• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of aacPermissionsJournals.ASM.orgJournalAAC ArticleJournal InfoAuthorsReviewers
Antimicrob Agents Chemother. Nov 2010; 54(11): 4893–4895.
Published online Aug 23, 2010. doi:  10.1128/AAC.00482-10
PMCID: PMC2976121

Characteristics and Dissemination of Mosaic Penicillin-Binding Protein 2-Harboring Multidrug-Resistant Neisseria gonorrhoeae Isolates with Reduced Cephalosporin Susceptibility in Northern Taiwan[down-pointing small open triangle]

Abstract

Among 254 Neisseria gonorrhoeae isolates from a sexually transmitted infection (STI) clinic in northern Taiwan, 69 isolates were found to contain the mosaic penA (MA) gene and were associated with elevated cefixime and ceftriaxone MICs. Most of these MA gene-harboring isolates were also resistant to penicillin (71.4%) and ciprofloxacin (100%) and were from men who have sex with men (MSM) or from bisexual men (81.2%). Three major sequence types (ST835, ST2180, and ST2253) constituted 55.7% of these isolates. The major sequence types harboring the mosaic penA gene may represent major sexual networks responsible for the emergence/introduction and the spread of the multidrug-resistant clones in Taiwan.

Neisseria gonorrhoeae infection continues to be one of the most globally prevalent sexually transmitted infections (STIs) (2). In Taiwan, the incidence of gonorrhea has increased in the last decade, as it has in many other developed countries (4, 5). Recently, however, the number of N. gonorrhoeae isolates with reduced susceptibility to cephalosporin antibiotics has gradually increased (12, 15, 16). Cases of treatment failures with ceftibuten and cefixime, but not with ceftriaxone, have been reported (8, 14). The major mechanisms of N. gonorrhoeae resistance to oral cephalosporins have been determined to be associated with the presence of a mosaic penicillin-binding protein 2 (PBP2) encoded by a penA gene which was generated by homologous recombination with other related commensal Neisseria species (1, 7); resistance is determined by this element, combined with other determinants (17). We explored the antibiotic susceptibilities, risk characteristics, and clonal relatedness of mosaic penA (MA) gene-containing gonococcal isolates in Taiwan.

From June 2006 to July 2009, a total of 273 patients, including 254 males and 19 females, were diagnosed with gonorrhea at the Taipei City Hospital, KunMing Branch, Taiwan. A total of 93.0% (254/273) of the N. gonorrhoeae isolates were collected from male patients (median age, 31 years; range, 17 to 79 years); the remaining 20 isolates were from female patients (median age, 24 years; range, 14 to 34 years). Due to the low percentage of female patients in this population, this study mainly analyzes the male subset of the population with gonorrhea.

Sequencing of the PBP2 gene was conducted to learn the distribution of mosaic penA genotypes among N. gonorrhoeae clinical isolates in Taiwan. Two distinct MA types were identified, and these were categorized into MA1 (68 isolates) and MA2 (1 isolate) types. The translated amino acid sequence from the MA1 gene is identical to that from the MA allele previously identified as mosaic pattern X (GenBank accession no. AB071984) and associated with reduced susceptibility to cephalosporins (7). The amino acid sequence of the MA2 gene is identical to the penA gene uploaded into the NCBI database by Corkill et al. (GenBank accession no. DQ335216), and N. gonorrhoeae isolates with the MA2 gene were recently discovered in San Francisco, CA (10).

The susceptibilities of the 254 isolates to cefixime and ceftriaxone were evaluated by Etest with breakpoints defined as recommended by the Clinical and Laboratory Standards Institute (CLSI) (11). The MICs for cefixime ranged from 0.016 to 0.5 mg/liter, with 5, 26, 21, 15, and 4 isolates having MICs of 0.125, 0.19, 0.25, 0.38, and 0.5 mg/liter, respectively. The distribution of cefixime MICs for the 185 isolates with the nonmosaic penA (NM) gene ranged from 0.016 to 0.094 mg/liter, with the exception of 2 isolates with MICs of 0.125 mg/liter. In contrast, the cefixime MICs for the 69 isolates with the MA gene ranged from 0.125 to 0.5 mg/liter and constituted 97.2% (69/71) of all isolates having MICs of 0.125 to 0.5 mg/liter. The detection of MA alleles in N. gonorrhoeae clinical isolates could be an effective alternative method for the surveillance of isolates with reduced cefixime susceptibility.

The MICs for ceftriaxone ranged from 0.002 to 0.19 mg/liter, with 14 isolates having MICs of 0.125 mg/liter and 1 isolate having an MIC of 0.19 mg/liter. The distribution of ceftriaxone MICs for the isolates with the NM gene ranged from 0.002 to 0.094 mg/liter, with the exception of 4 isolates with MICs of 0.125 mg/liter. In contrast, the ceftriaxone MICs for the isolates with the MA gene ranged from 0.023 to 0.19 mg/liter, with 10 isolates having MICs of 0.125 mg/liter and 1 isolate having an MIC of 0.19 mg/liter. The presence of mosaic PBP2 seemed to have only a partial effect on the reduced ceftriaxone susceptibility. The report of Zhao et al. also showed that the mosaic penA gene was one of several resistance determinants that resulted in the reduced susceptibility to ceftriaxone in N. gonorrhoeae (17). Ceftriaxone is still frequently used for the treatment of gonorrhea in Taiwan. According to the medical records, clinical treatments in this study were successful. However, continuous monitoring of the MIC for ceftriaxone is still critical for the detection of the emergence of ceftriaxone-resistant gonococcal isolates, which may erode the last resort of gonococcal therapy.

The overall rates of resistance to penicillin and ciprofloxacin were found to be 62.2% (158/254) and 88.6% (225/254), respectively, by disk susceptibility tests. The rates of reduced susceptibility of clinical isolates with the MA gene to penicillin and ciprofloxacin were higher than those of isolates with the NM gene (71.0% versus 58.9% and 100% versus 84.3%, respectively) (Table (Table1).1). Therefore, the isolates with the MA gene not only exhibited reduced susceptibilities to cephalosporins but also are more likely to have multiple-drug resistance.

TABLE 1.
Characteristics of male patients infected with isolates categorized by two PBP2 genotypes

Analysis of demographic features of each patient showed that, among the 69 isolates with the MA gene, 56 (81.2%; P < 0.001) were from men who have sex with men (MSM) or from bisexual men and 32 (46.4%; P < 0.001) and 15 (21.7%) reflected coinfection with HIV and syphilis, respectively (Table (Table1).1). In contrast, the isolates with the NM gene were mostly from heterosexual men (67.0%; 124/185), and the proportions of these isolates reflecting concurrent HIV (20/185; 10.8%) and syphilis (18/185; 9.7%) infections were lower than those of isolates with the MA gene. Similar findings involving clinical isolates with reduced cefotaxime susceptibility, mainly obtained from MSM patients, were also reported in the Netherlands (3). The detection of the mosaic penA gene in N. gonorrhoeae may be helpful in identifying core groups for transmission of multidrug-resistant (MDR) gonococci and at high risk of concomitantly contracting HIV and syphilis.

The 69 isolates with the MA gene were divided by Neisseria gonorrhoeae multiantigen sequence typing (NG-MAST) into 29 sequence types (STs). The three major STs were ST2180 (23.2%, 16/69), ST835 (17.4%, 12/69), and ST2253 (15.9%, 11/69), which constituted 55.7% of these isolates. It was discovered that the penA allele of 28 STs was of the MA1 type, and only one isolate, belonging to ST1407, was of the MA2 type. The major ST clusters may represent large sexual networks with clonal transmission of specific genotypes. The clonal relationship of isolates of ST835, ST2253, and ST2180 constructed by pulsed-field gel electrophoresis (PFGE) correlated well with NG-MAST and supported the high-level similarity of strains within each major clonal cluster from a common ancestor (data not shown). ST835 strains with reduced susceptibility to oral cephalosporin and resistance to azithromycin have been discovered in Australia and Hong Kong and in Italy, respectively (8, 11, 13), and ST2180 is closely related to ST835. ST1407, harboring the MA2-type mosaic penA allele, was one of the major STs in Australia, and in this study, we also report the first isolation of ST1407 and MA2 in Taiwan (13).

N. gonorrhoeae clinical isolates harboring the mosaic penA gene were first identified in 2001 by Ito et al. (7). We deduce from a previous report on resistant N. gonorrhoeae strains in Taiwan that this kind of clinical isolate might have emerged in or been introduced to Taiwan between 1999 and 2003 (6). Temporal analysis of the strain replacement dynamic of N. gonorrhoeae with mosaic penA from June 2006 to July 2009 showed that strains of ST835 and ST2180 were predominant in 2006 and were gradually replaced by other STs (ST2253 and ST3082) during 2007 to 2009 (data not shown). Selection pressure exerted by antibiotic usage or host immunity might have contributed to such a transition. One recent report indicates that the horizontal transfer of the mosaic penA from a strain with reduced cefixime susceptibility to a strain susceptible to cefixime was proved by an in vitro test (9). It might explain why the major penA allele, the MA1 type, was widely discovered in different STs in this study besides the closely related major clones. Continuous molecular epidemiology may shed more light on the emergence and dissemination of clinical isolates with higher MICs for cephalosporin antibiotics in Taiwan.

Acknowledgments

This work was supported by grants DOH98-DC-2006 and DOH99-DC-2010 from the Centers for Disease Control, Department of Health and National Research Program for Genome Medicine 98-0324-01-F-12, National Science Council, Taiwan.

Footnotes

[down-pointing small open triangle]Published ahead of print on 23 August 2010.

REFERENCES

1. Ameyama, S., S. Onodera, M. Takahata, S. Minami, N. Maki, K. Endo, H. Goto, H. Suzuki, and Y. Oishi. 2002. Mosaic-like structure of penicillin-binding protein 2 gene (penA) in clinical isolates of Neisseria gonorrhoeae with reduced susceptibility to cefixime. Antimicrob. Agents Chemother. 46:3744-3749. [PMC free article] [PubMed]
2. Centers for Disease Control and Prevention. 2007. Increase in gonorrhea—eight western states, 2000-2005. MMWR Morb. Mortal. Wkly. Rep. 56:222-225. [PubMed]
3. de Vries, H. J., J. J. van der Helm, M. F. Schim van der Loeff, and A. P. van Dam. 2009. Multidrug-resistant Neisseria gonorrhoeae with reduced cefotaxime susceptibility is increasingly common in men who have sex with men, Amsterdam, the Netherlands. Euro Surveill. 14:pii:19330. [PubMed]
4. Fenton, K. A., and J. Imrie. 2005. Increasing rates of sexually transmitted diseases in homosexual men in Western Europe and the United States: why? Infect. Dis. Clin. North Am. 19:311-331. [PubMed]
5. Fenton, K. A., and C. M. Lowndes. 2004. Recent trends in the epidemiology of sexually transmitted infections in the European Union. Sex. Transm. Infect. 80:255-263. [PMC free article] [PubMed]
6. Hsueh, P. R., S. P. Tseng, L. J. Teng, and S. W. Ho. 2005. High prevalence of ciprofloxacin-resistant Neisseria gonorrhoeae in northern Taiwan. Clin. Infect. Dis. 40:188-192. [PubMed]
7. Ito, M., T. Deguchi, K. S. Mizutani, M. Yasuda, S. Yokoi, S. Ito, Y. Takahashi, S. Ishihara, Y. Kawamura, and T. Ezaki. 2005. Emergence and spread of Neisseria gonorrhoeae clinical isolates harboring mosaic-like structure of penicillin-binding protein 2 in central Japan. Antimicrob. Agents Chemother. 49:137-143. [PMC free article] [PubMed]
8. Lo, J. Y., K. M. Ho, A. O. Leung, F. S. Tiu, G. K. Tsang, A. C. Lo, and J. W. Tapsall. 2008. Ceftibuten resistance and treatment failure of Neisseria gonorrhoeae infection. Antimicrob. Agents Chemother. 52:3564-3567. [PMC free article] [PubMed]
9. Ohnishi, M., Y. Watanabe, E. Ono, C. Takahashi, H. Oya, T. Kuroki, K. Shimuta, N. Okazaki, S. Nakayama, and H. Watanabe. 2010. Spread of a chromosomal cefixime-resistant penA gene among different Neisseria gonorrhoeae lineages. Antimicrob. Agents Chemother. 54:1060-1067. [PMC free article] [PubMed]
10. Pandori, M., P. M. Barry, A. Wu, A. Ren, W. L. Whittington, S. Liska, and J. D. Klausner. 2009. Mosaic penicillin-binding protein 2 in Neisseria gonorrhoeae isolates collected in 2008 in San Francisco, California. Antimicrob. Agents Chemother. 53:4032-4034. [PMC free article] [PubMed]
11. Starnino, S., and P. Stefanelli. 2009. Azithromycin-resistant Neisseria gonorrhoeae strains recently isolated in Italy. J. Antimicrob. Chemother. 63:1200-1204. [PubMed]
12. Tanaka, M., H. Nakayama, K. Huruya, I. Konomi, S. Irie, A. Kanayama, T. Saika, and I. Kobayashi. 2006. Analysis of mutations within multiple genes associated with resistance in a clinical isolate of Neisseria gonorrhoeae with reduced ceftriaxone susceptibility that shows a multidrug-resistant phenotype. Int. J. Antimicrob. Agents 27:20-26. [PubMed]
13. Tapsall, J. W., S. Ray, and A. Limnios. 2010. Characteristics and population dynamics of mosaic penA allele-containing Neisseria gonorrhoeae isolates collected in Sydney, Australia, in 2007-2008. Antimicrob. Agents Chemother. 54:554-556. [PMC free article] [PubMed]
14. Wang, S. A., M. V. Lee, N. O'Connor, C. J. Iverson, R. G. Ohye, P. M. Whiticar, J. A. Hale, D. L. Trees, J. S. Knapp, P. V. Effler, and H. S. Weinstock. 2003. Multidrug-resistant Neisseria gonorrhoeae with decreased susceptibility to cefixime—Hawaii, 2001. Clin. Infect. Dis. 37:849-852. [PubMed]
15. Whiley, D. M., E. A. Limnios, S. Ray, T. P. Sloots, and J. W. Tapsall. 2007. Diversity of penA alterations and subtypes in Neisseria gonorrhoeae strains from Sydney, Australia, that are less susceptible to ceftriaxone. Antimicrob. Agents Chemother. 51:3111-3116. [PMC free article] [PubMed]
16. WHO Western Pacific Gonococcal Antimicrobial Surveillance Programme. 2006. Surveillance of antibiotic resistance in Neisseria gonorrhoeae in the WHO Western Pacific Region, 2005. Commun. Dis. Intell. 30:430-433. [PubMed]
17. Zhao, S., M. Duncan, J. Tomberg, C. Davies, M. Unemo, and R. A. Nicholas. 2009. Genetics of chromosomally mediated intermediate resistance to ceftriaxone and cefixime in Neisseria gonorrhoeae. Antimicrob. Agents Chemother. 53:3744-3751. [PMC free article] [PubMed]

Articles from Antimicrobial Agents and Chemotherapy are provided here courtesy of American Society for Microbiology (ASM)
PubReader format: click here to try

Formats: