex qPCR, incorporating channels for the wild-type (WT) gyrA 91S allele, the mutant gyrA 91F allele, the N. gonorrhoeae opa gene, the N. gonorrhoeae porA gene, and a PCR positive control. Across multiple studies, the ResistancePlus assay detects gonorrhea with a sensitivity of 94-100% 8,18,19 and specificity of 99-100% 8,19 . The test is also highly sensitive (>99%) and specific (>97%) for ciprofloxacin resistance 18 .The ResistancePlus GC assay has been approved in Europe and received a break-through designation from the FDA 20 .…”

Section: Introductionmentioning

confidence: 99%

Neisseria gonorrhoeae diagnostic escape from a gyrA-based test for ciprofloxacin susceptibility can lead to increased zoliflodacin resistance

Rubin

Mortimer

Grad

2022

Preprint

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The etiologic agent of gonorrhea, Neisseria gonorrhoeae, has become resistant to each of the first line antibiotics used to treat it. However, most isolates in the US remain susceptible to ciprofloxacin, leading to the hypothesis that a diagnostic that accurately reports susceptibility to ciprofloxacin may allow its reintroduction into clinical use. One diagnostic approach is to identify codon 91 in gyrA, where coding for the wildtype serine is associated with ciprofloxacin susceptibility and phenylalanine with resistance. Here, we found that escape from this diagnostic method can occur through gyrA F91S reversion and expansion of pre-existing ciprofloxacin non-susceptible N. gonorrhoeae lineages with gyrA 91S. Furthermore, we found that mutations conferring ciprofloxacin resistance in gyrA 91S isolates led to cross-resistance with zoliflodacin, an antibiotic in phase 3 trial for treatment of gonorrhea. These findings define N. gonorrhoeae mutations that should be monitored via surveillance systems, particularly in populations and regions deploying gyrA 91-based diagnostics, and demonstrate how diagnostics that guide antibiotic therapy can have unintended consequences.

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Reflex Detection of Ciprofloxacin Resistance in Neisseria gonorrhoeae by Use of the SpeeDx ResistancePlus GC Assay

Cited by 15 publications

References 12 publications

Tolerance to Ceftriaxone in Neisseria gonorrhoeae: Rapid Induction in WHO P Reference Strain and Detection in Clinical Isolates

Tolerance to Ceftriaxone in Neisseria gonorrhoeae: Rapid Induction in WHO P Reference Strain and Detection in Clinical Isolates

Characterization of a Neisseria gonorrhoeae Ciprofloxacin panel for an antimicrobial resistant Isolate Bank

Neisseria gonorrhoeae diagnostic escape from a gyrA-based test for ciprofloxacin susceptibility can lead to increased zoliflodacin resistance

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