RESUMEN
Clinical resistance to the currently recommended extended-spectrum cephalosporins (ESCs), the last remaining treatment options for gonorrhea, is being reported. Gonorrhea may become untreatable, and new treatment options are crucial. We investigated the in vitro activity of ertapenem, relative to ceftriaxone, against N. gonorrhoeae isolates and the effects of ESC resistance determinants on ertapenem. MICs were determined using agar dilution technique or Etest for international reference strains (n = 17) and clinical N. gonorrhoeae isolates (n = 257), which included the two extensively drug-resistant (XDR) strains H041 and F89 and additional isolates with high ESC MICs, clinical ESC resistance, and other types of clinical high-level and multidrug resistance (MDR). Genetic resistance determinants for ESCs (penA, mtrR, and penB) were sequenced. In general, the MICs of ertapenem (MIC(50) = 0.032 µg/ml; MIC(90) = 0.064 µg/ml) paralleled those of ceftriaxone (MIC(50) = 0.032 µg/ml; MIC(90) = 0.125 µg/ml). The ESC resistance determinants mainly increased the ertapenem MIC and ceftriaxone MIC at similar levels. However, the MIC ranges for ertapenem (0.002 to 0.125 µg/ml) and ceftriaxone (<0.002 to 4 µg/ml) differed, and the four (1.5%) ceftriaxone-resistant isolates (MIC = 0.5 to 4 µg/ml) had ertapenem MICs of 0.016 to 0.064 µg/ml. Accordingly, ertapenem had in vitro advantages over ceftriaxone for isolates with ceftriaxone resistance. These in vitro results suggest that ertapenem might be an effective treatment option for gonorrhea, particularly for the currently identified ESC-resistant cases and possibly in a dual antimicrobial therapy regimen. However, further knowledge regarding the genetic determinants (and their evolution) conferring resistance to both antimicrobials, and clear correlates between genetic and phenotypic laboratory parameters and clinical treatment outcomes, is essential.
Asunto(s)
Antibacterianos/farmacología , Ceftriaxona/farmacología , Gonorrea/microbiología , Neisseria gonorrhoeae/efectos de los fármacos , beta-Lactamas/farmacología , Farmacorresistencia Bacteriana , Ertapenem , Humanos , Pruebas de Sensibilidad Microbiana , Neisseria gonorrhoeae/patogenicidadRESUMEN
With increasing concerns regarding diminishing treatment options for gonorrhea, maintaining the efficacy of currently used treatments and ensuring optimal Neisseria gonorrhoeae antimicrobial resistance surveillance are of the utmost importance. Penicillin is still used to treat gonorrhea in some parts of the world. In this study, we developed and validated a real-time PCR assay for the detection of penicillinase-producing N. gonorrhoeae (PPNG) in noncultured clinical samples with the aim of enhancing penicillin resistance surveillance. The assay (PPNG-PCR2) was designed to be an indirect marker of penicillinase activity, by targeting a region of sequence predicted to be conserved across all N. gonorrhoeae plasmid types harboring the beta-lactamase gene while not specifically targeting the actual beta-lactamase-encoding sequence. The assay was evaluated by using a total of 118 N. gonorrhoeae clinical isolates and 1,194 clinical specimens, including 239 N. gonorrhoeae-positive clinical samples from which N. gonorrhoeae cells were isolated and for which phenotypic penicillinase results are available. Overall, the PPNG-PCR2 assay provided 100% sensitivity and 98.7% specificity compared to bacterial culture results for the detection of PPNG in clinical specimens. PPNG-PCR2 false-positive results, presumably due to cross-reactions with unrelated bacterial species, were observed for up to 1.3% of clinical samples but could be distinguished on the basis of high cycle threshold values. In tandem with phenotypic surveillance, the PPNG-PCR2 assay has the potential to provide enhanced epidemiological surveillance of N. gonorrhoeae penicillin resistance and is of particular relevance to regions where penicillin is still used to treat gonorrhea.
Asunto(s)
Farmacorresistencia Bacteriana , Gonorrea/microbiología , Neisseria gonorrhoeae/enzimología , Neisseria gonorrhoeae/genética , Penicilinasa/biosíntesis , Reacción en Cadena de la Polimerasa/métodos , Secuencia Conservada , Cartilla de ADN/genética , ADN Bacteriano/genética , Humanos , Pruebas de Sensibilidad Microbiana/métodos , Neisseria gonorrhoeae/efectos de los fármacos , Neisseria gonorrhoeae/aislamiento & purificación , Penicilinasa/genética , Plásmidos , Sensibilidad y EspecificidadRESUMEN
Molecular detection of Neisseria gonorrhoeae in extragenital samples may result in false-positive results due to cross-reaction with commensal Neisseria species or Neisseria meningitidis. This study examined 450 characterized clinical culture isolates, comprising 216 N. gonorrhoeae isolates and 234 isolates of nongonococcal Neisseria species (n = 218) and 16 isolates of other closely related bacteria, with six commercial nucleic acid amplification tests (NAATs). The six NAATs tested were Gen-Probe APTIMA COMBO 2 and APTIMA GC, Roche COBAS Amplicor CT/NG and COBAS 4800 CT/NG tests, BD ProbeTec GC Qx amplified DNA assay, and Abbott RealTime CT/NG test. All assays except COBAS Amplicor CT/NG test where four (1.9%) isolates were not detected showed a positive result with all N. gonorrhoeae isolates (n = 216). Among the 234 nongonococcal isolates examined, initial results from all assays displayed some false-positive results due to cross-reactions. Specifically, the COBAS Amplicor and ProbeTec tests showed the highest number of false-positive results, detecting 33 (14.1%) and 26 (11%) nongonococcal Neisseria isolates, respectively. On the first testing, APTIMA COMBO 2, APTIMA GC, Abbott RealTime, and Roche COBAS 4800 showed lower level of cross-reactions with five (2.1%), four (1.7%), two (1%), and two (1%) of the isolates showing low-level positivity, respectively. Upon retesting of these nine nongonococcal isolates using freshly cultured colonies, none were positive by the APTIMA COMBO 2, Abbott RealTime, or COBAS 4800 test. In conclusion, the COBAS Amplicor and ProbeTec tests displayed high number of false-positive results, while the remaining NAATs showed only sporadic low-level false-positive results. Supplementary testing for confirmation of N. gonorrhoeae NAATs remains recommended with all samples tested, in particular those from extragenital sites.
Asunto(s)
Técnicas Bacteriológicas/métodos , Técnicas de Diagnóstico Molecular/métodos , Neisseria/clasificación , Neisseria/aislamiento & purificación , Infecciones por Neisseriaceae/diagnóstico , Infecciones por Neisseriaceae/microbiología , Técnicas de Amplificación de Ácido Nucleico/métodos , Reacciones Falso Positivas , Humanos , Neisseria/genética , Sensibilidad y EspecificidadRESUMEN
Eighteen hundred Neisseria gonorrhoeae isolates collected in Sydney, Australia, in 2007 and 2008 were examined for mosaic penA alleles that mediated cephalosporin resistance, and the genotypes of the isolates were evaluated. In 2008, there were substantial increases in numbers (from 15 to 85) and proportions (from 1.5 to 10.3%) of mosaic-containing gonococci and major shifts in genotypic patterns, with 10 new genotypes representing 74 of the 85 mosaic-containing isolates and genotypes detected between 2001 and 2005 having disappeared. Enhanced surveillance of gonococcal resistance to cephalosporins is necessary.
Asunto(s)
Genes Bacterianos/genética , Gonorrea/microbiología , Neisseria gonorrhoeae/genética , Proteínas de Unión a las Penicilinas/genética , Alelos , Antibacterianos/farmacología , Australia/epidemiología , Cefalosporinas/farmacología , Farmacorresistencia Bacteriana/genética , Genotipo , Gonorrea/epidemiología , Humanos , Estudios LongitudinalesRESUMEN
OBJECTIVES: Gonorrhoea remains a global public health problem and the treatment options are diminishing through the emergence of gonococci resistant to most antimicrobials. Previous in vitro studies have indicated a role for Neisseria gonorrhoeae pilQ alterations in conferring resistance to antimicrobials, including penicillin. In this study, we investigated whether pilQ polymorphisms were associated with decreased susceptibility to extended-spectrum cephalosporins (ESCs) in clinical gonococcal strains. METHODS: Full-length pilQ nucleotide and PilQ amino acid sequences from geographically and temporally diverse gonococcal clinical isolates (nâ=â63), including the 2008 WHO reference strains, representing a range of ceftriaxone and cefixime MICs (≤0.008-0.25 and <0.016-0.5 mg/L, respectively) and 38 N. gonorrhoeae multiantigen sequence types, were examined. Previously described alterations associated with decreased ESC susceptibility (mosaic penA, mtrR and penB alterations) were also examined. RESULTS: Fifteen different pilQ nucleotide sequence types and nine different PilQ amino acid sequence types were observed, with two PilQ types accounting for 53 (84%) of the isolates. Independent of other genetic resistance determinants (penA mosaic, mtrR promoter deletion and penB), only one pilQ alteration, a D526N substitution, provided a statistically significant association with ceftriaxone (Pâ<â0.01) and cefixime (Pâ<â0.05) MICs. However, the two isolates exhibiting D526N lacked all three previously described alterations associated with decreased ESC susceptibility, thereby providing an alternative basis for the low MICs (≤0.008 mg/L) observed for these strains. The previously described E666K (pilQ2) and F595L (pilQ1) mutations were absent in all 63 isolates. CONCLUSIONS: pilQ polymorphisms are unlikely contributors to decreased susceptibility to ESCs in clinical gonococcal strains.
Asunto(s)
Antibacterianos/farmacología , Cefixima/farmacología , Ceftriaxona/farmacología , Resistencia a las Cefalosporinas/genética , Proteínas Fimbrias/genética , Neisseria gonorrhoeae/efectos de los fármacos , Secuencia de Aminoácidos , Gonorrea/microbiología , Humanos , Pruebas de Sensibilidad Microbiana , Datos de Secuencia Molecular , Mutación , Neisseria gonorrhoeae/genéticaRESUMEN
OBJECTIVES: Reduced susceptibility to extended-spectrum cephalosporins in Neisseria gonorrhoeae has, to date, been associated with three alterations: a mosaic penA allele encoding the penicillin-binding protein 2 (PBP2); A-del-mtrR, an adenine deletion in the mtrR promoter; and penB, comprising mutated alleles of PorBIb. In this study, we examined an association between reduced susceptibility to ceftriaxone and additional mutations in gonococcal PBP2. METHODS: N. gonorrhoeae isolates (n = 76) exhibiting reduced susceptibility to ceftriaxone but lacking the mosaic penA sequence were investigated for A-del-mtrR and penB as well as substitutions at PBP2 501, 542 and 551 using a previously described real-time PCR approach. To further investigate PBP2 542 and 551 substitutions, we reanalysed penA sequence data from a previous study of 98 gonococci exhibiting a range of ceftriaxone MICs. RESULTS: Of 76 N. gonorrhoeae isolates exhibiting reduced susceptibility to ceftriaxone and lacking the mosaic penA sequence, a 501 (A501V or A501T) substitution was present in 9/76, a 542 substitution in 39/76 and a 551 substitution in 26/76 isolates. Reanalysis of 98 gonococcal isolates from a previous study showed that substitutions at PBP2 542 (G542S) and 551 (P551S or P551L) were significantly associated with raised MICs to ceftriaxone (P = 0.0186 and 0.001, respectively) and penicillin (P = 0.0231 and 0.0007, respectively). CONCLUSIONS: Our findings provide strong evidence for the involvement of PBP2 G542S and P551S/P551L in reduced susceptibility to ceftriaxone and to penicillin. Further studies are needed to investigate the precise and relative roles played by these mutations.
Asunto(s)
Antibacterianos/farmacología , Proteínas Bacterianas/genética , Ceftriaxona/farmacología , Mutación Missense , Neisseria gonorrhoeae/efectos de los fármacos , Proteínas de Unión a las Penicilinas/genética , Resistencia betalactámica , Sustitución de Aminoácidos/genética , Pruebas de Sensibilidad Microbiana , Análisis de Secuencia de ADNRESUMEN
OBJECTIVES: The Neisseria gonorrhoeae multi-antigen sequence typing (NG-MAST) system, based on PCR amplification and sequence analysis of the gonococcal porB and tbpB genes, is widely used for molecular typing of gonococcal isolates but is not validated for non-cultured clinical samples. This study sought to examine the performance of the NG-MAST system on a range of non-cultured samples. METHODS: Nucleic acid extracts of 73 N gonorrhoeae-positive samples, comprising eight cervical swabs, nine urethral swabs, 35 urine samples, one vaginal swab, 13 rectal swabs and seven throat swabs, were analysed by NG-MAST. For 27 specimens, corresponding gonococcal isolates were also analysed and the results compared. A panel of 44 non-gonococcal Neisseria strains and 100 N gonorrhoeae-negative clinical samples were used to investigate further the specificity of the NG-MAST PCR reactions. RESULTS: PCR amplification and DNA sequencing of gonococcal porB and tbpB genes was successful for all N gonorrhoeae-positive urogenital specimens, 11 of 13 rectal swabs and four of seven throat swabs. For the 27 N gonorrhoeae-positive specimens with corresponding gonococcal isolates, the porB and tbpB sequences obtained from the non-cultured specimen were identical to those obtained from the isolate. Cross-reaction with non-gonococcal Neisseria species was observed for both the porB and tbpB PCR reactions, and proved to be problematical for NG-MAST typing of throat swab specimens. CONCLUSIONS: The NG-MAST system can successfully be applied directly to non-cultured urogenital samples, but is less suitable for extragenital specimens, particularly throat swabs, due to cross-reaction with commensal Neisseria species.
Asunto(s)
Antígenos Bacterianos/genética , Técnicas de Tipificación Bacteriana/métodos , Gonorrea/microbiología , Neisseria gonorrhoeae/clasificación , Cuello del Útero/microbiología , ADN Bacteriano/genética , Femenino , Humanos , Masculino , Neisseria gonorrhoeae/genética , Neisseria gonorrhoeae/inmunología , Neisseria gonorrhoeae/aislamiento & purificación , Faringe/microbiología , Reacción en Cadena de la Polimerasa/métodos , Recto/microbiología , Sensibilidad y Especificidad , Uretra/microbiología , Orina/microbiologíaRESUMEN
We report a duplex real-time PCR assay for the simultaneous screening of mutations involved in fluoroquinolone resistance within gyrA and parC quninolone resistance-determining regions (QRDRs) in Neisseria gonorrhoeae. Our assay clearly detects all mutated QRDRs and allows the identification of common genotypes, whether the QRDRs contain single or double mutations, providing valuable epidemiological tools. When this method is used in conjunction with similar assays and in vitro analyses, essential antibiotic resistance surveillance can be performed for public health purposes.
Asunto(s)
Girasa de ADN/genética , Farmacorresistencia Bacteriana/genética , Genes Bacterianos , Neisseria gonorrhoeae/genética , Quinolonas/farmacología , ADN Bacteriano/química , ADN Bacteriano/genética , Susceptibilidad a Enfermedades , Reacción en Cadena de la Polimerasa , Valor Predictivo de las Pruebas , Quinolonas/uso terapéuticoRESUMEN
Neisseria gonorrhoeae has developed resistance to multiple classes of antimicrobials. There is now growing concern that without the availability of appropriate public health strategies to combat this problem, gonorrhea could become untreatable. For this reason, surveillance for gonococcal antimicrobial resistance must be optimal both in terms of obtaining a representative sample of gonococcal isolates and in terms of having the appropriate tools to identify resistance. To aid with this surveillance, molecular tools are increasingly being used. In the present study, we investigated the use of a simple heat denaturation protocol for isolate DNA preparation combined with SYBR green-based real-time PCR for the identification of mutations associated with N. gonorrhoeae antimicrobial resistance. A total of 109 clinical gonococcal isolates were tested by high-resolution melting (HRM) curve analysis for chromosomal mutations associated with gonococcal resistance to beta-lactam antibiotics: a penA 345A insertion, ponA L421P, mtrR G45D, substitutions at positions 120 and 121 in porB1b, and an adenine deletion in the mtrR promoter. An allele-specific PCR assay was also investigated for its ability to detect the adenine deletion in the mtrR promoter. The results were compared to those obtained by DNA sequencing. Our HRM assays provided the accurate discrimination of heat-treated isolates in which the sequence types differed in GC content, including isolates with the penA 345A insertion and the ponA L421P and mtrR G45D mutations. The allele-specific PCR assay accurately identified isolates with the adenine deletion in the mtrR promoter. Heat-denatured DNA combined with SYBR green-based real-time PCR offers a simple, rapid, and inexpensive means of detecting gonococcal resistance mechanisms. These methods may have broader application in the detection of polymorphisms associated with phenotypes of interest.
Asunto(s)
Farmacorresistencia Bacteriana Múltiple/genética , Neisseria gonorrhoeae/genética , Reacción en Cadena de la Polimerasa/métodos , Antibacterianos/farmacología , Proteínas Bacterianas/genética , Proteínas Bacterianas/fisiología , Secuencia de Bases , Datos de Secuencia Molecular , Mutación , Neisseria gonorrhoeae/efectos de los fármacos , Reproducibilidad de los Resultados , Homología de Secuencia de Ácido NucleicoRESUMEN
PURPOSE OF REVIEW: Antibiotic resistance in Neisseria gonorrhoeae poses on-going problems for individual case management and disease control for gonorrhoea. Considerable reliance is now placed on third-generation cephalosporins for the treatment of gonorrhoea following the loss of efficacy of penicillins and quinolones. Current clinical and laboratory perspectives on N. gonorrhoeae with decreased susceptibility to third-generation cephalosporins are provided. RECENT FINDINGS: Treatment failures following therapy with the oral third-generation cephalosporins cefixime and ceftibuten have been reported, but not with the injectable ceftriaxone. The gonococci involved have raised minimal inhibitory concentrations to these agents, including to ceftriaxone. The presence of multiple chromosomal changes form the basis for this 'resistance', prominent among which is a mosaic penicillin-binding protein 2 found in association with additional known and unknown mutations in other genes. The imprecise nature of laboratory criteria for detecting these gonococci means that the distribution and prevalence of these strains is also uncertain. SUMMARY: Concerns regarding the appearance of gonococci associated with treatment failure with oral cephalosporins are increasing. The origins, causes and patterns of spread of these clinically resistant gonococci are reminiscent of the earlier experiences with quinolone-resistant gonococci. Preventive measures require simultaneous implementation of disease-control principles, coupled with those for antimicrobial resistance.
Asunto(s)
Antibacterianos/farmacología , Cefalosporinas/farmacología , Gonorrea/microbiología , Neisseria gonorrhoeae/efectos de los fármacos , Resistencia betalactámica , Antibacterianos/uso terapéutico , Cefalosporinas/uso terapéutico , Gonorrea/tratamiento farmacológico , Humanos , Insuficiencia del TratamientoRESUMEN
OBJECTIVES: Emergence and spread of antimicrobial resistance (AMR) in Neisseria gonorrhoeae remain a major global problem and expanded, but valid, AMR surveillance is crucial for public health purposes. The World Health Organization (WHO) Collaborating Centre in Sydney, Australia, continually evaluates N. gonorrhoeae strains used in quality control and assurance aspects of the national, WHO regional and international programmes for AMR surveillance it conducts. Here we phenotypically and genetically characterized the 2008 WHO N. gonorrhoeae reference panel, widely used under existing WHO AMR surveillance protocols. MATERIALS AND METHODS: The eight N. gonorrhoeae WHO reference strains were phenotypically characterized by antibiogram, auxotype, serovar and prolyliminopeptidase screening; and genetically with regard to resistance plasmid types, polymorphisms in divergent genetic resistance-mediating loci (n = 9), porB sequencing and N. gonorrhoeae multi-antigen sequence typing. RESULTS: The 2008 WHO reference strains represented all the important susceptible and resistant phenotypes, including corresponding resistance genotypes, and the range of resistances currently seen for relevant antimicrobials. Several pertinent additional phenotypic and genotypic markers, for example, epidemiological markers, were also determined. CONCLUSIONS: The 2008 WHO N. gonorrhoeae reference strain panel was extensively characterized, which is crucial for the expansion of gonococcal AMR surveillance nationally and internationally. The panel is available through WHO sources for quality assurance and quality control aspects of current phenotypic testing protocols, to allow valid comparison of AMR data derived by divergent methods, and also for the control of present and future molecular assays for AMR detection. Additional WHO reference strains can be included as required by the emergence of additional resistant phenotypes and/or genotypes.
Asunto(s)
ADN Bacteriano/genética , Pruebas de Sensibilidad Microbiana/normas , Neisseria gonorrhoeae/fisiología , Estándares de Referencia , Antígenos Bacterianos/genética , Australia , Técnicas de Tipificación Bacteriana , Humanos , Neisseria gonorrhoeae/genética , Fosfotransferasas (aceptor de Grupo Carboxilo)/genética , Plásmidos , Garantía de la Calidad de Atención de Salud/métodos , Control de Calidad , Análisis de Secuencia de ADN , SerotipificaciónRESUMEN
OBJECTIVES: To investigate the presence of resistance genes in nosocomial multidrug-resistant (MDR) Acinetobacter baumannii isolated from outbreak and sporadic settings. METHODS: Thirty-two A. baumannii isolates were collected, 13 of which were involved in two outbreaks from different hospitals, which resulted in four deaths. The remaining 19 isolates were collected sporadically over 5 years from two other hospitals. The MICs of 25 antibiotics were determined for each isolate. PCR screening was carried out to identify possible genes that contributed to each resistance phenotype. Repetitive extragenic palindromic-PCR (REP-PCR) was performed to assess isolate clonality in conjunction with genotype data. RESULTS: Between eight and 12 resistance determinants were detected in the 32 MDR A. baumannii isolates examined. These resistance determinants included the genes blaOXA-23 and ampC, with the upstream element ISAba1 promoting increased gene expression and subsequent resistance to carbapenems and cephalosporins, respectively. In all isolates, resistance to quinolones and fluoroquinolones was conferred by an S83L mutation in GyrA. Twenty-eight of the 32 isolates were also positive for tet(B), a tetracycline resistance determinant, blaTEM-1, which contributed to beta-lactam resistance, and strB, which contributed to aminoglycoside resistance. Class 1 integrons that harboured aacC1, aadA1, qacEDelta1 and sul1 were identified in 10 of the 32 isolates (31%) together with the kanamycin resistance gene, aphA1. A putative trimethoprim resistance gene, folA, was also identified in all isolates. REP-PCR together with genotyping identified three main clonal types. CONCLUSIONS: Isolates of A. baumannii from both outbreak and sporadic cases possess at least eight resistance gene determinants that give rise to the MDR phenotype.
Asunto(s)
Infecciones por Acinetobacter/epidemiología , Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/efectos de los fármacos , Infección Hospitalaria/epidemiología , Infección Hospitalaria/microbiología , Brotes de Enfermedades , Farmacorresistencia Bacteriana Múltiple , Acinetobacter baumannii/genética , Acinetobacter baumannii/aislamiento & purificación , Técnicas de Tipificación Bacteriana , Análisis por Conglomerados , Dermatoglifia del ADN , ADN Bacteriano/genética , Genes Bacterianos , Genotipo , Hospitales , Humanos , Pruebas de Sensibilidad Microbiana , Reacción en Cadena de la Polimerasa/métodosRESUMEN
Diagnostic, genotypic and antibiotic-resistance determinants of Neisseria gonorrhoeae were analysed by molecular methods to verify the failure of ceftriaxone treatment in two cases of pharyngeal gonorrhoea. Monoplex assays were needed to define competitive inhibition of a positive Chlamydia PCR in a duplex assay. Different penA changes were detected in the N. gonorrhoeae isolated from the two cases. These were associated with raised ceftriaxone MICs of 0.03 and 0.016 mg l(-1), which may have contributed to the treatment failures in these cases.
Asunto(s)
Ceftriaxona/uso terapéutico , Chlamydia trachomatis/aislamiento & purificación , Gonorrea/tratamiento farmacológico , Neisseria gonorrhoeae/aislamiento & purificación , Enfermedades Faríngeas/tratamiento farmacológico , Enfermedades Faríngeas/microbiología , Adulto , Antibacterianos/uso terapéutico , Chlamydia trachomatis/genética , Femenino , Homosexualidad , Humanos , Masculino , Reacción en Cadena de la Polimerasa , Insuficiencia del TratamientoRESUMEN
Neisseria gonorrhoeae infections have been empirically treated in Hong Kong with a single oral 400-mg dose of ceftibuten since 1997. Following anecdotal reports of the treatment failure of gonorrhea with oral extended-spectrum cephalosporins, the current study was undertaken to determine the antimicrobial susceptibility pattern and molecular characteristics of isolates of N. gonorrhoeae among patients with putative treatment failure in a sexually transmitted disease clinic setting. Between October 2006 and August 2007, 44 isolates of N. gonorrhoeae were studied from patients identified clinically to have treatment failure with empirical ceftibuten. The ceftibuten MICs for three strains were found to have been 8 mg/liter. These strains were determined by N. gonorrhoeae multiantigen sequence typing to belong to sequence type 835 (ST835) or the closely related ST2469. The testing of an additional eight archived ST835 strains revealed similarly elevated ceftibuten MICs. The penA gene sequences of these 11 isolates all had the mosaic pattern previously described as pattern X. Of note is that the ceftriaxone susceptibility results of these strains all fell within the susceptible range. It is concluded that ceftibuten resistance may contribute to the empirical treatment failure of gonorrhea caused by strains harboring the mosaic penA gene, which confers reduced susceptibility to oral extended-spectrum cephalosporins. Screening for such resistance in the routine clinical laboratory may be undertaken by the disk diffusion test. The continued monitoring of antimicrobial resistance and molecular characteristics of N. gonorrhoeae isolates is important to ensure that control and prevention strategies remain effective.
Asunto(s)
Antibacterianos/farmacología , Resistencia a las Cefalosporinas , Cefalosporinas/farmacología , Gonorrea/tratamiento farmacológico , Gonorrea/microbiología , Neisseria gonorrhoeae/efectos de los fármacos , Ceftibuteno , Resistencia a las Cefalosporinas/genética , Femenino , Genes Bacterianos , Humanos , Masculino , Pruebas de Sensibilidad Microbiana , Neisseria gonorrhoeae/clasificación , Neisseria gonorrhoeae/genética , Neisseria gonorrhoeae/aislamiento & purificación , Insuficiencia del TratamientoRESUMEN
Nucleic acid amplification tests (NAATs) for the detection of Neisseria gonorrhoeae became available in the early 1990s. Although offering several advantages over traditional detection methods, N. gonorrhoeae NAATs do have some limitations. These include cost, risk of carryover contamination, inhibition, and inability to provide antibiotic resistance data. In addition, there are sequence-related limitations that are unique to N. gonorrhoeae NAATs. In particular, false-positive results are a major consideration. These primarily stem from the frequent horizontal genetic exchange occurring within the Neisseria genus, leading to commensal Neisseria species acquiring N. gonorrhoeae genes. Furthermore, some N. gonorrhoeae subtypes may lack specific sequences targeted by a particular NAAT. Therefore, NAAT false-negative results because of sequence variation may occur in some gonococcal populations. Overall, the N. gonorrhoeae species continues to present a considerable challenge for molecular diagnostics. The need to evaluate N. gonorrhoeae NAATs before their use in any new patient population and to educate physicians on the limitations of these tests is emphasized in this review.
Asunto(s)
Gonorrea/diagnóstico , Técnicas de Diagnóstico Molecular , Neisseria gonorrhoeae/genética , Técnicas de Amplificación de Ácido Nucleico/métodos , Reacciones Cruzadas , Sondas de ADN , ADN Bacteriano/química , Gonorrea/economía , Gonorrea/epidemiología , Humanos , Enfermedades Faríngeas/diagnóstico , Enfermedades Faríngeas/microbiología , Enfermedades del Recto/diagnóstico , Enfermedades del Recto/microbiología , Sensibilidad y EspecificidadAsunto(s)
Antibacterianos/farmacología , Ceftriaxona/farmacología , Gonorrea/microbiología , Gonorrea/transmisión , Neisseria gonorrhoeae/efectos de los fármacos , Resistencia betalactámica , Técnicas de Tipificación Bacteriana , Genotipo , Humanos , Pruebas de Sensibilidad Microbiana , Epidemiología Molecular , Tipificación de Secuencias Multilocus , Neisseria gonorrhoeae/clasificación , Neisseria gonorrhoeae/genética , Neisseria gonorrhoeae/aislamiento & purificaciónRESUMEN
AIMS: The primary aim of the study was to determine if the gonococcal porA pseudogene is a stable sequence target for the detection of Neisseria gonorrhoeae by PCR. METHODS: A total of 240 gonococcal strains from various geographic locations were tested by porA pseudogene PCR. In addition, porA pseudogene PCR positivity rates were compared with established gonococcal assays in three Australian states. RESULTS: All N. gonorrhoeae isolates provided positive results in the porA pseudogene PCR. Positivity rates compared favourably with established gonococcal assays, with increased N. gonorrhoeae detection in the Northern Territory and Western Australia. CONCLUSIONS: The results of this multicentre study provide further evidence that the porA pseudogene is highly conserved across a diverse range N. gonorrhoeae strains and is a suitable PCR target for routine detection of N. gonorrhoeae.
Asunto(s)
Técnicas de Diagnóstico Urológico , Gonorrea/diagnóstico , Neisseria gonorrhoeae/genética , Neisseria gonorrhoeae/aislamiento & purificación , Porinas/genética , Reacciones Falso Positivas , Gonorrea/microbiología , Humanos , Neisseria gonorrhoeae/clasificación , Reacción en Cadena de la Polimerasa , Reproducibilidad de los Resultados , Especificidad de la EspecieRESUMEN
The incidence of gonorrhea is increasing in developed countries and remains high elsewhere. This untenable disease burden, the complication rate in women and newborns, and the amplification of human immunodeficiency virus transmission that accompanies gonorrhea makes control of gonococcal disease a priority. However, antibiotic resistance in Neisseria gonorrhoeae has severely compromised the successful treatment of gonorrhea. Older therapies are ineffective, whereas those that remain efficacious are unaffordable in many high-incidence settings. Penicillins, tetracyclines, and newer macrolides have limited utility, and spectinomycin (and in many parts of the world, quinolones) have been withdrawn because of resistance. Of the usually recommended treatments, only the third-generation cephalosporins, and most notably ceftriaxone, have retained their efficacy, but decreased susceptibility to these antibiotics has also appeared. A sustained decrease in gonococcal disease requires an integrated approach combining improved prevention, better diagnosis, and effective treatment. Without continued commitment and effort, gonorrhea may well become untreatable.
Asunto(s)
Antibacterianos/farmacología , Farmacorresistencia Bacteriana , Gonorrea/tratamiento farmacológico , Gonorrea/epidemiología , Neisseria gonorrhoeae/efectos de los fármacos , Antibacterianos/uso terapéutico , Control de Enfermedades Transmisibles/normas , Control de Enfermedades Transmisibles/tendencias , Femenino , Gonorrea/microbiología , Humanos , Cooperación Internacional , Masculino , Pruebas de Sensibilidad Microbiana , Salud Pública/normas , Garantía de la Calidad de Atención de Salud/normas , Enfermedades de Transmisión Sexual/tratamiento farmacológico , Enfermedades de Transmisión Sexual/microbiología , Enfermedades de Transmisión Sexual/transmisiónRESUMEN
AIMS: To describe the development and evaluation of a diagnostic enzyme immunoassay (EIA) for IgM antibody to serogroup C capsular polysaccharide of Neisseria meningitidis (CCPS). METHODS: Purified CCPS was used as the antigen. The optical densities (OD) that determined the cut-off values for the assay were derived using sera from blood bank donors, and the accuracy was evaluated with sera from patients with culture-confirmed serogroup C and non-serogroup C invasive meningococcal disease (IMD), other infectious diseases, culture-confirmed pharyngeal carriage of N. meningitidis and immunoproliferative disorders. RESULTS: In sera collected between days 5 and 20 following positive culture, the assay had a sensitivity of 92%. The two negative sera were from a single patient but showed a significant rise in OD. The sensitivity declined to 80% on days 21-40 then to 75% on days 41-70. No positive reactions were detected in five samples collected after 71 days. The specificity of the assay was at least 97%. CONCLUSIONS: An EIA for IgM antibody to CCPS was evaluated for local conditions and had acceptable sensitivity and specificity. Use of the assay, based on a single blood sample, provided clinically and epidemiologically relevant information for individual and public health management of IMD.
Asunto(s)
Anticuerpos Antibacterianos/sangre , Técnicas para Inmunoenzimas/métodos , Inmunoglobulina M/inmunología , Infecciones Meningocócicas/diagnóstico , Neisseria meningitidis Serogrupo C/inmunología , Humanos , Infecciones Meningocócicas/sangre , Infecciones Meningocócicas/inmunología , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Pruebas Serológicas/métodosRESUMEN
Analysis of atypical meningococci from invasive disease that either (i) did not produce acid from maltose and glucose or (ii) were gamma-glutamyl peptidase test negative for porA and porB DNA variable region (VR) type, multilocus sequence type, and for presence of capsule transport gene ctrA, conclusively demonstrated that these are Neisseria meningitidis.