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872300.9903Unraveling the Basis of Neonicotinoid Resistance in Whitefly Species Complex: Role of Endosymbiotic Bacteria and Insecticide Resistance Genes. Bemisia tabaci (whitefly) is one of the most detrimental agricultural insect pests and vectors of many plant viruses distributed worldwide. Knowledge of the distribution patterns and insecticide resistance of this cryptic species is crucial for its management. In this study, genetic variation of mitochondrial cytochrome oxidase subunit 1 (MtCoI) gene of B. tabaci was analyzed followed by a study of the infection profile of various endosymbionts in 26 whitefly populations collected from West Bengal, India. Phylogenetic analysis revealed Asia I as the major cryptic species (65.38%), followed by Asia II 5, China 3, and Asia II 7, which were diversified into 20 different haplotypes. In addition to the primary endosymbiont (C. poriera), each of the four whitefly species showed a variable population of three secondary endosymbionts, majorly Arsenophonus with the highest infection rate (73.07%), followed by Wolbachia and Rickettsia. Further phylogenetic analyses revealed the presence of two subgroups of Arsenophonus, viz., A1 and A2, and one each in Wolbachia (W1) and Rickettsia (R3). Resistance to thiamethoxam, imidacloprid, and acetamiprid insecticides was analyzed for a clear picture of pesticide resistance status. The highest susceptibility was noted toward thiamethoxam (LC(50) = 5.36 mg/L), followed by imidacloprid and acetamiprid. The whitefly population from Purulia and Hooghly districts bearing Asia II 7 and Asia II 5 cryptic species, respectively, shows maximum resistance. The differences in mean relative titer of four symbiotic bacteria among field populations varied considerably; however, a significant positive linear correlation was observed between the resistance level and relative titer of Arsenophonus and Wolbachia in the case of imidacloprid and thiamethoxam, while only Wolbachia was found in case of acetamiprid. Expression analysis demonstrated differential upregulation of insecticide resistance genes with Purulia and Hooghly populations showing maximally upregulated P450 genes. Moreover, thiamethoxam and imidacloprid resistance ratio (RR) showed a significant correlation with CYP6CM1, CYP6DZ7, and CYP4C64 genes, while acetamiprid RR correlated with CYP6CX1, CYP6DW2, CYP6DZ7, and CYP4C64 genes. Taken together, these findings suggested that P450 mono-oxygenase and symbiotic bacteria together affected whitefly resistance to neonicotinoids. Hence, a symbiont-oriented management programme could be a better alternative to control or delay resistance development in whitefly and can be used for pesticide clean-up in an agricultural field.202235814684
872410.9898Effects of different salinity on the transcriptome and antibiotic resistance of two Vibrio parahaemolyticus strains isolated from Penaeus vannamei cultured in seawater and freshwater ponds. The transcriptome and antibiotic resistance of Vibrio parahaemolyticus isolated from Penaeus vannamei cultured in seawater (strain HN1)and freshwater (strain SH1) ponds were studied at different salinity (2‰ and 20‰). At different salinity, 623 differentially expressed genes (DEGs) significantly upregulated and 1,559 DEGs significantly downregulated in SH1. In HN1, 466 DEGs significantly upregulated and 1,930 DEGs significantly downregulated, indicating high salinity can lead to the downregulation of most genes. In KEGG analysis, the expression of DEGs annotated to starch and sucrose metabolism pathway was higher at 2‰ salinity than at 20‰ salinity in HN1 and SH1, implying salinity affected bacterial growth mainly through this pathway. In the enrichment analysis of upregulated DEGs, two pathways (Valine, leucine, and isoleucine degradation, and Butanoate metabolism) were significantly enriched at different salinity. Antibiotic-susceptibility test discovered that SH1 isolated from P. vannamei cultured in freshwater was resistant to multiple drugs, including kanamycin, gentamicin, medemycin, and azithromycin, at a salinity of 2‰, whereas at 20‰ salinity, SH1 was not resistant to the drugs. The HN1 strain isolated from P. vannamei cultured in mariculture was resistant to polymyxin B and clindamycin at 20‰ salinity. Whereas, HN1 was intermediately susceptible to these two antibiotics at 2‰ salinity. These results indicate that the drug resistance of bacteria was affected by salinity. Furthermore, beta-lactam resistance was significantly enriched in SH1 at different salinity, and the inhibition zone of penicillin G was consistent with the results of a beta-lactam resistance pathway.202134496040
872220.9896Symbiotic Fungus Affected the Asian Citrus Psyllid (ACP) Resistance to Imidacloprid and Thiamethoxam. The Asian citrus psyllid (ACP), Diaphorina citri (Kuwayama) (Hemiptera: Liviidae), is a notorious Rutaceae plant pest. Frequent and extensive use of pesticides has resulted in severe insecticide resistance in ACP populations. Fully understanding the mechanism of ACP resistance to pesticides is vital for us to control or delay the development of resistance. Therefore, we compared the difference in resistance to imidacloprid and thiamethoxam between Hunan (Yongzhou, Chenzhou) and Guangdong (Guangzhou) ACP populations and analyzed the correlations between the resistance level and genes and symbiotic fungi. The results showed that the resistance of the Guangdong ACP population to imidacloprid and thiamethoxam was lower than that of Hunan ACP population, and the relative expression of genes associated with P450 mono-oxygenase and acetylcholinesterase was significantly lower in the Guangdong ACP population than in Hunan ACP population. The differences of mean relative abundances of four symbiotic bacteria among three populations were marginally significant; however, the mean relative abundance of 16 fungi among three populations was significantly different, and positive linear correlations were observed between the resistance level and two fungi (Aspergillus niger and Aureobasidium pullulans) and two genes (CYP4C70 and CYP4DB1). Negative correlations were only observed between the resistance level and two fungi (Golubevia pallescens and Acremonium sclerotigenum). Moreover, four fungi were unique to the Chenzhou population which was the highest resistance to imidacloprid and thiamethoxam. These findings suggested the P450 mono-oxygenase and symbiotic fungi together affected ACP resistance to imidacloprid and thiamethoxam. In the future, we may use environmental G. pallescens and A. sclerotigenum to control or delay ACP resistance.202033391190
636830.9892Antibacterial effects of curcumin encapsulated in nanoparticles on clinical isolates of Pseudomonas aeruginosa through downregulation of efflux pumps. Curcumin as a flavonoid from the rhizome of Curcuma longa has antibacterial, antiviral and antifungal activity. Multidrug resistance in pathogenic bacteria is continuously increasing in hospitals. The aim of this study was to investigate the effect of curcumin encapsulated in micellar/polymersome nanoparticles as an efflux pump inhibitor (EPI) on the expression of mexX and oprM genes in curcumin-treated and -untreated isolates of Pseudomonas aeruginosa. Clinical isolates of Pseudomonas aeruginosa were treated with ciprofloxacin (sub-MICs) alone and/or in combination with curcumin-encapsulated in micellar/polymersome nanoparticles. The expression of mexX and oprM genes was quantitatively evaluated by qRT-PCR in curcumin-treated and -untreated bacteria after 24 h. Curcumin-encapsulated in nanoparticles (400 µg/mL) induced cell death up to 50% in ciprofloxacin-treated (1/2MIC) resistant isolates during 24 h, while the bacteria treated with ciprofloxacin (without curcumin) were not inhibited. Also, curcumin in different concentrations increased effect of ciprofloxacin (sub-MICs). Downregulation of mexX and oprM genes was observed in cells treated with curcumin and ciprofloxacin compared to cells treated with ciprofloxacin alone. It seems that curcumin can be used as complementary drug in ciprofloxacin-resistant isolates through downregulating genes involved in efflux pumps and trapping ciprofloxacin on bacterial cells and increasing the effects of drug.201930778922
575340.9890Sensitization of Gram-Negative Bacteria to Aminoglycosides with 2-Aminoimidazole Adjuvants. In 2019, five million deaths associated with antimicrobial resistance were reported by The Centers for Disease Control and Prevention (CDC). Acinetobacter baumannii, a Gram-negative bacterial pathogen, is among the list of urgent threats. Previously, we reported 2-aminoimidazole (2-AI) adjuvants that potentiate macrolide activity against A. baumannii. In this study, we identify several of these adjuvants that sensitize A. baumannii to aminoglycoside antibiotics. Lead compounds 1 and 7 lower the tobramycin (TOB) minimum inhibitory concentration (MIC) against the TOB-resistant strain AB5075 from 128 μg/mL to 2 μg/mL at 30 μM. In addition, the lead compounds lower the TOB MIC against the TOB-susceptible strain AB19606 from 4 μg/mL to 1 μg/mL and 0.5 μg/mL, respectively, at 30 μM and 15 μM. The evolution of resistance to TOB and 1 in AB5075 revealed mutations in genes related to protein synthesis, the survival of bacteria under environmental stressors, bacteriophages, and proteins containing Ig-like domains.202337998765
628950.9888Pseudomonas aeruginosa is oxygen-deprived during infection in cystic fibrosis lungs, reducing the effectiveness of antibiotics. Pseudomonas aeruginosa infects the lungs of patients with cystic fibrosis. Sputum expectorated from the lungs of patients contains low levels of oxygen, indicating that P. aeruginosa may be oxygen-deprived during infection. During in vitro growth under oxygen-limiting conditions, a P. aeruginosa reference strain increases expression of a cytochrome oxidase with a high affinity for oxygen, and of nitrate and nitrite reductases that enable it to use nitrate instead of oxygen during respiration. Here, we quantified transcription of the genes encoding these three enzymes in sputum samples from 18 infected patients, and in bacteria isolated from the sputum samples and grown in aerobic and anaerobic culture. In culture, expression of all three genes was increased by averages of 20- to 500-fold in anaerobically grown bacteria compared with those grown aerobically, although expression levels varied greatly between isolates. Expression of the same genes in sputum was similar to that of the corresponding bacteria in anaerobic culture. The isolated bacteria were less susceptible to tobramycin and ciprofloxacin, two widely used anti-pseudomonal antibiotics, when grown anaerobically than when grown aerobically. Our findings show that P. aeruginosa experiences oxygen starvation during infection in cystic fibrosis, reducing the effectiveness of antibiotic treatment.202337516450
66260.9887Gene expression and physiological role of Pseudomonas aeruginosa methionine sulfoxide reductases during oxidative stress. Pseudomonas aeruginosa PAO1 has two differentially expressed methionine sulfoxide reductase genes: msrA (PA5018) and msrB (PA2827). The msrA gene is expressed constitutively at a high level throughout all growth phases, whereas msrB expression is highly induced by oxidative stress, such as sodium hypochlorite (NaOCl) treatment. Inactivation of either msrA or msrB or both genes (msrA msrB mutant) rendered the mutants less resistant than the parental PAO1 strain to oxidants such as NaOCl and H2O2. Unexpectedly, msr mutants have disparate resistance patterns when exposed to paraquat, a superoxide generator. The msrA mutant had a higher paraquat resistance level than the msrB mutant, which had a lower paraquat resistance level than the PAO1 strain. The expression levels of msrA showed an inverse correlation with the paraquat resistance level, and this atypical paraquat resistance pattern was not observed with msrB. Virulence testing using a Drosophila melanogaster model revealed that the msrA, msrB, and, to a greater extent, msrA msrB double mutants had an attenuated virulence phenotype. The data indicate that msrA and msrB are essential genes for oxidative stress protection and bacterial virulence. The pattern of expression and mutant phenotypes of P. aeruginosa msrA and msrB differ from previously characterized msr genes from other bacteria. Thus, as highly conserved genes, the msrA and msrB have diverse expression patterns and physiological roles that depend on the environmental niche where the bacteria thrive.201323687271
575070.9887Effects of in vitro-induced drug resistance on the virulence of Streptococcus. This study aimed to evaluate the effects of in vitro-induced drug resistance on the virulence of Streptococcus. Micro-dilution method was used to determine the minimal inhibitory concentration (MIC). In vitro-induced drug resistance was conducted for S. agalactiae (CVCC1886) and S. dysgalactiae (CVCC3701) by gradually increasing the antimicrobial concentration (strains were from IVDC, China). PCR was used to detect the resistance and virulence genes of the strains before and after resistance induction. Colony morphology was observed to compare the physiological and biochemical properties of the strains. A total of 88 clean-grade Kunming mice (obtained from Inner Mongolia University, Hohhot, China) were used in half of the lethal dose (LD50) test for detecting the changes in virulence of strains. The results showed that S. agalactiae (CVCC1886) and S. dysgalactiae (CVCC3701) developed resistance against seven kinds of antibiotics, respectively. Resistance and virulence genes of CVCC3701 were changed when treated by the Penicillin-inducing. The growth of the CVCC3701-PEN was decreased compared to the CVCC3701. Virulence test in mice indicated that the LD50 of CVCC3701 before induction and CVCC3701-PEN after induction were 5.45 × 10(6) and 5.82 × 10(8)  CFU/ml, respectively. Compared with the untreated bacteria, the bacterial virulence was reduced 1.1 × 10(2) times after resistance induction. In conclusion, S. dysgalactiae (CVCC3701) is a susceptible strain of drug resistance to antibiotics, in vitro-induced drug resistance reduced the virulence of CVCC3701, but the virulence is still existing and also could result in the death of mice. For public health safety, it must be alert to the emergence of drug resistance of Streptococcus in animal production.202133314727
880480.9887A single exposure to a sublethal pediocin concentration initiates a resistance-associated temporal cell envelope and general stress response in Listeria monocytogenes. Listeria monocytogenes can cause the potentially fatal food-borne disease listeriosis, and the use of bacteriocin-producing lactic acid bacteria to control L. monocytogenes holds great promise. However, the development of bacteriocin resistance is a potential challenge, and the purpose of this study was to determine if exposure to sublethal concentrations of pediocin-containing Lactobacillus plantarum WHE 92 supernatant could prime L. monocytogenes for resistance. By transcriptomic analysis, we found two, 55 and 539 genes differentially expressed after 10, 60 and 180 min of exposure to L. plantarum WHE 92 supernatant as compared with control exposures. We observed temporal expression changes in genes regulated by the two component system LisRK and the alternative sigma factors SigB and SigL. Additionally, several genes involved in bacteriocin resistance were induced. ΔlisR, ΔsigB and ΔsigL mutants were all more resistant than wild types to L. plantarum WHE 92 supernatant. Conclusively, LisRK, SigB and SigL regulation and genes associated with resistance are involved in the temporal adaptive response to pediocin, and all three regulatory systems affect pediocin resistance. Thus, a single exposure to a sublethal pediocin concentration initiates a response pointing to resistance, and indicates that further research exploring the link between adaptive responses and resistance is needed.201524920558
588490.9885Early genetic diagnosis of clarithromycin resistance in Helicobacter pylori. BACKGROUND: The drug resistance rate of clinical Helicobacter pylori (H. pylori) isolates has increased. However, the mechanism of drug resistance remains unclear. In this study, drug-resistant H. pylori strains were isolated from different areas and different populations of Chinese for genomic analysis. AIM: To investigate drug-resistant genes in H. pylori and find the genes for the early diagnosis of clarithromycin resistance. METHODS: Three drug-resistant H. pylori strains were isolated from patients with gastritis in Bama County, China. Minimal inhibitory concentrations of clarithromycin, metronidazole, and levofloxacin were determined and complete genome sequencing was performed with annotation. Hp1181 and hp1184 genes were found in these strains and then detected by reverse transcription polymerase chain reaction. The relationships between hp1181 or hp1184 and clarithromycin resistance were ascertained with gene mutant and drug-resistant strains. The homology of the strains with hp26695 was assessed through complete genome detection and identification. Differences in genome sequences, gene quantity, and gene characteristics were detected amongst the three strains. Prediction and analysis of the function of drug-resistant genes indicated that the RNA expression of hp1181 and hp1184 increased in the three strains, which was the same in the artificially induced clarithromycin-resistant bacteria. After gene knockout, the drug sensitivity of the strains was assessed. RESULTS: The strains showing a high degree of homology with hp26695, hp1181, and hp1184 genes were found in these strains; the expression of the genes hp1184 and hp1181 was associated with clarithromycin resistance. CONCLUSION: Hp1181 and hp1184 mutations may be the earliest and most persistent response to clarithromycin resistance, and they may be the potential target genes for the diagnosis, prevention, and treatment of clarithromycin resistance.202134239272
8985100.9884High Wolbachia density correlates with cost of infection for insecticide resistant Culex pipiens mosquitoes. In the mosquito Culex pipiens, insecticide resistance genes alter many life-history traits and incur a fitness cost. Resistance to organophosphate insecticides involves two loci, with each locus coding for a different mechanism of resistance (degradation vs. insensitivity to insecticides). The density of intracellular Wolbachia bacteria has been found to be higher in resistant mosquitoes, regardless of the mechanism involved. To discriminate between costs of resistance due to resistance genes from those associated with elevated Wolbachia densities, we compared strains of mosquito sharing the same genetic background but differing in their resistance alleles and Wolbachia infection status. Life-history traits measured included strength of insecticide resistance, larval mortality, adult female size, fecundity, predation avoidance, mating competition, and strength of cytoplasmic incompatibility (CI). We found that: (1) when Wolbachia are removed, insecticide resistance genes still affect some life-history traits; (2) Wolbachia are capable of modifying the cost of resistance; (3) the cost of Wolbachia infections increases with their density; (4) different interactions occurred depending on the resistance alleles involved; and (5) high densities of Wolbachia do not increase the strength of CI or maternal transmission efficiency relative to low Wolbachia densities. Insecticide resistance genes generated variation in the costs of Wolbachia infections and provided an interesting opportunity to study how these costs evolve, a process generally operating when Wolbachia colonizes a new host.200616610322
6288110.9884Regulation of ofloxacin resistance in Escherichia coli strains causing calf diarrhea by quorum-sensing acyl-homoserine lactone signaling molecules. Escherichia coli is a major pathogen responsible for calf diarrhea. However, it has developed resistance to many antimicrobial drugs for their inappropriate usage. The bacterial quorum sensing system transmits information between bacteria, it's important in regulating bacterial virulence, drug and acid resistance and so on. This system can found in Gram-negative bacteria and operates through acyl-homoserine lactone (AHL) signaling molecules. In this study, a type I quorum sensing AHL, N-Octanoyl-L-Homoserine lactone (C8), was added to E. coli growth medium to investigate its regulatory functions in drug resistance. After screening out the strains of E. coli that showed an obvious regulatory effect to the drug ofloxacin (OFX), transcriptomic sequencing was performed on the E. coli strains from the sub-inhibitory concentration group that concentration plus C8 group, and the control group. It shows that C8 significantly influenced resistance to OFX and the minimum inhibitory concentration of OFX in the tested strain was significantly increased. To Analyze transcriptome sequencing results identified 415 differentially expressed genes between the control and sub-inhibitory concentration groups, of which 201 were up-regulated and 214 were down. There were 125 differentially expressed genes between bacteria treated with a sub-inhibitory concentration of OFX and those treated with C8, of which 102 were up-regulated and 23 were down. Finally, It found that to adding the C8 significantly increased the resistance of tested bacteria to OFX. Data from transcriptome sequencing on differently expressed genes helps to explain how the type I quorum sensing system controls drug resistance in E. coli.202539974163
6292120.9884Genome-Wide Screening and Characterization of Genes Involved in Response to High Dose of Ciprofloxacin in Escherichia coli. The global emergence of antibiotic resistance, especially in Gram-negative bacteria, is an urgent threat to public health. Inevitably, considering its extensive use and misuse, resistance toward ciprofloxacin has increased in almost all clinically relevant bacteria. This study aimed to investigate the transcriptome changes at a high concentration of ciprofloxacin in Escherichia coli. In brief, 1,418 differentially expressed genes (DEGs) were identified, from which 773 genes were upregulated by ciprofloxacin, whereas 651 genes were downregulated. Enriched biological pathways reflected the upregulation of biological processes such as DNA damage and repair system, toxin/antitoxin systems, formaldehyde detoxification system. With kyoto encyclopedia of genes and genomes pathway analysis, higher expressed DEGs were associated with "LPS biosynthesis," "streptomycin biosynthesis," and "polyketide sugar unit biosynthesis." Lower expressed DEGs were associated with "biosynthesis of amino acids" and "flagellar assembly" pathways. After treatment of ciprofloxacin, lipopolysaccharide (LPS) release was increased by two times, and the gene expression level of LPS synthesis was elevated (p < 0.05) in both reference and clinical strains. Our results demonstrated that transient exposure to high-dose ciprofloxacin is a double-edged sword. Cautions should be taken when administering high-dose antibiotic treatment for infectious diseases.202235512736
2282130.9884Cross resistance of quinolone derivatives in gram-negative bacteria. A total of 127 Gram-negative bacteria resistant to nalidixic acid were isolated from as many patients affected by urinary tract infections and hospitalized in the first Clinic of Infectious Diseases, University of Naples. Enterobacteria were identified by Enterotube system (Roche) and API 20 system (Ayerst). Non-fermentative bacteria were identified by OXI/FERM system (Roche). The following bacteria were collected: Escherichia coli 50, Proteus spp. 35, Enterobacter agglomerans 12, Serratia sp. 5, Pseudomonas aeruginosa 25. The in vitro antibacterial activity of nalidixic acid and three other quinoline derivatives (pipemidic acid, oxolinic acid and ciprofloxacin) were studied by determining the MICs by a miniaturized dilution broth method. The MICs were compared to evaluate the eventual cross resistance to the drugs under examination within each bacterial species. The results showed that 23% of bacteria were resistant to nalidixic acid, pipemidic acid and oxolinic acid; 49.6% to nalidixic and pipemidic acid and 0.7% to nalidixic acid and oxolinic acid. On the other hand none of the bacteria were resistant to ciprofloxacin. The last showed very low MICs against all the bacteria under examination, including Pseudomonas and Serratia. The high antibacterial activity of ciprofloxacin even against bacteria highly resistant to the other quinolines could be due to a greater affinity of the target sites or to the better permeability of resistant strains to the newer drug or because it is unaffected until now by mutations of genes responsible for cross resistance.19853159488
6362140.9884The role of midgut symbiotic bacteria in resistance of Anopheles stephensi (Diptera: Culicidae) to organophosphate insecticides. In the current study, the effects of the presence of symbiotic bacteria on the activity of the enzymes involved in An. stephensi resistance to temephos are evaluated for the first time. Four different strains (I. susceptible strain, II. resistant strain, III. resistant strain + antibiotic, and IV. resistant strain + bacteria) were considered in order to determine the possible effects of the symbiotic bacteria on their hosts' resistance to temephos. The median values of all enzymes of susceptible strain were compared with those of other resistant strains. The results of this study indicated a direct relationship between the presence of bacteria in the symbiotic organs of An. stephensi and resistance to temephos. The profile of enzymatic activities in the resistant strain changed to a susceptible status after adding antibiotic. The resistance of An. stephensi to temephos could be completely broken artificially by removing their bacterial symbionts in a resistant population.201728745553
5747150.9884Synergistic effects of antimicrobial peptide DP7 combined with antibiotics against multidrug-resistant bacteria. Antibiotic-resistant bacteria present a great threat to public health. In this study, the synergistic effects of antimicrobial peptides (AMPs) and antibiotics on several multidrug-resistant bacterial strains were studied, and their synergistic effects on azithromycin (AZT)-resistance genes were analyzed to determine the relationships between antimicrobial resistance and these synergistic effects. A checkerboard method was used to evaluate the synergistic effects of AMPs (DP7 and CLS001) and several antibiotics (gentamicin, vancomycin [VAN], AZT, and amoxicillin) on clinical bacterial strains (Staphylococcus aureus, Pseudomonas aeruginosa, Acinetobacter baumannii, and Escherichia coli). The AZT-resistance genes (ermA, ermB, ermC, mefA, and msrA) were identified in the resistant strains using quantitative polymerase chain reaction. For all the clinical isolates tested that were resistant to different antibiotics, DP7 had high antimicrobial activity (≤32 mg/L). When DP7 was combined with VAN or AZT, the effect was most frequently synergistic. When we studied the resistance genes of the AZT-resistant isolates, the synergistic effect of DP7-AZT occurred most frequently in highly resistant strains or strains carrying more than two AZT-resistance genes. A transmission electron microscopic analysis of the S. aureus strain synergistically affected by DP7-AZT showed no noteworthy morphological changes, suggesting that a molecular-level mechanism plays an important role in the synergistic action of DP7-AZT. AMP DP7 plus the antibiotic AZT or VAN is more effective, especially against highly antibiotic-resistant strains.201728356719
6376160.9884Mechanisms of mepA Overexpression and Membrane Potential Reduction Leading to Ciprofloxacin Heteroresistance in a Staphylococcus aureus Isolate. Heteroresistance has seriously affected the evaluation of antibiotic efficacy against pathogenic bacteria, causing misjudgment of antibiotics' sensitivity in clinical therapy, leading to treatment failure, and posing a serious threat to current medical health. However, the mechanism of Staphylococcus aureus heteroresistance to ciprofloxacin remains unclear. In this study, heteroresistance to ciprofloxacin in S. aureus strain 529 was confirmed by antimicrobial susceptibility testing and population analysis profiling (PAP), with the resistance of subclonal 529_HR based on MIC being 8-fold that of the original bacteria. A 7-day serial MIC evaluation and growth curves demonstrate that their phenotype was stable, with 529_HR growing more slowly than 529, but reaching a plateau in a similar proportion. WGS analysis showed that there were 11 nonsynonymous mutations and one deletion gene between the two bacteria, but none of these SNPs were directly associated with ciprofloxacin resistance. Transcriptome data analysis showed that the expression of membrane potential related genes (qoxA, qoxB, qoxC, qoxD, mprF) was downregulated, and the expression of multidrug resistance efflux pump gene mepA was upregulated. The combination of ciprofloxacin and limonene restored the 529_HR MIC from 1 mg/L to 0.125 mg/L. Measurement of the membrane potential found that 529_HR had a lower potential, which may enable it to withstand the ciprofloxacin-induced decrease in membrane potential. In summary, we demonstrated that upregulation of mepA gene expression and a reduction in membrane potential are the main heteroresistance mechanisms of S. aureus to ciprofloxacin. Additionally, limonene may be a potentially effective agent to inhibit ciprofloxacin heteroresistance phenotypes.202540076991
6190170.9883Identifying Escherichia coli genes involved in intrinsic multidrug resistance. Multidrug resistance is a major cause of clinical failure in treating bacterial infections. Increasing evidence suggests that bacteria can resist multiple antibiotics through intrinsic mechanisms that rely on gene products such as efflux pumps that expel antibiotics and special membrane proteins that block the penetration of drug molecules. In this study, Escherichia coli was used as a model system to explore the genetic basis of intrinsic multidrug resistance. A random mutant library was constructed in E. coli EC100 using transposon mutagenesis. The library was screened by growth measurement to identify the mutants with enhanced or reduced resistance to chloramphenicol (Cm). Out of the 4,000 mutants screened, six mutants were found to be more sensitive to Cm and seven were more resistant compared to the wild-type EC100. Mutations in 12 out of the 13 mutants were identified by inverse polymerase chain reaction. Mutants of the genes rob, garP, bipA, insK, and yhhX were more sensitive to Cm compared to the wild-type EC100, while the mutation of rhaB, yejM, dsdX, nagA, yccE, atpF, or htrB led to higher resistance. Overexpression of rob was found to increase the resistance of E. coli biofilms to tobramycin (Tob) by 2.7-fold, while overexpression of nagA, rhaB, and yccE significantly enhanced the susceptibility of biofilms by 2.2-, 2.5-, and 2.1-fold respectively.200818807027
5379180.9883Membrane-Targeting Triphenylphosphonium Functionalized Ciprofloxacin for Methicillin-Resistant Staphylococcus aureus (MRSA). Multidrug-resistant (MDR) bacteria have become a severe problem for public health. Developing new antibiotics for MDR bacteria is difficult, from inception to the clinically approved stage. Here, we have used a new approach, modification of an antibiotic, ciprofloxacin (CFX), with triphenylphosphonium (TPP, PPh(3)) moiety via ester- (CFX-ester-PPh(3)) and amide-coupling (CFX-amide-PPh(3)) to target bacterial membranes. In this study, we have evaluated the antibacterial activities of CFX and its derivatives against 16 species of bacteria, including MDR bacteria, using minimum inhibitory concentration (MIC) assay, morphological monitoring, and expression of resistance-related genes. TPP-conjugated CFX, CFX-ester-PPh(3), and CFX-amide-PPh(3) showed significantly improved antibacterial activity against Gram-positive bacteria, Staphylococcus aureus, including MDR S. aureus (methicillin-resistant S. aureus (MRSA)) strains. The MRSA ST5 5016 strain showed high antibacterial activity, with MIC values of 11.12 µg/mL for CFX-ester-PPh(3) and 2.78 µg/mL for CFX-amide-PPh(3). The CFX derivatives inhibited biofilm formation in MRSA by more than 74.9% of CFX-amide-PPh(3). In the sub-MIC, CFX derivatives induced significant morphological changes in MRSA, including irregular deformation and membrane disruption, accompanied by a decrease in the level of resistance-related gene expression. With these promising results, this method is very likely to combat MDR bacteria through a simple TPP moiety modification of known antibiotics, which can be readily prepared at clinical sites.202033143023
3610190.9883Quantitative real-time PCR study of the expression and regulation of the tetracycline resistance gene in Riemerella anatipestifer. Riemerella anatipestifer (RA) is one of the most important pathogens of 1- to 8-wk-old ducklings that severely affects the development of the duck industry in China. Every year, antibiotic medicines including tetracycline and doxycycline are used in the duck industry. Few reports compare the expression of multidrug-resistant genes in RA before and after addition of chemical drugs. With this in mind, the direct effects of gradient concentration of tetracyclines on the expression of tetracycline resistance genes (TETr) in RA at the cDNA level were studied by using a quantitative real-time PCR method. The expression of TETr, tetA, tetC, and tetM was investigated in ATCC11845 and in 30 RA isolated from different samples. Using a range of doxycycline concentrations up to 50% of the minimum inhibitory concentration (MIC), the optimal induction concentration of 0.0625 μg/mL was selected. Under the optimal inducible expression, concentrations of TETr, tetC, and tetM cDNA were detected in all isolates, and the highest mRNA expression level of TETr genes was shown. Additionally, the expression levels of 3 TETr genes in RA14 (tetA and tetC) and RA17 (tetM and tetC) were compared. Both tetC and tetA found in isolate RA14 was found to express both tetC and tetA, and tetC cDNA was detected in isolate RA17 at all doxycycline concentrations tested, whereas tetM cDNA was not detected at any concentration. We can conclude that resistance pump is the main mechanism of tetracycline antibiotic resistance, and under the action of drug resistance pump tetC, the expression of tetM was not activated in RA17. These data suggest that the mRNA expression level of TETr genes was correlated with the MIC values, indicating that the degree of drug resistance is determined by the expression levels of TETr genes. Also, the induction of TETr is the major tetracycline resistance mechanism in RA, especially the resistance pump. However, lower concentrations of doxycycline induced higher TETr expression, and higher concentrations inhibited TETr expression. Maybe that is the reason for selection mutation to make tolerated bacteria survive.201323687151