High-level Tetracycline Resistance Research Articles

An updated molecular diagnostic for surveillance of tetM in Neisseria gonorrhoeae.

Doxycycline post-exposure prophylaxis (doxy-PEP) for sexually transmitted bacterial infections reduces the risk of syphilis and chlamydia, but effectiveness against gonorrhea is variable, likely attributable to varying resistance rates. As doxy-PEP is incorporated into clinical practice, an urgent unanswered question is whether increased doxycycline use will drive tetracycline-class resistance in Neisseria gonorrhoeae . Here, we report an updated RT-PCR molecular diagnostic to detect the tetM gene that confers high-level tetracycline resistance in N. gonorrhoeae .

2687. Tetracycline Resistance in Neisseria gonorrhoeae Isolates Among Transgender Women and Men Who Have Sex with Men — eGISP and SURRG, 2018–2021

Abstract Background Doxycycline, a tetracycline analog, is being considered for use as post-exposure prophylaxis (PEP) to reduce bacterial sexually transmitted infections (STIs) among gay, bisexual, and other men who have sex with men (MSM) and transgender women. Pre-existing tetracycline resistance may contribute to the varied effectiveness of doxycycline in preventing Neisseria gonorrhoeae (GC). We examined trends in tetracycline resistance (TetR) and tetM plasmid-mediated high-level tetracycline resistance (TetHLR) among GC isolates collected from MSM and transgender women through national sentinel surveillance. Methods Genital (urethral, urine, vaginal or endocervical), pharyngeal and rectal GC isolates were collected from MSM and transgender women during 2018–2021 at 12 Enhanced Gonococcal Isolate Surveillance Project (eGISP) and 8 Strengthening the U.S. Response to Resistant Gonorrhea (SURRG) sites. We determined overall prevalence of TetR and TetHLR (minimum inhibitory concentration, MIC ≥2 µg/mL and MIC ≥16 µg/mL, respectively) among MSM and transgender women by anatomic site of infection and described annual trends among MSM. Results Among 10,853 isolates collected from 8,667 MSM, overall TetR prevalence was 31% (genital: 31%, pharyngeal: 32%, rectal: 29%), and annual TetR prevalence decreased during 2018–2021 (Table 1). Overall TetHLR prevalence among MSM was 13% (genital: 13%, pharyngeal: 12%, rectal: 12%) and annual TetHLR prevalence for genital and pharyngeal infections increased during 2018–2021. Among 108 isolates collected from 89 transgender women, overall TetR prevalence was 23% (genital: 19%, pharyngeal: 23%, rectal: 25%) and TetHLR prevalence was 9% (genital: 0%, pharyngeal: 10%, rectal: 12%), though numbers were small. Conclusion Approximately 1/3 of GC isolates collected from MSM and 1/4 of GC isolates collected from transgender women were tetracycline resistant. Prevalence of tetracycline resistance was similar across anatomic sites. Providers should continue to follow recommended screening and treatment guidelines for gonorrhea among persons using doxycycline as STI PEP. Additionally, given high levels of TetHLR, enhanced tetracycline resistance surveillance is needed with implementation of doxycycline as STI PEP. Disclosures All Authors: No reported disclosures

Predominance of High-Level Tetracycline-Resistant Neisseria gonorrhoeae in Kenya: Implications for Global Implementation of Doxycycline Postexposure Prophylaxis for Prevention of Sexually Transmitted Infections.

Using archived Neisseria gonorrhoeae samples from 2008 to 2012, the prevalence of tet (M) genemediating high-level tetracycline resistance in N. gonorrhoeae was 96% among 50 Kenyan women. Determining the local and national prevalence of gonococcal tetracycline resistance and surveillance of gonococcal antimicrobial resistance can inform the implementation of doxycycline postexposure prophylaxis for STI prevention.

SCREENING OF ANTIBIOTIC RESISTANCE AND VIRULENCE GENES OF ENTEROCOCCUS SPP. STRAINS ISOLATED FROM URFA CHEESE

Enterococcus faecium, E. durans and E. faecalis species were isolated and identified from traditional Urfa cheese samples which were produced from unpasteurized raw milk. The enterococcal load of the cheese samples was between 4.4-5.6 log cfu g−1. High-level tetracycline, streptomycin, erythromycin, gentamycin, and penicillin resistance was determined in the enterococcal isolates. Multiple antibiotic resistance was also determined in E. faecalis (20.4%) and E. faecium (16.3%) strains. 36.7% of the enterococcal isolates were greater than 0.2 MAR index ratio in this study. The gelE and agg2 genes were found in 40 (81.63%) of the enterococcal isolates, whereas the vanB gene was found in 3 (6.12%) of the enterococcal isolates. The results indicate that the consumption of Urfa cheese, which is produced using raw milk, may have public health risk because of its antibiotic resistance characteristics and virulence genes of enterococcal biota.

Resistance Profile of Neisseria gonorrhoeae in KwaZulu-Natal, South Africa Questioning the Effect of the Currently Advocated Dual Therapy.

We report on the antimicrobial resistance profile of Neisseria gonorrhoeae isolates and the distribution of tetM genes in isolates with high-level tetracycline resistance in KwaZulu-Natal, South Africa. Male and female patients presenting with urethral and/or vaginal discharge were recruited into the study. Urethral and cervical secretions were cultured on New York City agar. Confirmatory tests for N. gonorrhoeae included Gram stain, catalase, oxidase, and carbohydrate utilization tests. Beta-lactamase was tested by means of the chromogenic cephalosporin test. Minimum inhibitory concentrations were determined using agar dilution with multipoint inoculation. Polymerase chain reaction with gel electrophoresis was used to detect the presence and type of the tetM gene. N. gonorrhoeae was isolated from the specimens of 319 (26%) of the 1220 recruited patients. Of these 319 isolates, 71% were resistant to 3 or more drugs. Resistance to azithromycin was found in 68% of the isolates. All isolates showed high-level tetracycline resistance with minimum inhibitory concentration values of 16 and 32 mg/mL. The tetM gene was present in 293 (92%). The American type was found in 264 (90%) and the Dutch type in 29 (10%). Twenty-six (8%) did not carry a tetM gene. The current syndromic management with dual ceftriaxone and azithromycin is due to the high level of azithromycin resistance factually single-drug therapy. High-level tetracycline resistance based on a resistance mechanism other than ribosome protection by the tetM gene product is present in N. gonorrhoeae infecting South African patients.

Prevalence and genetic profiling of tetracycline resistance (Tet-R) genes and transposable element (Tn916) in environmental Enterococcus species

Resistance against antimicrobial agents in enterococci is a global concern that not only challenges infection therapy but also make them reservoir of antibiotic resistance in human and animal alike. This study was conducted to establish tetracycline resistance profiles, prevalence of tet genes and transposable element (Tn916) in enterococcal soil and clinical isolates. Enterococci (n = 1210) from different environmental niche were collected and subjected to molecular identification. In total, 361 isolates showed tetracycline resistance at the breakpoint of 32 μg ml−1. MICs (32–512 μg ml−1) were established by both agar and micro-broth dilution methods. Soil isolates (n = 76) were further investigated for Tet genes (tet-A, C, K, L, M, S, O) and Tn916. Major resistance was observed in E. faecium 67% followed by E. faecalis 22%, E. hirae 8% and E. casseliflavus 2.6%. Results revealed that tet(L) was more frequently found in E. faecium 74.5%, while tet(M) was in high prevalence in E. faecalis 82.3%. Tn916 was detected in both clinical and soil isolates (i.e. 43.3% and 19.7%, respectively). RAPD-PCR analysis showed high diversity among the investigated isolates. Cumulatively, our results revealed high-level tetracycline resistance and the presence of multiple Tet genes and transposable element Tn916 in enterococci.

In Vitro Activity of Eravacycline against Carbapenem-Resistant Enterobacteriaceae and Acinetobacter baumannii.

Eravacycline and comparators were tested against carbapenem- and tigecycline-resistant Enterobacteriaceae and Acinetobacter isolates received at the United Kingdom's national reference laboratory. Eravacycline MICs correlated closely with those of tigecycline but mostly were around 2-fold lower; both molecules retained full activity against isolates with high-level tetracycline and minocycline resistance. MIC90s of eravacycline and tigecycline were raised ca. 2-fold for carbapenem-resistant Enterobacteriaceae compared with carbapenem-susceptible controls, probably reflecting subsets of isolates with increased efflux.

The Tetracycline Destructases: A Novel Family of Tetracycline-Inactivating Enzymes

Enzymes capable of inactivating tetracycline are paradoxically rare compared with enzymes that inactivate other natural-product antibiotics. We describe a family of flavoenzymes, previously unrecognizable as resistance genes, which are capable of degrading tetracycline antibiotics. From soil functional metagenomic selections, we discovered nine genes that confer high-level tetracycline resistance by enzymatic inactivation. We also demonstrate that a tenth enzyme, an uncharacterized homolog in the human pathogen Legionella longbeachae, similarly inactivates tetracycline. These enzymes catalyze the oxidation of tetracyclines in vitro both by known mechanisms and via previously undescribed activity. Tetracycline-inactivation genes were identified in diverse soil types, encompass substantial sequence diversity, and are adjacent to genes implicated in horizontal gene transfer. Because tetracycline inactivation is scarcely observed in hospitals, these enzymes may fill an empty niche in pathogenic organisms, and should therefore be monitored for their dissemination potential into the clinic.

Twenty-five-year changing pattern of gonococcal antimicrobial susceptibility in Shanghai: surveillance and its impact on treatment guidelines.

BackgroundAntimicrobial resistance of Neisseria gonorrhoeae is a serious health problem in China. Gonococcal antimicrobial susceptibility has been monitored in Shanghai since 1988. In this study, we examined the changing pattern of gonococcal antimicrobial susceptibility based on data from N. gonorrhoeae isolates collected over the past 25 years.MethodsApproximately 100–200 isolates each year (1988–2013) were tested for their susceptibility to penicillin (PEN), tetracycline (TET), ciprofloxacin (CIP), ceftriaxone (CRO) and spectinomycin (SPT), using the agar dilution method. Plasmid-mediated N. gonorrhoeae antimicrobial resistance, comprising penicillinase-producing N. gonorrhoeae (presumed PPNG) and high-level tetracycline resistance N. gonorrhoeae (presumed TRNG), were also determined. Breakpoints for susceptibilities followed those described by the Clinical and Laboratory Standard Institute and the European Committee on Antimicrobial Susceptibility Testing.ResultsA high proportion of isolates were resistant to PEN, TET and CIP, ranging from less than 20% at the beginning of the survey, increasing in the late 1990s and reaching over 90% in recent years. The proportion of isolates exhibiting plasmid-mediated resistance exceeded 38% for presumed PPNG and 20% for presumed TRNG in recent years. The proportion of CRO nonsusceptible isolates (MIC ≥ 0.125 mg/L) ranged from 7% to 13% in most of the study years. Almost all isolates were susceptible to SPT. The SPT MIC90 was 16–32 mg/L for 2008–2013. The proportion of CRO nonsusceptible-associated multiple-drug-resistant (MDR) isolates was over 5% in most of the study years.ConclusionsN. gonorrhoeae isolates in Shanghai were resistant to PEN, TET and CIP. Furthermore, CRO nonsusceptible and MDR isolates were prevalent. N. gonorrhoeae isolates were also found to be susceptible to SPT. It is recommended that the CRO dose be increased from currently recommended 250 mg to 500 mg and that SPT be an alternative in treating urogenital gonorrhea. Our findings highlight the importance of both regional and national surveillance programs for the prompt modification of treatment guidelines, vital in responding to the changing pattern of gonococcal antimicrobial susceptibility.Electronic supplementary materialThe online version of this article (doi:10.1186/s12879-014-0731-9) contains supplementary material, which is available to authorized users.

High-level tetracycline resistance mediated by efflux pumps Tet(A) and Tet(A)-1 with two start codons.

Efflux is the most common mechanism of tetracycline resistance. Class A tetracycline efflux pumps, which often have high prevalence in Enterobacteriaceae, are encoded by tet(A) and tet(A)-1 genes. These genes have two potential start codons, GTG and ATG, located upstream of the genes. The purpose of this study was to determine the start codon(s) of the class A tetracycline resistance (tet) determinants tet(A) and tet(A)-1, and the tetracycline resistance level they mediated. Conjugation, transformation and cloning experiments were performed and the genetic environment of tet(A)-1 was analysed. The start codons in class A tet determinants were investigated by site-directed mutagenesis of ATG and GTG, the putative translation initiation codons. High-level tetracycline resistance was transferred from the clinical strain of Klebsiella pneumoniae 10-148 containing tet(A)-1 plasmid pHS27 to Escherichia coli J53 by conjugation. The transformants harbouring recombinant plasmids that carried tet(A) or tet(A)-1 exhibited tetracycline MICs of 256-512 µg ml(-1), with or without tetR(A). Once the ATG was mutated to a non-start codon, the tetracycline MICs were not changed, while the tetracycline MICs decreased from 512 to 64 µg ml(-1) following GTG mutation, and to ≤4 µg ml(-1) following mutation of both GTG and ATG. It was presumed that class A tet determinants had two start codons, which are the primary start codon GTG and secondary start codon ATG. Accordingly, two putative promoters were predicted. In conclusion, class A tet determinants can confer high-level tetracycline resistance and have two start codons.

Antimicrobial susceptibility and genetic characterisation of oxytetracycline-resistant Edwardsiella tarda isolated from diseased eels

Edwardsiellosis is one of the most important bacterial diseases in eels. Edwardsiella tarda (E. tarda) isolates (n=94) from diseased eels were screened against the eight most commonly used antimicrobial agents...

P47 High-level tetracycline resistance caused by efflux pumps Tet(A) and Tet(A)-1 with two start codons

P47 High-level tetracycline resistance caused by efflux pumps Tet(A) and Tet(A)-1 with two start codons

Antimicrobial susceptibility patterns of gonococcal isolates in Pretoria, South Africa, over a 20-year period (1984-2004)

This paper reviews the susceptibility profles of Neisseria gonorrhoeae over a 20-year period in the Pretoria region. Endourethral specimens were collected from adult men with symptoms of urethritis attending primary health care clinics and private medical practitioners. These swabs were plated on enriched media for isolation of N. gonorrhoeae. Antimicrobial susceptibility of the organisms was performed using the disc diffusion and agar dilution methods. Plasmid analyses were performed on beta-lactamase-producing isolates. Penicillase-producing N. gonorrhoeae strains increased from 4% to 16%, whilst chromosomally mediated penicillin-resistant strains increased dramatically from 0% to 16% from 1984 to 2004. There was an equal distribution of the 3.2 MDa African and 4.4 MDa Asian plasmids. High-level tetracycline-resistant strains (36%) were detected for the frst time in 2004. Ciprofoxacin resistance emerged at 7% in the same year. Gonococcal isolates remained susceptible to cefoxitin, ceftriaxone, cefpodoxime, and spectinomycin. However, the minimum inhibitory concentration values for spectinomycin were very close to the breakpoint. We have shown a continuing increase in resistance to penicillin (plasmid and chromosomal), the emergence of high-level tetracycline resistance and an emergence of resistance to ciprofoxacin. Susceptibility testing is essential for successful therapeutic outcomes and needs to be performed in an ongoing basis.

Tetracycline resistance in Chilean clinical isolates of Helicobacter pylori

Since high-level tetracycline resistance in Helicobacter pylori has been associated with a AGA926-928-->TTC substitution in the 16S rRNA genes rrnA/B, the aim of the study was to screen for tetracycline resistance in H. pylori clinical isolates obtained from Santiago, Chile by using a recently reported molecular assay. A PCR-restriction fragment length polymorphism (PCR-RFLP) assay of the conserved 535 bp region of the H. pylori 16S rRNA genes rrnA/B (between nucleotides 710 and 1245) using HinfI was followed by DNA sequencing of the same fragment obtained from tetracycline-resistant H. pylori clinical isolates. The PCR-RFLP assay revealed that the tetracycline-resistant H. pylori isolates lacked the AGA926-928-->TTC substitution. In contrast, DNA sequencing of the 535 bp PCR fragment from 11 tetracycline-resistant H. pylori Chilean clinical isolates showed an association of low-level tetracycline resistance with 1 bp (A928C) or 2 bp (AG926-927-->GT and/or A926G/A928C) substitutions in both 16S rRNA genes. The PCR-RFLP (HinfI) assay alone is unreliable for the detection of tetracycline resistance in Chilean clinical isolates of H. pylori. To that end, it must be complemented by sequencing of the 535 bp PCR fragment.

English

In India antibiotics are frequently used for preventing and controlling bacterial pathogens in carp aquaculture system, yet no studies have been performed to evaluate the ecological impact of its intensive and prolonged use. In this work the frequency of oxytetracycline-resistant bacteria from water, palletized feed and different life stages of fish from Indian freshwater carp aquaculture system as well as the level of resistance of selected strains was investigated. Viable as well as antibiotic-resistant bacterial counts were performed by spread plate method in culture media supplemented with the oxytetracycline. Sixty two resistant Gram negative isolates which represented the oxytetracycline-resistant bacterial population, were randomly selected on nutrient agar supplemented with oxytetracycline (50μg/ml) from carp farms and feed pellet samples. Among these bacterial isolates Flavobacterium (21%), Alcaligenes (14.5%), Aeromonas (11%), Pseudomonas (10%) and Enterobacteriace (19%) were the most frequent. The Escherichia, Serratia, Citrobacter, Enterobacter, Shigella and Proteus from Enterobacteriace were recovered. Twelve isolates of oxytetracycline resistant bacteria were mainly dominated in adult fishes by the genus Flavobacterium (23%) and Enterobacteriace(41%). Selected strains exhibited high levels of oxytetracycline resistance with minimum inhibitory concentrations (MICs) ranging from 50 to 600μg/ml.This study shows the presence of an important population of oxytetracycline-resistant bacteria in the microflora of Indian carp aquaculture farms. Therefore the environment of these farms might play important roles as reservoirs of bacteria carrying genetic determinants for high level tetracycline resistance, prompting an important risk to public health.

Multidrug-resistant Providencia stuartii expressing extended-spectrum -lactamase PER-1, originating in Kosovo

triple-base-pair mutation involved in the high-level tetracycline resistance in H. pylori, whereas nucleotide 1054 contacts the A-site tRNA. In contrast, the G346A mutation is located in a 16S rRNA region not closely associated with tetracycline binding. For M. pneumoniae, two mutations were found in the brothselected mutants and were associated with 2to 16-fold increases in MICs of the three tetracyclines studied, in comparison with those of the parental strain FH. The two mutations described for this species, G1193A and T968C, are located very close to or contact the primary tetracycline binding site, respectively. In summary, this is the first description of mutations in 16S rRNA associated with decreased susceptibility to tetracyclines in human mycoplasmas. However, what real effect these new mutations have on the tetracycline susceptibility of both mycoplasmas has yet to be determined.

A Combination ofsbmAandtolCMutations inEscherichia coliK-12 Tn10-Carrying Strains Results in Hypersusceptibility to Tetracycline

Previously, we demonstrated that Escherichia coli tolC mutations reduce the high-level resistance to tetracycline afforded by the transposon Tn10-encoded TetA pump from resistance at 200 microg/ml to resistance at 40 microg/ml. In this study, we found that the addition of an sbmA mutation to a tolC::Tn10 mutant exacerbates this phenotype: the double mutant did not form colonies, even in the presence of tetracycline at a concentration as low as 5 microg/ml. Inactivation of sbmA alone partially inhibited high-level tetracycline resistance, from resistance at 200 microg/ml to resistance at 120 microg/ml. There thus appears to be an additive effect of the mutations, resulting in almost complete suppression of the phenotypic expression of Tn10 tetracycline resistance.

Analysis of trends in antimicrobial resistance in Neisseria gonorrhoeae isolated in Australia, 1997 2006

To analyse trends in antimicrobial resistance (AMR) in Neisseria gonorrhoeae (GC) isolated in Australia between 1997 and 2006 and to identify factors influencing emergence and spread of AMR in GC. AMR data were generated in reference laboratories in each state and territory of Australia using the methods of the Australian Gonococcal Surveillance Programme from a comprehensive sample of GC. Trends in the proportion of strains resistant to penicillin, ciprofloxacin, spectinomycin and ceftriaxone or with high-level tetracycline resistance (TRNG) were determined from aggregated national data and were also disaggregated by region. Further analyses of additional AMR, demographic, transmission and antibiotic use data were also performed. More than 36,000 GC were examined. Significant increases in resistance to penicillin and ciprofloxacin and in TRNG occurred in national data and in urban populations. Approximately half of the GC tested in larger urban centres were penicillin and/or ciprofloxacin resistant by 2006. These high rates of resistance arose despite low (penicillin) or absent (ciprofloxacin) exposure. In contrast, in rural and remote areas with very high disease rates and high rates of penicillin use, <5% of GC tested were penicillin (or quinolone) resistant. No spectinomycin-resistant GC were detected. Low numbers of GC with raised MICs of ceftriaxone were present in urban centres each year from 2001 onwards. Significant increases in AMR in GC occurred in parts of Australia in the 10 years to 2006. The data suggest that the AMR seen in GC in urban populations were the result of their repeated importation into Australia and ultimate introduction into established sexual networks rather than originating de novo or as a result of selection by antibiotic use or misuse.

Gonococcal resistance: evolving from penicillin, tetracycline to the quinolones in South Africa – implications for treatment guidelines

Resistant Neisseria gonorrhoeae has been evolving. This study assessed the antimicrobial susceptibility profile of isolates in the Pretoria region, South Africa. Isolates of N. gonorrhoeae from men with urethritis were tested for susceptibility to eight antimicrobial agents by disc diffusion, Etest and agar dilution methods. Chromosomal resistance to penicillin was found in 16% of isolates, 16% showed plasmid-mediated resistance and decreased susceptibility was seen in 73% of isolates. For the first time, there is evidence of high-level tetracycline resistance (36%). Ciprofloxacin resistance emerged at 7%. All isolates remained susceptible to ceftriaxone. In view of these findings of the emergence of quinolone-resistant N. gonorrhoeae, national treatment guidelines for syndromic management of sexually transmitted infections need to be urgently reviewed. The injectable preparation, ceftriaxone has to be considered as a first-line agent for the management of gonococcal infections. Overall, the gonococcal isolates in the Pretoria region remain susceptible to ceftriaxone, cefoxitin, cefpodoxime and spectinomycin.

Role of the plasmid-encoded tet(O) gene in tetracycline-resistant clinical isolates of Campylobacter jejuni and Campylobacter coli

The prevalence of tetracycline resistance, tetracycline MICs and tet(O) gene localization were investigated in 83 Campylobacter isolates from patients suffering from acute gastroenteritis in Germany. Combined biochemical and molecular markers identified 74 isolates (89 %) as Campylobacter jejuni, including seven atypical isolates that failed to hydrolyse hippurate, and nine isolates (11 %) as Campylobacter coli. Tetracycline resistance was detected in six out of nine Campylobacter coli isolates (67 %) and 13 out of 74 C. jejuni isolates (18 %). Low-level tetracycline resistance was observed for C. coli (MIC 16 microg ml(-1) for all strains), whereas C. jejuni showed high-level resistance (MIC >256 microg ml(-1) for all strains). Both low- and high-level tetracycline resistance was associated with the presence of the tet(O) gene. In C. jejuni, tet(O) was plasmid-encoded in 54 % of tetracycline-resistant isolates, whereas in C. coli, tet(O) appeared to be located on the chromosome.