<i>In Vitro</i>
            Efficacy of Gentamicin Alone and in Combination with Ceftriaxone, Ertapenem, and Azithromycin against Multidrug-Resistant Neisseria gonorrhoeae

Li, Xuechun; Le, Wei; Lou, Xiangdi; Wang, Biwei; Genco, Caroline Attardo; Rice, Peter A.; Su, Xiaohong

doi:10.1128/spectrum.00181-21

Cited by 7 publications

(9 citation statements)

References 36 publications

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“…54 Over time, N. gonorrhoeae acquired resistance to these antimicrobials due to cumulative mutations in the PBPs and multidrug MtrCDE efflux pumps. 55 In 2018, an extreme drug-resistant gonococcal strain was reported in the UK, requiring 6,40 the search for a newer set of targets and inhibitors. The NgA, which remained highly conserved within the N. gonorrhoeae pathogen sequence database, appears to be serving the purpose (Figures 1B and 5A).…”

Section: Discussionmentioning

confidence: 99%

“…N. gonorrhoeae was initially cultured on Thayer Martin agar (HiMedia Laboratories), a selective media supplemented with vancomycin, colistin, nystatin, and trimethoprim for screening and maintaining pure N. gonorrhoeae colonies. For further experiments, N. gonorrhoeae was preinoculated in GC chocolate agar (HiMedia Laboratories) supplemented with 2% haemoglobin and vitamin growth supplement (HiMedia Laboratories) at 37°C in a humid candle jar overnight 40 . The inhibitory concentration (IC 50 ) for N2, N8, and N9 was studied by broth microdilution method using 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) and resazurin‐based cell viability assay (REMA) described earlier 21,41,42 .…”

Section: Methodsmentioning

confidence: 99%

“…For further experiments, N. gonorrhoeae was preinoculated in GC chocolate agar (HiMedia Laboratories) supplemented with 2% haemoglobin and vitamin growth supplement (HiMedia Laboratories) at 37°C in a humid candle jar overnight. 40 The inhibitory concentration (IC 50 ) for N2, N8, and N9 was studied by broth microdilution method using 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT) and resazurin-based cell viability assay (REMA) described earlier. 21,41,42 Ceftriaxone (Caymen Chemical) used in treating gonorrhoea infections was utilised as a control in the experiments.…”

Section: Cell Viability Assaymentioning

confidence: 99%

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Antimicrobial‐resistant Neisseria gonorrhoeae can be targeted using inhibitors against evolutionary conservedl‐asparaginase

Vashishtha

Singh

Kundu

2022

J of Cellular Biochemistry

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The emergence of multiple drug‐resistant “super gonorrhoea” complicates the management and treatment of Neisseria gonorrhoeae infections due to the progressive accumulation of mutations in the biological targets of frontline antimicrobials. Continuous evaluation and reporting of newer molecular targets and their inhibitors are necessary. Here, we present l‐asparaginase of N. gonorrhoeae (NgA) as a new molecular target based on structure‐based high‐throughput screening, molecular dynamics(MD) simulations, and validation by biophysical, biochemical, and cell viability assays. We observed that the NgA is evolutionarily conserved in both the drug‐resistant and susceptible strains of N. gonorrhoeae, indicating its importance in the growth and survival of the pathogen. Three Food and Drug Administration‐approved drugs, pemirolast, thalidomide, and decitabine, were identified as potential inhibitors of NgA using high‐throughput screening. The binding energies of the drugs with NgA were −20.14, −19.67, and −16.47 kcal/mol, respectively, compared to −6.82 ± 1.46 for enzyme–substrate l‐Asn, as obtained through MD simulations. Subsequently, fluorescence quenching and differential scanning calorimetry experiments validated the in silico data. The observance of inhibition of NgA activity at micromolar drug concentrations further strengthened our findings. Conclusive evidence came from the cell viability assays where these drugs were found to impede the growth of N. gonorrhoeae culture effectively. Thus, our study establishes l‐asparaginase as a new molecular target against gonococcal infections. From this study, we propose that targeting of NgA can be explored to control N. gonorrhoeae infections in combination therapy.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Cell Viability Assaymentioning

confidence: 99%

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Antimicrobial‐resistant Neisseria gonorrhoeae can be targeted using inhibitors against evolutionary conservedl‐asparaginase

Vashishtha

Singh

Kundu

2022

J of Cellular Biochemistry

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“…By the agar dilution method, we found that 36.60% of the isolates were fully susceptible to gentamicin, while 61.30% of the strains showed intermediate susceptibility. In two other studies from China on N. gonorrhoeae susceptibility to gentamicin, moderate susceptibility rates of 97.8% [ 25 ] and 91.6% [ 21 ] were also reported. The possible reason was that these two studies were both from eastern provinces of China, while the isolates of our study were from the eastern, northern and western regions of China.…”

Section: Discussionmentioning

confidence: 99%

“…The criteria for azithromycin and ceftriaxone susceptibility and resistance were interpreted based on the Clinical and Laboratory standards Institute (CLSI) M100-S29 breakpoints [ 20 ]. According to previous studies and interpretation criteria, the breakpoint for gentamicin MIC was set to be ≤4 mg/L as fully susceptible, 8 to 16 mg/L as intermediately susceptible, and ≥32 mg/L as resistant [ 14 , 21 ].…”

Section: Methodsmentioning

confidence: 99%

In Vitro Study of the Interaction of Gentamicin with Ceftriaxone and Azithromycin against Neisseria gonorrhoeae Using Agar Dilution Method

Zhou

Liu

et al. 2022

Antibiotics

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The susceptibility to gentamicin of N. gonorrhoeae isolates collected in three Chinese provinces and the correlation among the MICs of gentamicin, azithromycin, and ceftriaxone were investigated in this study. The effects of combinations from those three antibiotics were also in the scope of this study to determine the efficacy of gentamicin as a combination therapeutic drug. The agar dilution method was used to measure the minimum inhibitory concentrations (MICs) of ceftriaxone, azithromycin and gentamicin on N. gonorrhoeae isolates. The synergy between these three antimicrobials were determined using the agar dilution checkerboard method. Subgroup studies were conducted to explore differences between azithromycin- and ceftriaxone-sensitive and resistant isolates. A total of 139 (36.60%) and 233 (61.30%) isolates demonstrated full susceptibility and intermediate susceptibility to gentamicin, respectively. The correlation analysis showed that the MICs of ceftriaxone and azithromycin weakly correlated with the value of gentamicin. The overall results of the three antibiotic combinations revealed indifferent effects. Combination therapy established a significant reduction on the MIC value. Most of the N. gonorrhoeae isolates tested in this study demonstrated a certain degree of susceptibility to gentamicin. Overall, antimicrobial combinations of gentamicin with ceftriaxone or azithromycin demonstrate indifferent effects.

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Evolutionarily conserved heat shock protein, HtpX, as an adjunct target against antibiotic‐resistant Neisseria gonorrhoeae

Vashishtha,

Thakur,

Singh

et al. 2023

J of Cellular Biochemistry

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The emergence of multiple drug resistance and extreme drug resistance pathogens necessitates the continuous evaluation of the pathogenic genome to identify conserved molecular targets and their respective inhibitors. In this study, we mapped the global mutational landscape of Neisseria gonorrhoeae (an intracellular pathogen notoriously known to cause the sexually transmitted disease gonorrhoea). We identified highly variable amino acid positions in the antibiotic target genes like the penA, ponA, 23s rRNA, rpoB, gyrA, parC, mtrR and porB. Some variations are directly reported to confer resistance to the currently used front‐line drugs like ceftriaxone, cefixime, azithromycin and ciprofloxacin. Further, by whole genome comparison and Shannon entropy analysis, we identified a completely conserved protein HtpX in the drug‐resistant as well as susceptible isolates of N. gonorrhoeae (NgHtpX). Comparison with the only available information of Escherichia coli HtpX suggested it to be a transmembrane metalloprotease having a role in stress response. The critical zinc‐binding residue of NgHtpX was mapped to E141. By applying composite high throughput screening followed by MD simulations, we identified pemirolast and thalidomide as high‐energy binding ligands of NgHtpX. Following cloning and expression of the purified metal‐binding domain of NgHtpX (NgHtpXd), its Zn2+‐binding (Kd = 0.4 µM) and drug‐binding (pemirolast, Kd = 3.47 µM; and thalidomide, Kd = 1.04 µM) potentials were determined using in‐vitro fluorescence quenching experiment. When tested on N. gonorrhoeae cultures, both the ligands imposed a dose‐dependent reduction in viability. Overall, our results provide high entropy positions in the targets of presently used antibiotics, which can be further explored to understand the AMR mechanism. Additionally, HtpX and its specific inhibitors identified can be utilised effectively in managing gonococcal infections.

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In Vitro Efficacy of Gentamicin Alone and in Combination with Ceftriaxone, Ertapenem, and Azithromycin against Multidrug-Resistant Neisseria gonorrhoeae

Abstract: Antimicrobial-resistant Neisseria gonorrhoeae is a major global public health concern. New treatment options are urgently needed to successfully treat multidrug-resistant (MDR) Neisseria gonorrhoeae infections.

Cited by 7 publications

References 36 publications

Antimicrobial‐resistant Neisseria gonorrhoeae can be targeted using inhibitors against evolutionary conservedl‐asparaginase

Antimicrobial‐resistant Neisseria gonorrhoeae can be targeted using inhibitors against evolutionary conservedl‐asparaginase

In Vitro Study of the Interaction of Gentamicin with Ceftriaxone and Azithromycin against Neisseria gonorrhoeae Using Agar Dilution Method

Evolutionarily conserved heat shock protein, HtpX, as an adjunct target against antibiotic‐resistant Neisseria gonorrhoeae

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