To the Editor: Spread of multidrug-resistant Neisseria gonorrhoeae is a major public
health concern. Effective antimicrobial therapy is a key element in gonorrhea control.
However, N. gonorrhoeae has developed resistance to multiple classes of antimicrobial
drugs, including β-lactams, tetracyclines, and fluoroquinolones (
1
–
3
). Even an extended-spectrum oral cephalosporin-resistant, cefixime-resistant N. gonorrhoeae
has emerged, and cefixime has now been withdrawn from use in Japan. Best practice
treatment is limited to injectable extended-spectrum cephalosporins, such as ceftriaxone
and spectinomycin. The emergence of ceftriaxone-resistant N. gonorrhoeae threatens
effective disease control.
We identified a novel ceftriaxone-resistant N. gonorrhoeae isolated from a 31-year-old
female commercial sex worker; MIC of ceftriaxone for this isolate was high (2 µg/mL).
The woman visited a clinic in Kyoto for a routine examination for sexually transmitted
infections in January 2009. Although she had no obvious symptoms or signs, a throat
sample collected on her first visit yielded a positive result for N. gonorrhoeae by
the strand displacement amplification test (ProbeTec ET, Becton Dickinson, Franklin
Lakes, NJ, USA), but a vaginal sample taken at the same time was negative. After 2
weeks, another throat sample was positive for N. gonorrhoeae when cultured on Thayer-Martin
medium, and the patient subsequently received 1 g ceftriaxone intravenously. Her pharyngeal
sample was also N. gonorrhoeae positive by strand displacement amplification test
on the third visit 2 weeks later, and further ceftriaxone treatment was prescribed.
However, a culture for test of cure was not conducted because reinfection was considered.
A negative result was finally obtained in April 2009.
The culture showed positive reactions in oxidase and catalase tests. Gram staining
showed gram-negative diplococci. The ID-test HN-20 Rapid system (Nissui, Tokyo, Japan)
classified the bacterium as N. gonorrhoeae. Susceptibility was determined by the agar
dilution method (
4
). For this strain, named H041, MIC of ceftriaxone was high (2 µg/mL), and the strain
was highly resistant to penicillin G (4 µg/mL), cefixime (8 µg/mL), and levofloxacin
(32 µg/mL). However, it demonstrated susceptibility to spectinomycin (16 µg/mL) and
reduced susceptibility to azithromycin (0.5 µg/mL).
To characterize the ceftriaxone-resistant N. gonorrhoeae H041, multilocus sequence
typing characterized the strain as ST7363 (
5
), which is the predominant sequence type (ST) among cefixime-resistant clones (
6
). N. gonorrhoea multiantigen sequence typing (NG-MAST) was also performed (
7
). The NG-MAST strategy uses 2 genes, por and tbpB, for porin and a transferrin-binding
protein, respectively. NG-MAST indicated that the strain H041 was ST4220 and contained
the por2594 allele and the tbpB10 allele. NG-MAST 4220 is a novel ST. However, the
tbpB10 allele is the most frequently observed allele (76.5%) among multilocus sequence
typing-ST7363 N. gonorrhoeae strains (n = 81) (M. Ohnishi, unpub. data).
Molecular typing suggested that the novel ceftriaxone-resistant N. gonorrhoeae, H041,
is closely related to the ST7363 cefixime-resistant N. gonorrhoeae. Therefore, we
compared SpeI-digested genomic DNA banding patterns of strain H041 with those of other
N. gonorrhoeae strains by using pulsed-field gel electrophoresis as described (
8
). Four ST7363 strains, including N. gonorrhoeae H041, and 4 ST1901 strains (another
major ST among cefixime-resistant N. gonorrhoeae strains) (
6
) were analyzed. The banding pattern of SpeI digested H041 genomic DNA was similar
to that of other ST7363 strains and indistinguishable from that of cefixime-resistant
but ceftriaxone-susceptible NG0207 (Figure).
Figure
Pulsed-field gel electrophoresis patterns of ceftriaxone-resistant Neisseria gonorrhoeae
strain H041 and other multilocus sequence typing (MLST) ST7363 and ST1901 strains.
SpeI-digested genomic DNA from ceftriaxone-resistant N. gonorrhoeae H041, 3 of the
MLST ST7363 strains and 4 of the MLST ST1901 strains were analyzed by pulsed-field
gel electrophoresis. A lambda ladder standard (Bio-Rad, Hercules, CA, USA) was used
as a molecular size marker.
We describe the emergence of ceftriaxone-resistant N. gonorrhoeae, isolated from a
pharyngeal specimen from a female commercial sex worker. At 2 µg/mL, the MIC was 4-fold
higher than that of the previously reported ceftriaxone nonsusceptible strain (
9
). Our susceptibility testing suggests that only azithromycin and spectinomycin are
effective drugs for treating this strain. In this case, eradication was successful,
although N. gonorrhoeae colonization of the pharynx may just be tempory because the
pharynx is not an ideal site for N. gonorrhoeae growth. From the routine examinations
of commercial sex workers during January–March 2009, 40 N. gonorrhoeae were isolated
in the clinic, but no other ceftriaxone-resistant strains were isolated. There is
no evidence of dissemination of this strain in Kyoto.
Three independent molecular subtyping methods indicated that the ceftriaxone-resistant
H041 strain was N. gonorrhoeae, and it might originate from an ST7363 cefixime-resistant
N. gonorrhoeae clone. There are several possible mechanisms for the acquisition of
resistance, including formation of a new mosaic type penA allele as penA-X cefixime
resistance and acquisition of an extended-spectrum β-lactamase gene. The H041 strain
did not produce β-lactamase in a nitrocephin test. Further molecular analysis is needed
to elucidate the precise mechanism of the ceftriaxone resistance of the H041 strain.
The emergence of ceftriaxone-resistant N. gonorrhoeae raises concerns for controlling
gonorrhea because ceftriaxone is widely recommended and the first-line treatment for
gonorrhea around the world. N. gonorrhoeae has a potential to gain an extraordinarily
high MIC to ceftriaxone. Surveillance for ceftriaxone-resistant N. gonorrhoeae should
be strengthened.