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Background  The spread of  Neisseria gonorrhoeae  strains with mosaic  penA  alleles and reduced susceptibility to extended-spectrum cephalosporins is a major public health problem. While much work has been performed internationally, little is known about the genetics or molecular epidemiology of  N. gonorrhoeae  isolates with reduced susceptibility to extended-spectrum cephalosporins in the United States. The majority of  N. gonorrhoeae  infections are diagnosed without a live culture. Molecular tools capable of detecting markers of extended-spectrum cephalosporin resistance are needed.    Methods  Urethral  N. gonorrhoeae  isolates were collected from 684 men at public health clinics in California in 2011. Minimum inhibitory concentrations (MICs) to ceftriaxone, cefixime, cefpodoxime and azithromycin were determined by Etest and categorized according to the U.S. Centers for Disease Control 2010 alert value breakpoints. 684 isolates were screened for mosaic  penA  alleles using real-time PCR (RTPCR) and 59 reactive isolates were subjected to DNA sequencing of their  penA  alleles and  Neisseria gonorrhoeae  multi-antigen sequence typing (NG-MAST). To increase the specificity of the screening RTPCR in detecting isolates with alert value extended-spectrum cephalosporin MICs, the primers were modified to selectively amplify the mosaic XXXIV  penA  allele.    Results  Three mosaic  penA  alleles were detected including two previously described alleles (XXXIV, XXXVIII) and one novel allele (LA-A). Of the 29 isolates with an alert value extended-spectrum cephalosporin MIC, all possessed the mosaic XXXIV  penA  allele and 18 were sequence type 1407, an internationally successful strain associated with multi-drug resistance. The modified RTPCR detected the mosaic XXXIV  penA  allele in urethral isolates and urine specimens and displayed no amplification of the other  penA  alleles detected in this study.    Conclusion   N. gonorrhoeae  isolates with mosaic  penA  alleles and reduced susceptibility to extended-spectrum cephalosporins are currently circulating in California. Isolates with the same NG-MAST ST,  penA  allele and extended-spectrum cephalosporin MICs have caused treatment failures elsewhere. The RTPCR assay presented here may be useful for the detection of  N. gonorrheoae  isolates and clinical specimens with reduced extended-spectrum cephalosporin MICs in settings where antimicrobial susceptibility testing is unavailable. In an era of increasing antimicrobial resistance and decreasing culture capacity, molecular assays capable of detecting extended-spectrum cephalosporin of resistance are essential to public health.

Neisseria gonorrhoeae and extended-spectrum cephalosporins in California: surveillance and molecular detection of mosaic penA