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Neisseria meningitidis (Nm) and Neisseria gonorrhoeae (Ng) are human pathogens that sometimes occupy the same anatomical niche. Ng, the causative agent of gonorrhea, infects 87 million individuals annually worldwide and is an urgent threat due to increasing drug resistance. Ng is a pathogen of the urogenital tract and may infect the oropharyngeal or rectal site, often asymptomatically. Conversely, Nm is an opportunistic pathogen. While often a commensal in the oropharyngeal tract, it is also the leading cause of bacterial meningitis with 1.2 million cases globally, causing significant morbidity and mortality. Horizontal gene transfer (HGT) is likely to occur between Ng and Nm due to their shared anatomical niches and genetic similarity, which poses challenges for accurate detection and treatment. Routine surveillance through the Gonococcal Isolate Surveillance Project and Strengthening the U.S. Response to Resistant Gonorrhea detected six concerning urogenital Neisseria isolates with contradicting species identification in Milwaukee (MIL). While all six isolates were positive for Ng using nucleic acid amplification testing (NAAT) and matrix-assisted laser desorption/ionization time of flight identified the isolates as Ng, two biochemical tests, Gonochek-II and API NH, classified them as Nm. To address this discrepancy, we performed whole-genome sequencing (WGS) using Illumina MiSeq on all isolates and employed various bioinformatics tools. Species detection analysis using BMScan, which uses WGS data, identified all isolates as Ng. Furthermore, Kraken revealed over 98% of WGS reads mapped to the Ng genome and <1% to Nm. Recombination analysis identified putative HGT in all MIL isolates within the γ-glutamyl transpeptidase ( ggt ) gene, a key component in the biochemical tests used to differentiate between Nm and Ng. Further analysis identified Nm as the source of HGT event. Specifically, the active Nm ggt gene replaced the Ng pseudogenes, ggt1 and ggt2 . Together, this study demonstrates that closely related Neisseria species sharing a niche underwent HGT, which led to the misidentification of species following biochemical testing. Importantly, NAAT accurately detected Ng. The misidentification highlights the importance of using WGS to continually evaluate diagnostic or bacterial identification tests., Competing Interests: The authors declare no conflict of interest.

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., Competing Interests: The authors have declared that no competing interests exist.

Neisseria gonorrhoeae multilocus sequence type (ST) 9363 core-genogroup isolates have been associated with reduced azithromycin susceptibility (AZMrs) and show evidence of clonal expansion in the United States. Here, we analyze a global collection of ST-9363 core-genogroup genomes to shed light on the emergence and dissemination of this strain. The global population structure of ST-9363 core-genogroup falls into three lineages: Basal, European, and North American; with 32 clades within all lineages. Although, ST-9363 core-genogroup is inferred to have originated from Asia in the mid-19th century; we estimate the three modern lineages emerged from Europe in the late 1970s to early 1980s. The European lineage appears to have emerged and expanded from around 1986 to 1998, spreading into North America and Oceania in the mid-2000s with multiple introductions, along with multiple secondary reintroductions into Europe. Our results suggest two separate acquisition events of mosaic mtrR and mtrR promoter alleles: first during 2009-2011 and again during the 2012-2013 time, facilitating the clonal expansion of this core-genogroup with AZMrs in the United States. By tracking phylodynamic evolutionary trajectories of clades that share distinct demography as well as population-based genomic statistics, we demonstrate how recombination and selective pressures in the mtrCDE efflux operon granted a fitness advantage to establish ST-9363 as a successful gonococcal lineage in the United States and elsewhere. Although it is difficult to pinpoint the exact timing and emergence of this young core-genogroup, it remains critically important to continue monitoring it, as it could acquire additional resistance markers., (Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution 2021.)

Background: Sexual networks are difficult to construct because of incomplete sexual partner data. The proximity of people within a network may be inferred from genetically similar infections. We explored genomic data combined with partner services investigation (PSI) data to extend our understanding of sexual networks affected by Neisseria gonorrhoeae (NG)., Methods: We used 2017-2019 PSI and whole-genome sequencing (WGS) data from 8 jurisdictions participating in Centers for Disease Control and Prevention's Strengthening the US Response to Resistant Gonorrhea (SURRG) project. Clusters were identified from sexual contacts and through genetically similar NG isolates. Sexual mixing patterns were characterized by describing the clusters by the individual's gender and gender of their sex partners., Results: Our study included 4627 diagnoses of NG infection (81% sequenced), 2455 people received a PSI, 393 people were negative contacts of cases, and 495 were contacts with an unknown NG status. We identified 823 distinct clusters using PSI data combined with WGS data. Of cases that were not linked to any other case using PSI data, 37% were linked when using WGS data. Overall, 40% of PSI cases were allocated to a larger cluster when PSI and WGS data were combined compared with PSI data alone. Mixed clusters containing women, men who report sex with women, and men who report sex with men were common when using the WGS data either alone or in combination with the PSI data., Conclusions: Combining PSI and WGS data improves our understanding of sexual network connectivity., Competing Interests: Conflict of Interest and Sources of Funding: The findings and conclusions in this report are those of the authors and do not necessarily represent the official position of the US Centers for Disease Control and Prevention (CDC) and the Agency for Toxic Substances and Disease Registry. No author reports any conflict of interest., (Copyright © 2021 Written work prepared by employees of the Federal Government as part of their official duties is, under the U.S. Copyright Act, a “work of the United States Government” for which copyright protection under Title 17 of the United States Code is not available. As such, copyright does not extend to the contributions of employees of the Federal Government.)

The recent emergence of strains of Neisseria gonorrhoeae associated with treatment failures to ceftriaxone, the foundation of current treatment options, has raised concerns over a future of untreatable gonorrhea. Current global data on gonococcal strains suggest that several lineages, predominately characterized by mosaic penA alleles, are associated with elevated minimum inhibitory concentrations (MICs) to extended spectrum cephalosporins (ESCs). Here we report on whole genome sequences of 813 N. gonorrhoeae isolates collected through the Gonococcal Isolate Surveillance Project in the United States. Phylogenomic analysis revealed that one persisting lineage (Clade A, multi-locus sequence type [MLST] ST1901) with mosaic penA-34 alleles, contained the majority of isolates with elevated MICs to ESCs. We provide evidence that an ancestor to the globally circulating MLST ST1901 clones potentially emerged around the early to mid-20th century (1944, credibility intervals [CI]: 1935-1953), predating the introduction of cephalosporins, but coinciding with the use of penicillin. Such results indicate that drugs with novel mechanisms of action are needed as these strains continue to persist and disseminate globally.

Background: Antimicrobial-resistant (AMR) Neisseria gonorrhoeae is an urgent threat to public health, as strains resistant to at least one of the two last-line antibiotics used in empiric therapy of gonorrhoea, ceftriaxone and azithromycin, have spread internationally. Whole genome sequencing (WGS) data can be used to identify new AMR clones and transmission networks and inform the development of point-of-care tests for antimicrobial susceptibility, novel antimicrobials and vaccines. Community-driven tools that provide an easy access to and analysis of genomic and epidemiological data is the way forward for public health surveillance., Methods: Here we present a public health-focussed scheme for genomic epidemiology of N. gonorrhoeae at Pathogenwatch ( https://pathogen.watch/ngonorrhoeae ). An international advisory group of experts in epidemiology, public health, genetics and genomics of N. gonorrhoeae was convened to inform on the utility of current and future analytics in the platform. We implement backwards compatibility with MLST, NG-MAST and NG-STAR typing schemes as well as an exhaustive library of genetic AMR determinants linked to a genotypic prediction of resistance to eight antibiotics. A collection of over 12,000 N. gonorrhoeae genome sequences from public archives has been quality-checked, assembled and made public together with available metadata for contextualization., Results: AMR prediction from genome data revealed specificity values over 99% for azithromycin, ciprofloxacin and ceftriaxone and sensitivity values around 99% for benzylpenicillin and tetracycline. A case study using the Pathogenwatch collection of N. gonorrhoeae public genomes showed the global expansion of an azithromycin-resistant lineage carrying a mosaic mtr over at least the last 10 years, emphasising the power of Pathogenwatch to explore and evaluate genomic epidemiology questions of public health concern., Conclusions: The N. gonorrhoeae scheme in Pathogenwatch provides customised bioinformatic pipelines guided by expert opinion that can be adapted to public health agencies and departments with little expertise in bioinformatics and lower-resourced settings with internet connection but limited computational infrastructure. The advisory group will assess and identify ongoing public health needs in the field of gonorrhoea, particularly regarding gonococcal AMR, in order to further enhance utility with modified or new analytic methods.

Competing Interests: All authors have completed and submitted the International Committee of Medical Journal Editors form for disclosure of potential conflicts of interest. No potential conflicts of interest were disclosed.

Background: The number of cases of gonorrhoea in the USA and worldwide caused by Neisseria gonorrhoeae is increasing (555 608 reported US cases in 2017, and 87 million cases worldwide in 2016). Many countries report declining in vitro susceptibility of azithromycin, which is a concern because azithromycin and ceftriaxone are the recommended dual treatment in many countries. We aimed to identify strain types associated with decreased susceptibility to azithromycin., Methods: We did a genomic analysis of N gonorrhoeae isolates obtained by the US Gonococcal Isolate Surveillance Project. Isolates were whole-genome sequenced based on decreased susceptibility to azithromycin (minimal inhibitory concentration [MIC] ≥2 μg/mL, using agar dilution antibiotic susceptibility testing) and geographical representation. Bioinformatic analyses established genomic diversity, strain population dynamics, and antimicrobial resistance profiles., Findings: 410 isolates were sorted into more than 20 unique phylogenetic clades. One predominant persistent clade (consisting of 97 isolates) included the most isolates with azithromycin MICs of 2 μg/mL or higher (61 of 97 [63%] vs 59 of 311 [19%]; p<0·0001) and carried a mosaic mtr (multiple transferable resistance) locus (68 of 97 [70%] vs two of 313 [1%]; p<0·0001). Of the remaining 313 isolates, 57 (18%) had decreased susceptibility to azithromycin (MIC ≥4 μg/mL), which was attributed to 23S rRNA variants (56 of 57 [98%]) and formed phylogenetically diverse clades, showing various levels of clonal expansion., Interpretation: Reduced azithromycin susceptibility was associated with expanding and persistent clades harbouring two well described resistance mechanisms, mosaic mtr locus and 23S rRNA variants. Understanding the role of recombination, particularly within the mtr locus, on the fitness and expansion of strains with decreased susceptibility has important implications for the public health response to minimise gonorrhoea transmission., Funding: US Centers for Disease Control and Prevention (CDC), CDC Combating Antibiotic Resistant Bacteria initiative, Oak Ridge Institute for Science Education, US Department of Energy/CDC/Emory University, National Institutes of Health, and Biomedical Laboratory Research and Development Service of the US Department of Veterans Affairs., Competing Interests: Declaration of interests We declare no competing interests.

U.S. gonorrhea rates are rising, and antibiotic-resistant Neisseria gonorrhoeae (AR-Ng) is an urgent public health threat. Since implementation of nucleic acid amplification tests for N. gonorrhoeae identification, the capacity for culturing N. gonorrhoeae in the United States has declined, along with the ability to perform culture-based antimicrobial susceptibility testing (AST). Yet AST is critical for detecting and monitoring AR-Ng. In 2016, the CDC established the Antibiotic Resistance Laboratory Network (AR Lab Network) to shore up the national capacity for detecting several resistance threats including N. gonorrhoeae AR-Ng testing, a subactivity of the CDC's AR Lab Network, is performed in a tiered network of approximately 35 local laboratories, four regional laboratories (state public health laboratories in Maryland, Tennessee, Texas, and Washington), and the CDC's national reference laboratory. Local laboratories receive specimens from approximately 60 clinics associated with the Gonococcal Isolate Surveillance Project (GISP), enhanced GISP (eGISP), and the program Strengthening the U.S. Response to Resistant Gonorrhea (SURRG). They isolate and ship up to 20,000 isolates to regional laboratories for culture-based agar dilution AST with seven antibiotics and for whole-genome sequencing of up to 5,000 isolates. The CDC further examines concerning isolates and monitors genetic AR markers. During 2017 and 2018, the network tested 8,214 and 8,628 N. gonorrhoeae isolates, respectively, and the CDC received 531 and 646 concerning isolates and 605 and 3,159 sequences, respectively. In summary, the AR Lab Network supported the laboratory capacity for N. gonorrhoeae AST and associated genetic marker detection, expanding preexisting notification and analysis systems for resistance detection. Continued, robust AST and genomic capacity can help inform national public health monitoring and intervention., (Copyright © 2020 American Society for Microbiology.)

Introduction: Neisseria gonorrhoeae (Ng) has successively developed resistance to all previously recommended antimicrobial therapies, with ceftriaxone being the last option for monotherapy of gonorrhea. Global emergence and international spread of the FC428 clone derived mosaic penA-60 allele, associated with highlevel ceftriaxone minimum inhibitory concentrations (MICs) in non FC428 clone Ng lineages, has become an increasing concern. The penA60 allele carrying Ng was first identified in the U.S. in Las Vegas, Nevada (2019; GCWGS-102723), with a multi-locus sequence type (MLST)-1901 strain, in a non FC428 clone Ng lineage, which is associated with a historically ceftriaxone susceptible core genogroup. Later in 2022, an allele genetically similar to penA-60, mosaic penA-237, was identified in the UK (H22-722) and France (F92) with high-level ceftriaxone MICs and both belonged to MLST-1901. Methods: In this study, we assessed phylogenomic relatedness and antimicrobial resistance (AMR) determinant profiles of these three isolates with high-level ceftriaxone MICs among a global collection of 2,104 genomes belonging to the MLST-1901 core genome cluster group 31, which includes strains separated by a locus threshold of 200 or fewer differences (Ng_cgc_200). Recombination events in and around the penA coding region were catalogued and potential sources of inter species recombinant DNA were also inferred. Results: The global population structure of MLST-1901 core genogroup falls into 4 major lineages. Isolates GCWGS-10723, F92, and H22-722 clustered within Lineage 1, which was dominated by non-mosaic penA-5 alleles. These three isolates formed a clade within Lineage 1 that consisted of isolates from North America and southeast Asia. Neisseria subflava and Neisseria sicca were identified as likely progenitors of two independent recombination events that may have led to the generation of mosaic penA-60 and penA-237, within a possible non-mosaic penA-5 background. Discussions: Our study suggests that there are multiple evolutionary pathways that could generate concerning mosaic penA alleles via homologous recombination of historically susceptible Ng lineages with Neisseria commensals. Enhanced surveillance of gonococcal strains and Neisseria commensals is crucial for understanding of the evolution of AMR, particularly in less-studied regions (e.g., Asia), where high-level ceftriaxone MICs and multi-drug resistance are more prevalent. [ABSTRACT FROM AUTHOR]

Background Disseminated gonococcal infections (DGIs) are thought to be uncommon; surveillance is limited, and case reports are analyzed retrospectively or in case clusters. We describe the population-level burden of culture-confirmed DGIs through the Active Bacterial Core surveillance (ABCs) system. Methods During 2015–2016, retrospective surveillance was conducted among residents in 2 ABCs areas and prospectively in 3 ABCs areas during 2017–2019. A DGI case was defined as isolation of Neisseria gonorrhoeae from a normally sterile site. A case report form was completed for each case and antimicrobial susceptibility testing (AST) was performed on available isolates. Results During 2015–2019, 77 DGI cases were identified (a rate of 0.13 cases per 100 000 population) and accounted for 0.06% of all reported gonorrhea cases in the 3 surveillance areas. Most DGI cases were male (64%), non-Hispanic Black (68%), and ranged from 16 to 67 years of age; blood (55%) and joint (40%) were the most common sterile sites. Among 29 isolates with AST results during 2017–2019, all were susceptible to ceftriaxone. Conclusions DGI is an infrequent complication of N gonorrhoeae ; because it can quickly develop antimicrobial resistance, continued DGI surveillance, including monitoring trends in antimicrobial susceptibility, could help inform DGI treatment recommendations. [ABSTRACT FROM AUTHOR]

Disseminated gonococcal infection (DGI) is a rare complication caused by the systemic dissemination of Neisseria gonorrhoeae to normally sterile anatomical sites. Little is known about the genetic diversity of DGI gonococcal strains and how they relate to other gonococcal strains causing uncomplicated mucosal infections. We used whole genome sequencing to characterize DGI isolates (n = 30) collected from a surveillance system in Georgia, United States, during 2017–2020 to understand phylogenetic clustering among DGI as well as uncomplicated uro- and extragenital gonococcal infection (UGI) isolates (n = 110) collected in Fulton County, Georgia, during 2017–2019. We also investigated the presence or absence of genetic markers related to antimicrobial resistance (AMR) as well as surveyed the genomes for putative virulence genetic factors associated with normal human-serum (NHS) resistance that might facilitate DGI. We found that DGI strains demonstrated significant genetic variability similar to the population structure of isolates causing UGI, with sporadic incidences of geographically clustered DGI strains. DGI isolates contained various AMR markers and genetic mechanisms associated with NHS resistance. DGI isolates had a higher frequency of the porB1A allele compared with UGI (67% vs 9%, P  < .0001); however, no single NHS resistance marker was found in all DGI isolates. Continued DGI surveillance with genome-based characterization of DGI isolates is necessary to better understand specific factors that promote systemic dissemination. [ABSTRACT FROM AUTHOR]