Response mechanisms of resistance in L-form bacteria to different target antibiotics: Implications from oxidative stress to metabolism.

Due to the specific action on bacterial cell wall, ß-lactam antibiotics have gained widespread usage as they exhibit a high degree of specificity in targeting bacteria, but causing minimal toxicity to host cells. Under antibiotic pressure, bacteria may opt to shed their cell walls and transform into L-form state as a means to evade the antibiotic effects. In this study, we explored and identified diverse optimal conditions for both Gram-negative bacteria (E. coli DH5α (CTX)) and Gram-positive bacteria (B. subtilis ATCC6633), which were induced to L-form bacteria using lysozyme (0.5 ppm) and meropenem (64 ppm). Notably, when bacteria transformed into L-form state, both bacterial strains showed varying degrees of increased resistance to antibiotics polymyxin E, meropenem, rifampicin, and tetracycline. E. coli DH5α (CTX) exhibited the most significant enhancement in resistance to tetracycline, with a 128-fold increase, while B. subtilis ATCC6633 showed a 32-fold increase in resistance to tetracycline and polymyxin E. Furthermore, L-form bacteria maintained their normal metabolic activity, combined with enhanced oxidative stress, served as an adaptive strategy promoting the sustained survival of L-form bacteria. This study provided a theoretical basis for comprehending antibiotic resistance mechanisms, developing innovative treatment strategies, and confronting global antibiotic resistance challenges.

Pre-exposure of Triclosan compromise tetracycline-derived antibiotic resistance in methanogenic digestion microbiome.

Triclosan (TCS) and tetracycline (TC) are commonly detected antibacterial agents in sewage and environment matrices. Nonetheless, the impact of sequential exposure to TCS and TC on the methanogenic digestion microbiome remains unknown. In this study, TCS was shown to reduce COD removal efficiency to 69.8%, but alleviated the inhibitive effect of consequent TC-amendment on the digestion microbiome. Interestingly, TCS pre-exposure resulted in abundance increase of acetotrophic Methanosaeta to 2.68%, being 2.91 folds higher than that without TCS amendment. Microbial network analyses showed that TCS pre-exposure caused microorganisms to establish a co-ecological relationship against TC disturbance. Further analyses of total antibiotic resistance genes (ARGs) showed the TCS-derived compromise of TC-induced ARGs enrichment in digestion microbiomes, e.g., 238.2% and 152.1% ARGs increase upon TC addition in digestion microbiomes without and with TCS pre-exposure, respectively. This study provides new insights into the impact of antibacterial agents on the methanogenic digestion microbiome.

45 years of tetracycline post exposure prophylaxis for STIs and the risk of tetracycline resistance: a systematic review and meta-analysis.

There is considerable interest in the use of doxycycline post exposure prophylaxis (PEP) to reduce the incidence of bacterial sexually transmitted infections (STIs). An important concern is that this could select for tetracycline resistance in these STIs and other species. We searched PubMed and Google Scholar, (1948-2023) for randomized controlled trials comparing tetracycline PEP with non-tetracycline controls. The primary outcome was antimicrobial resistance (AMR) to tetracyclines in all bacterial species with available data. Our search yielded 140 studies, of which three met the inclusion criteria. Tetracycline PEP was associated with an increasedprevalence of tetracycline resistance in Neisseria gonorrhoeae, but this effect was not statistically significant (Pooled OR 2.3, 95% CI 0.9-3.4). PEP had a marked effect on the N. gonorrhoeae tetracycline MIC distribution in the one study where this was assessed. Prophylactic efficacy was 100% at low MICs and 0% at high MICs. In the one study where this was assessed, PEP resulted in a significant increase in tetracycline resistance in commensal Neisseria species compared to the control group (OR 2.9, 95% CI 1.5-5.5) but no significant effect on the prevalence of tetracycline resistance in Staphylococcus aureus. The available evidence suggests that PEP with tetracyclines could be associated with selecting tetracycline resistance in N. gonorrhoeae and commensal Neisseria species.

Riboswitch-controlled IL-12 gene therapy reduces hepatocellular cancer in mice.

Hepatocellular carcinoma (HCC) and solid cancers with liver metastases are indications with high unmet medical need. Interleukin-12 (IL-12) is a proinflammatory cytokine with substantial anti-tumor properties, but its therapeutic potential has not been realized due to severe toxicity. Here, we show that orthotopic liver tumors in mice can be treated by targeting hepatocytes via systemic delivery of adeno-associated virus (AAV) vectors carrying the murine IL-12 gene. Controlled cytokine production was achieved in vivo by using the tetracycline-inducible K19 riboswitch. AAV-mediated expression of IL-12 led to STAT4 phosphorylation, interferon-γ (IFNγ) production, infiltration of T cells and, ultimately, tumor regression. By detailed analyses of efficacy and tolerability in healthy and tumor-bearing animals, we could define a safe and efficacious vector dose. As a potential clinical candidate, we characterized vectors carrying the human IL-12 (huIL-12) gene. In mice, bioactive human IL-12 was expressed in a vector dose-dependent manner and could be induced by tetracycline, suggesting tissue-specific AAV vectors with riboswitch-controlled expression of highly potent proinflammatory cytokines as an attractive approach for vector-based cancer immunotherapy.

Machine learning reveals the selection pressure exerted by nonantibiotic pharmaceuticals at environmentally relevant concentrations on antibiotic resistance genotypes.

The emergence and increasing prevalence of antibiotic resistance pose a global public risk for human health, and nonantimicrobial pharmaceuticals play an important role in this process. Herein, five nonantimicrobial pharmaceuticals, including acetaminophen (ACT), clofibric acid (CA), carbamazepine (CBZ), caffeine (CF) and nicotine (NCT), tetracycline-resistant strains, five ARGs (sul1, sul2, tetG, tetM and tetW) and one integrase gene (intI1), were detected in 101 wastewater samples during two typical sewage treatment processes including anaerobic-oxic (A/O) and biological aerated filter (BAF) in Harbin, China. The impact of nonantibiotic pharmaceuticals at environmentally relevant concentrations on both the resistance genotypes and resistance phenotypes were explored. The results showed that a significant impact of nonantibiotic pharmaceuticals at environmentally relevant concentrations on tetracycline resistance genes encoding ribosomal protection proteins (RPPs) was found, while no changes in antibiotic phenotypes, such as minimal inhibitory concentrations (MICs), were observed. Machine learning was applied to further sort out the contribution of nonantibiotic pharmaceuticals at environmentally relevant concentrations to different ARG subtypes. The highest contribution and correlation were found at concentrations of 1400-1800 ng/L for NCT, 900-1500 ng/L for ACT and 7000-10,000 ng/L for CF for tetracycline resistance genes encoding RPPs, while no significant correlation was found between the target compounds and ARGs when their concentrations were lower than 500 ng/L for NCT, 100 ng/L for ACT and 1000 ng/L for CF, which were higher than the concentrations detected in effluent samples. Therefore, the removal of nonantibiotic pharmaceuticals in WWTPs can reduce their selection pressure for resistance genes in wastewater.

Refractoriness to anti-Helicobacter pylori treatment attributed to phenotypic resistance patterns in patients with gastroduodenopathy in Guayaquil-Ecuador.

BACKGROUND: Treatment of Helicobacter pylori gastric infection is complex and associated with increased rates of therapeutic failure. This research aimed to characterize the H. pylori infection status, strain resistance to antimicrobial agents, and the predominant lesion pattern in the gastroduodenal mucosa of patients with clinical suspicion of refractoriness to first- and second-line treatment who were diagnosed and treated in a health center in Guayaquil, Ecuador. METHODS: A total of 374 patients with upper gastrointestinal symptoms and H. pylori infection were preselected and prescribed one of three triple therapy regimens for primary infection, as judged by the treating physician. Subsequently, 121 patients who returned to the follow-up visit with persistent symptoms after treatment were studied. RESULTS: All patients had H. pylori infection. Histopathological examination diagnosed chronic active gastritis in 91.7% of cases; premalignant lesions were observed in 15.8%. The three triple therapy schemes applied showed suboptimal efficacy (between 47.6% and 77.2%), with the best performance corresponding to the scheme consisting of a proton pump inhibitor + amoxicillin + levofloxacin. Bacterial strains showed very high phenotypic resistance to all five antimicrobials tested: clarithromycin, 82.9%; metronidazole, 69.7%; amoxicillin and levofloxacin, almost 50%; tetracycline, 38.2%. Concurrent resistance to clarithromycin-amoxicillin was 43.4%, to tetracycline-metronidazole 30.3%, to amoxicillin-levofloxacin 27.6%, and to clarithromycin-metronidazole 59.2%. CONCLUSIONS: In vitro testing revealed resistance to all five antibiotics, indicating that H. pylori exhibited resistance phenotypes to these antibiotics. Consequently, the effectiveness of triple treatments may be compromised, and further studies are needed to assess refractoriness in quadruple and concomitant therapies.

Detoxification of tetracycline and synthetic dyes by a newly characterized Lentinula edodes laccase, and safety assessment using proteomic analysis.

Fungal laccase has strong ability in detoxification of many environmental contaminants. A putative laccase gene, LeLac12, from Lentinula edodes was screened by secretome approach. LeLac12 was heterogeneously expressed and purified to characterize its enzymatic properties to evaluate its potential use in bioremediation. This study showed that the extracellular fungal laccase from L. edodes could effectively degrade tetracycline (TET) and the synthetic dye Acid Green 25 (AG). The growth inhibition of Escherichia coli and Bacillus subtilis by TET revealed that the antimicrobial activity was significantly reduced after treatment with the laccase-HBT system. 16 transformation products of TET were identified by UPLC-MS-TOF during the laccase-HBT oxidation process. Gas chromatography-mass spectrometry (GC-MS) analysis revealed that LeLac12 could completely mineralize ring-cleavage products. LeLac12 completely catalyzed 50 mg/L TET within 4 h by adding AG (200 mg/L), while the degradation of AG was above 96% even in the co-contamination system. Proteomic analysis revealed that central carbon metabolism, energy metabolism, and DNA replication/repair were affected by TET treatment and the latter system could contribute to the formation of multidrug-resistant strains. The results demonstrate that LeLac12 is an efficient and environmentally method for the removal of antibiotics and dyes in the complex polluted wastewater.

Powdered activated carbon facilitated degradation of complex organic compounds and tetracycline in stressed anaerobic digestion systems.

Tetracycline exerts an inhibitory effect on anaerobic digestion, inducing stressed microbial activities and even system failure. Continuous-flow reactors (CFRs) and sequencing batch reactors (SBRs) were employed along with the dosage of powdered activated carbon (PAC) to enhance tetracycline removal during anaerobic digestion of complex organic compounds. PAC increased the maximum methane production rate by 15.6% (CFRs) and 13.8% (SBRs), and tetracycline biodegradation by 24.4% (CFRs) and 19.2% (SBRs). CFRs showed higher tetracycline removal and methane production rates than SBRs. Geobacter was enriched in CFRs, where Methanothrix was enriched with the addition of PAC. Desulfomicrobium harbored abundant propionate degradation-related genes, significantly correlating with tetracycline removal. The genes encoding carbon dioxide reduction in Methanothrix along with the detection of Geobacter might indicate direct interspecies electron transfer for methanogenesis in CFRs and PAC-added reactors. The study offers new insights into anaerobic digestion under tetracycline-stressed conditions and strategies for optimizing tetracycline removal.

Facile fabrication of Ag decorated MnFeO3 catalyst: Comparative analysis of visible light driven antibiotic reduction and antibacterial performance.

Photocatalysis is an effective method with the potential to eliminate pharmaceutical compounds from water sources. Manganese ferrite (MnFeO3), a type of multiferroic perovskite catalyst, has attracted significant attention due to its small band gap, however its application was limited due to its high recombination rate and low quantum efficiency. It was therefore aimed to improve the properties of MnFeO3 by doping silver (Ag)-particles. In this study, Ag-MnFeO3 photocatalysts with different Ag content (1-3 mmol%) were synthesized by performing a facile hydrothermal method. The as-prepared samples were characterized using x-ray diffraction (XRD), scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM/EDS), transmission electron microscopy (TEM), x-ray photoelectron spectroscopy (XPS), ultraviolet-visible spectroscopy (DRS), photoluminescence spectroscopy (PL), electrochemical impedance spectroscopy (EIS) and Brunauer-Emmett-Teller (BET) method, showing successful addition of Ag-particles with the MnFeO3 structure. Then, the as-synthesized materials were investigated as: (i) photocatalysts for degradation tetracycline (TC) antibiotic and (ii) antibacterial agents for bacteria. The Ag-MnFeO3 catalyst demonstrated superior catalytic performance (95.7%), which was 1.6 times higher than that of pristine MnFeO3 (59.7%). The positive effect was ascribed to oxygen vacancies, enhanced light absorption ability, and lower recombination rate. The Ag-MnFeO3 catalyst also showed satisfactory removal performances in real water matrices. Furthermore, radical trapping tests depicted that the superoxide radicals played a dominant role in the photodegradation system. In addition, Box-Behnken design (BBD) was performed to determine the optimum conditions, which were determined as catalyst dosage of 0.45 g/L, initial TC concentration of 5.10 mg/L, and initial solution pH value of 3.69. In terms of antibacterial tests, the incorporation of Ag into the MnFeO3 structure greatly increased the antimicrobial resistance against bacteria. Our findings disclose that the incorporation of Ag into the MnFeO3 structure can be regarded as a feasible and promising approach to improve both photocatalytic degradation and antibacterial performances.

Community coalescence and plant host filtering determine the spread of tetracycline resistance genes from pig manure into the microbiome continuum of the soil-plant system.

The spread of livestock manure-borne antibiotic resistance genes (ARGs) into agroecosystems through manure application poses a potential threat to human health. However, there is still a knowledge gap concerning ARG dissemination in coalescing manure, soil and plant microbiomes. Here, we examined the fate of tetracycline resistance genes (TRGs) originating from pig manure microbiomes and spread in the soil-A thaliana system and explored the effects of microbial functions on TRGs spread at different interfaces. Our results indicate that the TRGs abundances in all microbiome continuum of the soil-A. thaliana system were significantly increased with the application of a living manure microbiome, although the addition of manure with both an active and inactive microbiome caused a shift in the microbial community composition. This was attributed to the increasing relative abundances of tetA, tetL, tetM, tetO, tetW and tolC in the system. The application of living manure with DOX residues resulted in the highest relative abundance of total TRGs (3.30×10-3 copies/16S rRNA gene copies) in the rhizosphere soil samples. Community coalescence of the manure and soil microbiomes increased the abundance of Firmicutes in the soil and root microbiome, which directly explains the increase in TRG abundance observed in these interfaces. In contrast, the leaf microbiome differed markedly from that of the remaining samples, indicating strong plant host filtering effects on Firmicutes and TRGs from pig manure. The random forest machine learning model revealed microbial functions and their significant positive correlation with TRG abundance in the microbiome continuum of the system. Our findings revealed that community coalescence is the main driver of TRG spread from manure to the soil and root microbiomes. Plant host filtering effects play a crucial role in allowing certain microbial groups to occupy ecological niches in the leaves, thereby limiting the establishment of manure-borne TRGs in aboveground plant tissues.

Effects of four antibiotics on the photosynthetic light reactions in the green alga Chlorella pyrenoidosa.

Antibiotics are ubiquitously present in aquatic environments, posing a serious ecological risk to aquatic ecosystems. However, the effects of antibiotics on the photosynthetic light reactions of freshwater algae and the underlying mechanisms are relatively less understood. In this study, the effects of 4 representative antibiotics (clarithromycin, enrofloxacin, tetracycline, and sulfamethazine) on a freshwater alga (Chlorella pyrenoidosa) and the associated mechanisms, primarily focusing on key regulators of the photosynthetic light reactions, were evaluated. Algae were exposed to different concentrations of clarithromycin (0.0-0.3 mg/L), enrofloxacin (0.0-30.0 mg/L), tetracycline (0.0-10.0 mg/L), and sulfamethazine (0.0-50.0 mg/L) for 7 days. The results showed that the 4 antibiotics inhibited the growth, the photosynthetic pigment contents, and the activity of antioxidant enzymes. In addition, exposure to clarithromycin caused a 118.4 % increase in malondialdehyde (MDA) levels at 0.3 mg/L. Furthermore, the transcripts of genes for the adenosine triphosphate (ATP) - dependent chloroplast proteases (ftsH and clpP), genes in photosystem II (psbA, psbB, and psbC), genes related to ATP synthase (atpA, atpB, and atpH), and petA (related to cytochrome b6/f complex) were altered by clarithromycin. This study contributes to a better understanding of the risk of antibiotics on primary producers in aquatic environment.

Antimicrobial Resistance Patterns in Patients with Vaginal Discharge: A 2019-2022 Analysis at the National Health Laboratory in Eritrea.

Background: Antimicrobial resistance poses a significant global threat to the treatment of bacterial infections, particularly in low- and middle-income regions such as Africa. This study is aimed at analyzing antimicrobial resistance patterns in vaginal swab samples from patients at the National Health Laboratory from 2019 to 2022. Methods: This retrospective study examined patient records from vaginal swab analyses performed at the National Health Laboratory between January 1, 2019, and December 31, 2022. Ethical approval was obtained from the Ministry of Health Research Ethical Approval and Clearance Committee on 15/02/2023. Results: Of the 622 samples, 83% underwent microbial isolation and identification. Citrobacter spp. exhibited high resistance (>43%) to antibiotics such as cephalexin, ceftazidime, nalidixic acid, ampicillin, gentamicin, and tetracycline. E. coli showed resistance rates of more than 50% to ampicillin, trimethoprim-sulfamethoxazole, and tetracycline. Klebsiella spp. and Proteus spp. exhibited resistance rates that exceeded 47% to specific antibiotics. Gram-positive bacteria have resistance rates of more than 49% with ampicillin, trimethoprim-sulfamethoxazole, tetracycline, oxacillin, vancomycin, and penicillin G. In particular, S. aureus demonstrated no resistance to rifampicin or clindamycin, while Streptococcus spp. showed 100% resistance to rifampicin and vancomycin. Several species, including Proteus species, Streptococcus spp., S. aureus, and Klebsiella spp. exhibited multidrug resistance. Conclusion: Most gram-negative bacteria displayed higher resistance of >45% to ampicillin, trimethoprim-sulfamethoxazole, and tetracycline. Among gram-positive bacteria, a higher resistance rate with ampicillin, trimethoprim-sulfamethoxazole, tetracycline, oxacillin, vancomycin, and penicillin G was recorded. S. aureus showed no resistance to rifampicin and clindamycin, and Strep. spp. indicated 100% resistance to rifampicin and vancomycin. This study highlights critical gaps and areas for further exploration. Expanding the spectrum of antibiotics tested and investigating underlying multidrug resistance mechanisms would provide a more comprehensive understanding of resistance patterns.

Possibility of transfer and activation of 'silent' tetracycline resistance genes among Enterococcus faecalis under high-pressure processing.

In this study, the tetracycline resistance of Enterococcus faecalis strains isolated from food was determined and molecular analyses of the resistance background were performed by determining the frequency of selected tetracycline resistance genes. In addition, the effect of high-pressure stress (400 and 500 MPa) on the expression of selected genes encoding tetracycline resistance was determined, as well as changes in the frequency of transfer of these genes in isolates showing sensitivity to tetracyclines. In our study, we observed an increase in the expression of genes encoding tetracyclines, especially the tet(L) gene, mainly under 400 MPa pressure. The study confirmed the possibility of transferring genes encoding tetracyclines such as tet(M), tet(L), tet(K), tet(W) and tet(O) by horizontal gene transfer in both control strains and exposed to high-pressure. Exposure of the strains to 400 MPa pressure had a greater effect on the possibility of gene transfer and expression than the application of a higher-pressure. To our knowledge, this study for the first time determined the effect of high-pressure stress on the expression of selected genes encoding tetracycline resistance, as well as the possibility and changes in the frequency of transfer of these genes in Enterococcus faecalis isolates showing sensitivity to tetracyclines and possessing silent genes. Due to the observed possibility of increased expression of some of the genes encoding tetracycline resistance and the possibility of their spread by horizontal gene transfer to other microorganisms in the food environment, under the influence of high-pressure processing in strains phenotypically susceptible to this antibiotic, it becomes necessary to monitor this ability in isolates derived from foods.

What role do biocontrol agents with Mg2+ play in the fate of antibiotic resistome and pathogenic bacteria in the phyllosphere?

The contamination of the plant phyllosphere with antibiotics and antibiotic resistance genes (ARGs), caused by application of antibiotics, is a significant environmental issue in agricultural management. Alternatively, biocontrol agents are environmentally friendly and have attracted a lot of interest. However, the influence of biocontrol agents on the phyllosphere resistome remains unknown. In this study, we applied biocontrol agents to control the wildfire disease in the Solanaceae crops and investigated their effects on the resistome and the pathogen in the phyllosphere by using metagenomics. A total of 250 ARGs were detected from 15 samples, which showed a variation in distribution across treatments of biocontrol agents (BA), BA with Mg2+ (T1), BA with Mn2+ (T2), and kasugamycin (T3) and nontreated (CK). The results showed that the abundance of ARGs under the treatment of BA-Mg2+ was lower than that in the CK group. The abundance of cphA3 (carbapenem resistance), PME-1 (carbapenem resistance), tcr3 (tetracycline antibiotic resistance), and AAC (3)-VIIIa (aminoglycoside antibiotic resistance) in BA-Mg2+ was significantly higher than that in BA-Mn2+ (P < 0.05). The abundance of cphA3, PME_1, and tcr3 was significantly negatively related to the abundance of the phyllosphere pathogen Pseudomonas syringae (P < 0.05). We also found that the upstream and downstream regions of cphA3 were relatively conserved, in which rpl, rpm, and rps gene families were identified in most sequences (92%). The Ka/Ks of cphA3 was 0 in all observed sequences, indicating that under the action of purifying selection, nonsynonymous substitutions are often gradually eliminated in the population. Overall, this study clarifies the effect of biocontrol agents with Mg2+ on the distribution of the phyllosphere resistome and provides evolutionary insights into the biocontrol process. IMPORTANCE: Our study applied metagenomics analysis to examine the impact of biocontrol agents (BAs) on the phyllosphere resistome and the pathogen. Irregular use of antibiotics has led to the escalating dissemination of antibiotic resistance genes (ARGs) in the environment. The majority of BA research has focused on the effect of monospecies on the plant disease control process, the role of the compound BA with nutrition elements in the phyllosphere disease, and the resistome is still unknown. We believe BAs are eco-friendly alternatives for antibiotics to combat the transfer of ARGs. Our results revealed that BA-Mg2+ had a lower relative abundance of ARGs compared to the CK group, and the phyllosphere pathogen Pseudomonas syringae was negatively related to three specific ARGs, cphA3, PME-1, and tcr3. These three genes also present different Ka/Ks. We believe that the identification of the distribution and evolution modes of ARGs further elucidates the ecological role and facilitates the development of BAs, which will attract general interest in this field.

Tetracycline hydrochloride loaded-alginate based nanoparticle-hydrogel beads for potential wound healing applications: In vitro drug delivery, release kinetics, and antibacterial activity.

Novel hydrogel beads based on nanocomposite with outstanding antibacterial and swelling capabilities have been successfully produced as an efficient drug carrier for potential drug delivery systems in wound healing applications. The beads were characterized by Fourier transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM), and EDX-Mapping analysis. Then, using tetracycline hydrochloride (TCH) as a model drug system, they were studied in vitro for their potential efficiency as pH and temperature dependent sustained drug delivery carriers. Moreover, they were assessed in terms of porosity, swelling degree, encapsulation efficiency, and in vitro release kinetics. Beads released drugs at their highest levels under alkaline circumstances (pH = 8) and at a temperature of 39 °C, with a cumulative TCH release of 96.2 % at 36 h and in accordance with the Weibull kinetics model (R2 = 0.98). Additionally, the disc diffusion experiment demonstrated the strong antibacterial activity of the synthesized beads and offered a feasible and cost-effective wound dressing material for treating infected wounds.

Sequence-structure-function characterization of the emerging tetracycline destructase family of antibiotic resistance enzymes.

Tetracycline destructases (TDases) are flavin monooxygenases which can confer resistance to all generations of tetracycline antibiotics. The recent increase in the number and diversity of reported TDase sequences enables a deep investigation of the TDase sequence-structure-function landscape. Here, we evaluate the sequence determinants of TDase function through two complementary approaches: (1) constructing profile hidden Markov models to predict new TDases, and (2) using multiple sequence alignments to identify conserved positions important to protein function. Using the HMM-based approach we screened 50 high-scoring candidate sequences in Escherichia coli, leading to the discovery of 13 new TDases. The X-ray crystal structures of two new enzymes from Legionella species were determined, and the ability of anhydrotetracycline to inhibit their tetracycline-inactivating activity was confirmed. Using the MSA-based approach we identified 31 amino acid positions 100% conserved across all known TDase sequences. The roles of these positions were analyzed by alanine-scanning mutagenesis in two TDases, to study the impact on cell and in vitro activity, structure, and stability. These results expand the diversity of TDase sequences and provide valuable insights into the roles of important residues in TDases, and flavin monooxygenases more broadly.

Interaction of tetracycline and copper co-intake in inducing antibiotic resistance genes and potential pathogens in mouse gut.

The widespread use of copper and tetracycline as growth promoters in the breeding industry poses a potential threat to environmental health. Nevertheless, to the best of our knowledge, the potential adverse effects of copper and tetracycline on the gut microbiota remain unknown. Herein, mice were fed different concentrations of copper and/or tetracycline for 6 weeks to simulate real life-like exposure in the breeding industry. Following the exposure, antibiotic resistance genes (ARGs), potential pathogens, and other pathogenic factors were analyzed in mouse feces. The co-exposure of copper with tetracycline significantly increased the abundance of ARGs and enriched more potential pathogens in the gut of the co-treated mice. Copper and/or tetracycline exposure increased the abundance of bacteria carrying either ARGs, metal resistance genes, or virulence factors, contributing to the widespread dissemination of potentially harmful genes posing a severe risk to public health. Our study provides insights into the effects of copper and tetracycline exposure on the gut resistome and potential pathogens, and our findings can help reduce the risks associated with antibiotic resistance under the One Health framework.

Using UV/peracetic acid as pretreatment for subsequent bio-treatment of antibiotic-containing wastewater treatment: Mitigating microbial inhibition and antibiotic resistance genes proliferation.

UV/peracetic acid (PAA) treatment presents a promising approach for antibiotic removal, but its effects on microbial community and proliferation of antibiotic resistance genes (ARGs) during the subsequent bio-treatment remain unclear. Thus, we evaluated the effects of the UV/PAA on tetracycline (TTC) degradation, followed by introduction of the treated wastewater into the bio-treatment system to monitor changes in ARG expression and biodegradability. Results demonstrated effective TTC elimination by the UV/PAA system, with carbon-centered radicals playing a significant role. Crucially, the UV/PAA system not only eliminated antibacterial activity but also inhibited potential ARG host growth, thereby minimizing the emergence and dissemination of ARGs during subsequent bio-treatment. Additionally, the UV/PAA system efficiently removed multi-antibiotic resistant bacteria and ARGs from the bio-treatment effluent, preventing ARGs from being released into the environment. Hence, we propose a multi-barrier strategy for treating antibiotic-containing wastewater, integrating UV/PAA pre-treatment and post-disinfection with bio-treatment. The inhibition of ARGs transmission by the integrated system was verified through actual soil testing, confirming its effectiveness in preventing ARGs dissemination in the surrounding natural ecosystem. Overall, the UV/PAA treatment system offers a promising solution for tackling ARGs challenges by controlling ARGs proliferation at the source and minimizing their release at the end of the treatment process.

Discovery of novel dihydronaphthalene-imidazole ligands as potential inhibitors of Staphylococcus aureus multidrug resistant NorA efflux pump: A combination of experimental and in silico molecular docking studies.

Overexpression of the efflux pump is a predominant mechanism by which bacteria show antimicrobial resistance (AMR) and leads to the global emergence of multidrug resistance (MDR). In this work, the inhibitory potential of library of dihydronapthyl scaffold-based imidazole derivatives having structural resemblances with some known efflux pump inhibitors (EPI) were designed, synthesized and evaluated against efflux pump inhibitor against overexpressing bacterial strains to study the synergistic effect of compounds and antibiotics. Out of 15 compounds, four compounds (Dz-1, Dz-3, Dz-7, and Dz-8) were found to be highly active. DZ-3 modulated the MIC of ciprofloxacin, erythromycin, and tetracycline by 128-fold each against 1199B, XU212 and RN4220 strains of S. aureus respectively. DZ-3 also potentiated tetracycline by 64-fold in E. coli AG100 strain. DZ-7 modulated the MIC of both tetracycline and erythromycin 128-fold each in S. aureus XU212 and S. aureus RN4220 strains. DZ-1 and DZ-8 showed the moderate reduction in MIC of tetracycline in E. coli AG100 only by 16-fold and 8-fold, respectively. DZ-3 was found to be the potential inhibitor of NorA as determined by ethidium bromide efflux inhibition and accumulation studies employing NorA overexpressing strain SA-1199B. DZ-3 displayed EPI activity at non-cytotoxic concentration to human cells and did not possess any antibacterial activity. Furthermore, molecular docking studies of DZ-3 was carried out in order to understand the possible binding sites of DZ-3 with the active site of the protein. These studies indicate that dihydronaphthalene scaffolds could serve as valuable cores for the development of promising EPIs.

Investigating the cause of increased tetracycline-resistant Neisseria gonorrhoeae in England, 2016-20.

BACKGROUND: Antimicrobial resistance in Neisseria gonorrhoeae is a global public health concern. Tetracycline resistance (TetR) increased from 39.4% to 75.2% between 2016 and 2021 in N. gonorrhoeae isolates collected through national surveillance in England, despite the absence of use of tetracyclines for the treatment of gonorrhoea. OBJECTIVES: We investigated whether there was correlation between bacterial sexually transmitted infection (STI) tests performed and treatment with antimicrobials, with increased TetR in N. gonorrhoeae. METHODS: We examined correlations between bacterial STI tests, antimicrobial treatment and TetR in N. gonorrhoeae, using national surveillance data from three large sexual health services (SHS) in London during 2016-20. Doxycycline prescribing data and antibiograms of a non-STI pathogen from distinct patient groups (sexual health, obstetric and paediatric), at a large London hospital, were analysed to identify if doxycycline use in SHS was associated with resistance in a non-STI organism. RESULTS: A substantial increase in TetR was observed, particularly in isolates from gay, bisexual and other MSM (GBMSM). Strong positive correlations were observed exclusively in GBMSM between N. gonorrhoeae TetR and both bacterial STI tests (r = 0.97, P = 0.01) and antimicrobial treatment (r = 0.87, P = 0.05). Doxycycline prescribing increased dramatically during the study period in SHS. Prevalence of TetR in Staphylococcus aureus was higher in isolates sourced from SHS attendees than those from other settings. CONCLUSIONS: Frequent screening of GBMSM at higher risk of STIs, such as those on pre-exposure prophylaxis (PrEP) leading to/and increased use of doxycycline for the treatment of diagnosed infections, may account for the increase in TetR in N. gonorrhoeae.