Emerging Microbes & Infections

2020, VOL. 9
https://doi.org/10.1080/22221751.2020.1732836

LETTER

Gonococcal urethritis caused by a multidrug resistant Neisseria gonorrhoeae

strain with high-level resistance to spectinomycin in China
Shao-Chun Chena,b*, Li-Hua Huc*, Xiao-Yu Zhua,b and Yue-Ping Yina,b
a
Institute of Dermatology, Chinese Academy of Medical Sciences & Peking Union Medical College, Nanjing, People’s Republic of China;
b
National Center for STD Control, China Center for Disease Control and Prevention, Nanjing, People’s Republic of China; cZhejiang Provincial
Institute of Dermatology, Deqing, People’s Republic of China

ABSTRACT
We report a recent (2018) gonorrhoeal urethritis caused by a multidrug-resistant Neisseria gonorrhoeae strain in China. The
isolated N. gonorrhoeae strain from a male and female pair expressed high-level resistance to spectinomycin (SPC),
azithromycin, and other antibiotics but was sensitive to ceftriaxone. The SPC high-level resistance (MIC = 2048 mg/L)
was due to a small deletion in rspE that caused two amino acid changes in ribosomal protein S5.

ARTICLE HISTORY Received 4 January 2020; Revised 12 February 2020; Accepted 15 February 2020

KEYWORDS Neisseria gonorrhoeae; spectinomycin; resistance; urethritis; treatment failure

Neisseria gonorrhoeae has become resistant to all anti- Pharmaceutical, Shandong, China) intramuscularly
biotics previously used for gonorrhoea treatment, com- and 100 mg doxycycline (Yung Shin Pharm. Ind. Co.,
promising our ability to effectively control gonococcal Ltd, Jiangsu, China) orally twice a day for five days.
infections. In China, ceftriaxone (CRO) 250 mg or As his urethral discharge was not resolved and his
spectinomycin (SPC) 2 g is used as first-line monother- wife had pain and discharge at her urogenital tract,
apy treatment options for urogenital gonorrhoea [1]. he and his wife visited the same STD clinic 10 days
However, an increasing level of decreased susceptibility later after his first visit. A urethral swab from the
of gonococcal strains to CRO and clinical resistant male patient and an endocervical swab from his wife
strains to third-generation cephalosporins (cefixime were collected and cultured on Thayer–Martin selec-
and CRO) have been reported in recent years [2]. tive media (BioMérieux, Shanghai, China). Although
Due to such resistance, SPC is frequently used in few leukocytes were found from both samples by
empiric therapy of gonorrhoea in many Chinese clinics microscopy, results from a nucleic acid amplification
[3]. Importantly, SPC resistance expressed by gono- test (Acon, Zhejiang, China) on both samples were
cocci isolated in China is rare making SPC an appropri- positive for N. gonorrhoeae. Considering the possible
ate replacement antibiotic for the treatment of SPC treatment failure may have occurred at the first
cephalosporin-resistant cases of gonorrhoea. We now visit of the male patient, an increased dose of SPC 4 g
describe a gonorrhoeal urethritis case caused by a intramuscularly was used to treat the couple. A tele-
high-level SPC-resistant strain that is also high-level phone follow-up one week after treatment revealed
resistant to multiple antibiotics. that they both were asymptomatic, but a follow-up
The patient with an SPC-resistant gonorrhoeal test of cure was not performed.
infection was a heterosexual male in his early 40s. He The bacterial samples from the infected male (pre-
presented at Zhejiang Provincial Institute of Dermatol- and post-treatment isolates) and female (pre-treatment
ogy in August 2018 with urethral discharge and dysuria isolate) were transferred to the reference laboratory at
two days after unprotected sex with a female sex the National Center for Sexually Transmitted Disease
worker. A urethral swab was collected for laboratory Control (NCSTDC), Chinese Center for Disease Con-
diagnosis of gonorrhoea and Gram-negative diplococci trol and Prevention. N. gonorrhoeae species were
were observed within leukocytes by microscopy. He confirmed for all three isolates by Gram stain and a
was diagnosed to have gonorrhoea and empirically carbohydrate (glucose, maltose, lactose, sucrose pre-
treated with a single dose of 2 g of SPC (Lukang pared at NCSTDC) utilization test. The agar dilution

CONTACT Yue-Ping Yin yinyp@ncstdlc.org; Shao-Chun Chen chensc@ncstdlc.org Institute of Dermatology, Chinese Academy of Medical
Sciences & Peking Union Medical College, 12 Jiangwangmiao Street, Nanjing 210042, People’s Republic of China; National Center for STD Control, China
Center for Disease Control and Prevention, 12 Jiangwangmiao Street, Nanjing 210042, People’s Republic of China
*These authors contributed equally to this work.
© 2020 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group, on behalf of Shanghai Shangyixun Cultural Communication Co., Ltd
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted
use, distribution, and reproduction in any medium, provided the original work is properly cited.
518 S.-C. Chen et al.

Table 1. Phenotypic and genetic characteristics of three N. gonorrhoeae isolates with high-level resistance to SPC identified in
China.
MIC (mg/L, resistance) Genotype Resistant determinants
Ribosomal protein 5S
Strains SPC AZM PEN CRO TET CIP PPNG MLST porB tbpB NG-MAST (coded by rpsE) 16S rRNA 23S rRNA
Male pre-treatment isolate 2048 >256 >256 0.015 >64 4 Yes 7822 4 75 ST304 27 V deletion WT A2059G
(R) (R) (R) (S) (R) (R) K28E alteration
Male post-treatment isolate 2048 >256 >256 0.015 >64 4 Yes 7822 4 75 ST304 27 V deletion WT A2059G
(R) (R) (R) (S) (R) (R) K28E alteration
Female pre-treatment isolate 2048 >256 >256 0.015 >64 4 Yes 7822 4 75 ST304 27 V deletion WT A2059G
(R) (R) (R) (S) (R) (R) K28E alteration
Notes: aSPT, spectinomycin; AZM, azithromycin; PEN, penicillin; CRO, ceftriaxone; TET, tetracycline; CIP, ciprofloxacin; PPNG, penicillinase-producing
N. gonorrhoeae; ST, sequence type; NGMAST, N. gonorrhoeae multi-antigen sequence type; MLST, multilocus sequence type; V, valine; K, lysine; E, glutamic
acid; A, alanine; G, glycine; WT, wild type. bSusceptibility (S) and resistance (R) were determined based on breakpoints from the European Committee on
Antimicrobial Susceptibility Testing (http://www.eucast.org/clinical_breakpoints). cReference gene (GenBank accession no. KC311362.1) numbering in the
ribosomal protein 5S of N. gonorrhoeae was used.

method was used to determine the antibiotic suscepti- 16S rRNA gene, but had a deletion of three nucleotides
bility of the gonococcal isolates, which included SPC, (TTA) in the rpsE gene at position 80–82 compared
azithromycin (AZM), penicillin (PEN), tetracycline with that of N. gonorrhoeae reference strain FA1090
(TET), ciprofloxacin (CIP) and CRO (USP Reference (GenBank accession no. AE004969.1). Three nucleo-
Standard, Rockville, MD, US). Production of penicilli- tide deletion in rpsE resulted in a deletion of amino
nase was assessed using a nitrocefin solution filter acid 27 (V) and a K28E amino acid alteration in the
paper test [4]. All three isolates had the same antimi- ribosomal protein S5. A nucleotide blast search found
crobial susceptibility profile. Briefly, the strains were the rpsE genes of all three isolates were identical to a
high-level resistant to SPC (MIC = 2048 mg/L), AZM high-level SPC-resistant strain previously reported by
(MIC > 256 mg/L), PEN (MIC > 256 mg/L, penicilli- Unemo (GenBank accession no. KC311362.1) [8].
nase-positive), TET (MIC > 64 mg/L) and resistant to SPC has been introduced and proved to be effective
CIP (MIC = 4 mg/L), but susceptible to CRO (MIC = for the treatment of urogenital and anorectal gonor-
0.015 mg/L) according to susceptibility and resistance rhoea for nearly 50 years. Although the discontinuation
breakpoints from the European Committee on Antimi- of SPC manufacturing has restricted its use to a few
crobial Susceptibility Testing (Table 1). countries including China, it is still an alternative anti-
Multilocus sequence typing (MLST) [5], biotic for patients with a cephalosporin or PEN allergy.
N. gonorrhoeae multi-antigen sequence typing (NG- WHO also recommended SPC as single therapy based
MAST), [6] and N. gonorrhoeae sequence typing for on the local antimicrobial susceptibility surveillance
antimicrobial resistance (NG-STAR) [7] were per- data [9]. In China, SPC is readily accessible and rec-
formed to identify the sequence type and resistant ommended by national treatment guideline and widely
determinants. All three isolates were assigned to used for gonococcal infections in China. In a nation-
MLST7822 and NG-MAST ST304. The NG-STAR wide survey of antibiotic use, 7.9% of 2121 physicians
type was a new ST, which contained a type 43 non- use SPC as the first option for gonorrhoea treatment
mosaic penA allele (penA 43.002) as well as previously [3]. The PK/PD data of SPC shows that the antimicro-
described mutations in other chromosomally located bial effect of SPC is concentration-dependent and Cmax
genes associated with antibiotic resistance: an A del- can reach 100 mg/L 1 h after 2 g dose intramuscular
etion in the mtrR promoter as well as a missense injection. When Cmax/MIC is between 8 and 10 folds,
mutation at codon 45 (G45D) in mtrR, two missense the SPC treatment is considered to be effective. As
mutation in porB resulting in PorB amino acid changes the MIC of the resistant strains are as high as
(G120K) and 121 (A121D), a missense mutation in 2048 mg/L, higher doses of SPC within a safe level or
ponA at codon 421 (L421P), missense mutations as CRO should be considered as an appropriate treatment
gyrA codons 91 (S91F) and 95 (D95A), a missense to effectively control spreading of the SPC-resistant
mutation in parC (S87R), and A2059G in 23S rRNA. clone in China and beyond. Although SPC resistance
The identical sequence types between pre- and post- may emerge quite quickly when it is routinely used
SPC treatment samples obtained from the male for gonorrhoea treatment, the emergence and trans-
suggested a probable SPC treatment failure. Further, mission of a high-level SPC-resistant clone, which
because the partner samples were identical using seemed to have occurred with the male and female
these tests, it is likely that a transmission of this resist- pair studied herein, were sporadic worldwide [8,10]
ant strain occurred between the couple. To determine and has not heretofore been reported in China since
the genetic basis for SPC resistance in the isolates we 2012. Notably, the isolates reported here were only sen-
sequenced the 16S rRNA and rpsE genes as previously sitive to CRO and expressed resistance to multiple anti-
described [8]. All three isolates contained a wild-type biotics including AZM. Although uncommon, the dual
 Emerging Microbes & Infections 519

SPC and AZM resistance expressed by the strains Dis. 2018 Apr;24(4):804–806. DOI: 10.3201/eid2404.
described in this report coupled with their possible 171817. PubMed PMID: 29553336; PubMed Central
spread might compromise the dual therapy strategy PMCID: PMCPMC5875255.
[3] Han Y, Yin YP, Zhou Y, et al. Nonadherence to
recommended by WHO in cases where AZM and national guidelines for antibiotic treatment of uncom-
CRO fail to cure the infection [9]. Accordingly, a plicated gonorrhea in China: results from a nationwide
strengthened and continuous national surveillance in survey. Sex Transm Dis. 2018 Sep;45(9):600–606. DOI:
China is necessary to detect the spread of SPC-resistant 10.1097/OLQ.0000000000000819. PubMed PMID:
isolates of N. gonorrhoeae. 30102261.
[4] Unemo M, Ballard R, Ison C, et al. Laboratory
diagnosis of sexually transmitted infections,
Ethics including human immunodeficiency virus. Geneva:
World Health Organization; 2013. [cited 10 Feb
The project has been approved by the Medical Ethics 2015]. Available from: http://wwwwhoint/
Committee at the Institute of Dermatology, the Chi- reproductivehealth/publications/rtis/9789241505840/
nese Academy of Medical Sciences & Peking Union en/
[5] Jolley KA, Maiden MC. BIGSdb: scalable analysis of
Medical College, and the National Center for Sexually bacterial genome variation at the population level.
Transmitted Disease Control (NCSTD) at Nanjing BMC Bioinf. 2010 Dec;11:595. DOI: 10.1186/1471-
(approval number 2014-LS-026). 2105-11-595. PubMed PMID: 21143983; PubMed
Central PMCID: PMCPMC3004885.
[6] Martin IM, Ison CA, Aanensen DM, et al. Rapid
Acknowledgements sequence-based identification of gonococcal trans-
mission clusters in a large metropolitan area. J Infect
We are grateful to Zhejiang provincial institute of dermatol-
Dis. 2004 Apr;189(8):1497–1505. DOI: 10.1086/
ogy for providing the isolates and making this study possible.
383047. PubMed PMID: 15073688.
We would like to thank William Shafer (Emory University)
[7] Demczuk W, Sidhu S, Unemo M, et al. Neisseria gonor-
for his valuable comments.
rhoeae sequence typing for antimicrobial resistance, a
novel antimicrobial resistance multilocus typing
scheme for tracking global dissemination of
Disclosure statement
N. gonorrhoeae strains. J Clin Microbiol. 2017
No potential conflict of interest was reported by the May;55(5):1454–1468. DOI: 10.1128/jcm.00100-17.
author(s). PubMed PMID: 28228492; PubMed Central PMCID:
PMCPMC5405263.
[8] Unemo M, Golparian D, Skogen V, et al. Neisseria gonor-
Funding rhoeae strain with high-level resistance to spectinomycin
due to a novel resistance mechanism (mutated ribosomal
The work was supported by grants from the Chinese Acad- protein S5) verified in Norway. Antimicrob Agents
emy Medical Sciences Initiative for Innovative Medicine Chemother. 2013 Feb;57(2):1057–1061. DOI: 10.1128/
[grant number 2016-I2M-3-021] and Jiangsu Natural AAC.01775-12. PubMed PMID: 23183436; PubMed
Science Foundation [grant number BK20171133]. Central PMCID: PMCPMC3553714.
[9] World Health Organization. WHO guidelines for the
treatment of Neisseria gonorrhoeae; 2016.
References [10] Boslego JW, Tramont EC, Takafuji ET, et al. Effect of
[1] Wang QQ, Zhang GC. Guidelines for diagnosis and spectinomycin use on the prevalence of spectinomycin-
treatment of sexually transmitted diseases. Shanghai: resistant and of penicillinase-producing Neisseria gonor-
Shanghai Science and Technology Press; 2007. Chinese rhoeae. N Engl J Med. 1987 Jul;317(5):272–278. DOI:
[2] Chen SC, Yin YP, Chen XS. Cephalosporin-resistant 10.1056/nejm198707303170504. PubMed PMID:
Neisseria gonorrhoeae clone, China. Emerging Infect 2955222.