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1.
Arch Razi Inst ; 77(2): 697-701, 2022 Apr.
Article in English | MEDLINE | ID: mdl-36284977

ABSTRACT

Plants have been long valuable sources of natural materials that have served to preserve human and animal health; as a result, pharmacological purposes have arisen from the use of plant compounds in most countries, according to a World Health Organization report. The present study aimed to assess the antimicrobial resistance of tannin extract against Escherichia coli (E. coli) isolates in sheep. A total of 100 samples from sheep were used to isolate E. coli and treated with tannin extract (90% purity) to investigate the in vitro effect, as compared to some antibiotics (Clindamycin, Cephalexin, Kanamycin, Tetracycline, and Vancomycin). The bacterial samples were cultured in a selective and differential medium, and Gram staining was used to examine them. The biochemical assays were performed to purify and expose these cultures; moreover, the API 20E system and RapidTM ONE kits were utilized to confirm the bacterial strain. Based on the findings, 50% of the samples showed a positive result for the presence of E. coli. The well diffusion technique was used to investigate the antibacterial activity to confirm the antibacterial action of tannin extract (from pomegranate peel) in different concentrations against E. coli. The highest zone of inhibition for the bacteria ranged from 12±0.5 to 30.3±0.2 at 50% concentrations, proving that tannins extract was significantly effective against E. coli. The presence of E. coli was detected in 50 % of the samples. The well-diffusion technique was used to evaluate the antimicrobial property of tannin extract through various concentrations with the highest zone of inhibition for the bacteria ranging from 12.5 to 30.30.2 at 50%, demonstrating that tannin extract was significantly effective on E. coli.


Subject(s)
Anti-Infective Agents , Tannins , Sheep , Humans , Animals , Tannins/pharmacology , Tannins/chemistry , Escherichia coli , Anti-Bacterial Agents/pharmacology , Vancomycin/pharmacology , Clindamycin/pharmacology , Plant Extracts/pharmacology , Plant Extracts/chemistry , Drug Resistance, Bacterial , Anti-Infective Agents/pharmacology , Cephalexin/pharmacology , Kanamycin/pharmacology , Tetracyclines/pharmacology
2.
Lab Med ; 53(3): e69-e73, 2022 May 05.
Article in English | MEDLINE | ID: mdl-34668538

ABSTRACT

OBJECTIVE: To indicate the ability to disaggregate platelet clumps by vortex mixing and kanamycin supplementation in EDTA-dependent pseudothrombocytopenia (EDTA-PTCP) specimens. MATERIALS AND METHODS: For patients with EDTA-PTCP, citrate-anticoagulated, primary EDTA-anticoagulated, vortex-mixed, and kanamycin-treated specimens were tested for complete blood count and platelet-related parameters. RESULTS: Forty-eight specimens were included. Nineteen (39.6%) of the vortex-mixed specimens and 42 (87.5%) of the kanamycin-treated specimens revealed platelet counts more than those of the primary EDTA specimens, with levels exceeding 100 × 109/L. The platelet count and platelet recovery of the kanamycin-treated specimens were higher than those of the vortex-mixed specimens. CONCLUSION: Kanamycin supplementation to EDTA-PTCP blood may be considered as an alternative approach when the recollection of specimens is impractical. Only platelet-related parameters from kanamycin treatment were suitable for guiding patient management. Further studies about the impact of these methods in patients with various conditions, such as in patients with advanced kidney disease, should be conducted.


Subject(s)
Blood Platelet Disorders , Thrombocytopenia , Dietary Supplements , Edetic Acid/pharmacology , Humans , Kanamycin/pharmacology , Platelet Aggregation , Platelet Count/methods
3.
ACS Appl Mater Interfaces ; 12(50): 55696-55709, 2020 Dec 16.
Article in English | MEDLINE | ID: mdl-33249831

ABSTRACT

Antibiotic-resistant bacteria are a severe threat to human health. The World Health Organization's Global Antimicrobial Surveillance System has revealed widespread occurrence of antibiotic resistance among half a million patients across 22 countries, with Staphylococcus aureus, Escherichia coli, and Klebsiella pneumoniae being the most common resistant species. Antimicrobial nanoparticles are emerging as a promising alternative to antibiotics in the fight against antimicrobial resistance. In this work, selenium nanoparticles coated with the antimicrobial polypeptide, ε-poly-l-lysine, (Se NP-ε-PL) were synthesized and their antibacterial activity and cytotoxicity were investigated. Se NP-ε-PL exhibited significantly greater antibacterial activity against all eight bacterial species tested, including Gram-positive, Gram-negative, and drug-resistant strains, than their individual components, Se NP and ε-PL. The nanoparticles showed no toxicity toward human dermal fibroblasts at the minimum inhibitory concentrations, demonstrating a therapeutic window. Furthermore, unlike the conventional antibiotic kanamycin, Se NP-ε-PL did not readily induce resistance in E. coli or S. aureus. Specifically, S. aureus began to develop resistance to kanamycin from ∼44 generations, whereas it took ∼132 generations for resistance to develop to Se NP-ε-PL. Startlingly, E. coli was not able to develop resistance to the nanoparticles over ∼300 generations. These results indicate that the multifunctional approach of combining Se NP with ε-PL to form Se NP-ε-PL is a highly efficacious new strategy with wide-spectrum antibacterial activity, low cytotoxicity, and significant delays in development of resistance.


Subject(s)
Anti-Infective Agents/pharmacology , Biocompatible Materials/chemistry , Drug Resistance, Bacterial/drug effects , Nanoparticles/chemistry , Peptides/chemistry , Selenium/chemistry , Anti-Infective Agents/chemistry , Biocompatible Materials/pharmacology , Cell Line , Cell Survival/drug effects , Escherichia coli/drug effects , Fibroblasts/cytology , Fibroblasts/drug effects , Fibroblasts/metabolism , Humans , Kanamycin/pharmacology , Klebsiella pneumoniae/drug effects , Microbial Sensitivity Tests , Reactive Oxygen Species/metabolism , Staphylococcus aureus/drug effects
4.
World J Microbiol Biotechnol ; 36(11): 167, 2020 Oct 06.
Article in English | MEDLINE | ID: mdl-33025172

ABSTRACT

Activities of plant polyphenols (PPs), resveratrol and quercetin, alone or in combination with four conventional antibiotics against Escherichia coli have been investigated. In medium without antibiotics, both polyphenols caused a dose-dependent growth inhibition. However, pretreatment with resveratrol (40 and 100 µg ml-1) and quercetin (40 µg ml-1) reduced the bacteriostatic effect of kanamycin, streptomycin, cefotaxime and partially of ciprofloxacin. With few exceptions, both PPs also reduced the bactericidal effect of tested antibiotics. Paradoxically, low doses of PPs enhanced the bactericidal effect of kanamycin and partially ciprofloxacin. Compared to quercetin, resveratrol showed a weaker effect on the induction of antioxidant genes and the resistance of E. coli to the oxidative stress generated by hydrogen peroxide treatment. Both polyphenols at high doses reduced membrane potential. Altogether, these findings suggest that the decrease in the bactericidal effect of antibiotics by high doses of polyphenols is mostly due to bacteriostatic action of the latter. In the case of quercetin, the contribution of its antioxidant activity for antibiotic protection may be significant. There is a growing interest in the use of plant-derived compounds to enhance the toxicity of traditional antibiotics. This and other studies show that, under certain conditions, the use of polyphenols as adjuvants may not exert the expected therapeutic effect, but rather to decrease antimicrobial activity of antibiotics.


Subject(s)
Antioxidants/pharmacology , Escherichia coli/drug effects , Quercetin/pharmacology , Resveratrol/pharmacology , Anti-Bacterial Agents/pharmacology , Cefotaxime/pharmacology , Ciprofloxacin/pharmacology , Drug Resistance, Multiple, Bacterial , Escherichia coli/metabolism , Kanamycin/pharmacology , Oxidative Stress/drug effects , Plant Extracts/pharmacology , Polyphenols/pharmacology , Streptomycin/pharmacology , Stress, Physiological/drug effects , beta-Galactosidase/metabolism
5.
J Med Microbiol ; 69(7): 1013-1019, 2020 Jul.
Article in English | MEDLINE | ID: mdl-32579102

ABSTRACT

Introduction. Multidrug-resistant tuberculosis (MDR-TB) is a major public health problem globally, including in Indonesia. Whole-genome sequencing (WGS) analysis has rarely been used for the study of TB and MDR-TB in Indonesia.Aim. We evaluated the use of WGS for drug-susceptibility testing (DST) and to investigate the population structure of drug-resistant Mycobacterium tuberculosis in Java, Indonesia.Methodology. Thirty suspected MDR-TB isolates were subjected to MGIT 960 system (MGIT)-based DST and to WGS. Phylogenetic analysis was done using the WGS data. Results obtained using MGIT-based DST and WGS-based DST were compared.Results. Agreement between WGS and MGIT was 93.33 % for rifampicin, 83.33 % for isoniazid and 76.67 % for streptomycin but only 63.33 % for ethambutol. Moderate WGS-MGIT agreement was found for second-line drugs including amikacin, kanamycin and fluoroquinolone (73.33-76.67 %). MDR-TB was more common in isolates of the East Asian Lineage (63.3%). No evidence of clonal transmission of DR-TB was found among members of the tested population.Conclusion. Our study demonstrated the applicability of WGS for DST and molecular epidemiology of DR-TB in Java, Indonesia. We found no transmission of DR-TB in Indonesia.


Subject(s)
Mycobacterium tuberculosis/drug effects , Mycobacterium tuberculosis/genetics , Tuberculosis, Multidrug-Resistant/genetics , Adult , Antitubercular Agents/pharmacology , Drug Evaluation, Preclinical/methods , Drug Resistance, Multiple, Bacterial/drug effects , Drug Resistance, Multiple, Bacterial/genetics , Female , Genotype , Humans , Indonesia/epidemiology , Kanamycin/pharmacology , Male , Microbial Sensitivity Tests , Middle Aged , Molecular Epidemiology , Mutation , Phenotype , Phylogeny , Rifampin/pharmacology , Streptomycin/pharmacology , Tuberculosis, Multidrug-Resistant/microbiology , Whole Genome Sequencing/methods
6.
Biotechnol Lett ; 41(8-9): 1023-1031, 2019 Sep.
Article in English | MEDLINE | ID: mdl-31203568

ABSTRACT

OBJECTIVE: The present study evaluated the antibacterial activity and mode of action of fibrauretine on Escherichia coli (E. coli) and Staphylococcus aureus, and synergistic effect with kanamycin against multi-drug resistant E. coli. RESULTS: The fibrauretine exhibited inhibitory effect on the growth of the tested bacteria with minimum inhibitory concentration (MIC) and minimum bactericidal concentration of 2.5-5 and 5-10 mg/ml, respectively. Morphological changes of cell microstructure were observed after adding fibrauretine at MIC. The mode of action was further confirmed by measuring release of 260-nm absorbing materials and extracellular potassium ions. Checkerboard dilution test suggested that fibrauretine exhibited synergistic activity when combined with kanamycin (FICI ranging from 0.5625 to 0.625). CONCLUSIONS: Our results indicated that fibrauretine exerted synergistic effect with kanamycin and its antibacterial mode of action mainly attributed to disruption of cell membrane integrity.


Subject(s)
Anti-Bacterial Agents/pharmacology , Drug Resistance, Multiple, Bacterial , Drug Synergism , Escherichia coli/drug effects , Kanamycin/pharmacology , Phytochemicals/pharmacology , Plant Extracts/pharmacology , Microbial Sensitivity Tests , Microbial Viability/drug effects , Phytochemicals/isolation & purification , Plant Extracts/isolation & purification , Staphylococcus aureus/drug effects
7.
Methods Mol Biol ; 1815: 475-492, 2018.
Article in English | MEDLINE | ID: mdl-29981143

ABSTRACT

Pentalinon andrieuxii is a species used in Mayan traditional medicine due to its biological properties. Recent studies indicate that it produces a pentacyclic triterpene-denominated betulinic acid, which presents various biological activities: antibacterial, antifungal, antiplasmodial, anti-inflammatory, antimalarial, anticancer, leishmanicidal, and antiviral, as well as steroids and sterols with leishmanicidal properties. A recent study also reported the presence of urechitol A and B in the roots; these are secondary metabolites whose biochemical function is as yet unknown. This plant therefore represents a natural source of metabolites with potential application in the pharmaceutical industry. In this chapter, a protocol is described for obtaining transgenic plants, at the reporter gene of the ß-glucuronidase (GUS) via Agrobacterium tumefaciens from hypocotyl and root explants. The protocol established herein could be employed for the manipulation of the genes involved in the biosynthesis of isoprenoids or secondary metabolites of interest. To our knowledge, this is the first report of stable transformation of Pentalinon andrieuxii via Agrobacterium tumefaciens.


Subject(s)
Apocynaceae/genetics , Tissue Culture Techniques/methods , Transformation, Genetic , Adaptation, Physiological , Agrobacterium tumefaciens/metabolism , Culture Media/chemistry , Genes, Reporter , Germination/drug effects , Glucuronidase/metabolism , Hypocotyl/growth & development , Kanamycin/pharmacology , Plant Shoots/physiology , Plasmids/metabolism , Polymerase Chain Reaction , Seeds/physiology
8.
Article in English | MEDLINE | ID: mdl-29133554

ABSTRACT

Rapid and accurate drug susceptibility testing (DST) is essential for the treatment of multi- and extensively drug-resistant tuberculosis (M/XDR-TB). We compared the utility of genotypic DST assays with phenotypic DST (pDST) using Bactec 960 MGIT or Löwenstein-Jensen to construct M/XDR-TB treatment regimens for a cohort of 25 consecutive M/XDR-TB patients and 15 possible anti-TB drugs. Genotypic DST results from Cepheid GeneXpert MTB/RIF (Xpert) and line probe assays (LPAs; Hain GenoType MTBDRplus 2.0 and MTBDRsl 2.0) and whole-genome sequencing (WGS) were translated into individual algorithm-derived treatment regimens for each patient. We further analyzed if discrepancies between the various methods were due to flaws in the genotypic or phenotypic test using MIC results. Compared with pDST, the average agreement in the number of drugs prescribed in genotypic regimens ranged from just 49% (95% confidence interval [CI], 39 to 59%) for Xpert and 63% (95% CI, 56 to 70%) for LPAs to 93% (95% CI, 88 to 98%) for WGS. Only the WGS regimens did not contain any drugs to which pDST showed resistance. Importantly, MIC testing revealed that pDST likely underestimated the true rate of resistance for key drugs (rifampin, levofloxacin, moxifloxacin, and kanamycin) because critical concentrations (CCs) were too high. WGS can be used to rule in resistance even in M/XDR strains with complex resistance patterns, but pDST for some drugs is still needed to confirm susceptibility and construct the final regimens. Some CCs for pDST need to be reexamined to avoid systematic false-susceptible results in low-level resistant isolates.


Subject(s)
Antitubercular Agents/pharmacology , Drug Resistance, Multiple, Bacterial/genetics , Extensively Drug-Resistant Tuberculosis/drug therapy , Genome, Bacterial , Mycobacterium tuberculosis/genetics , Tuberculosis, Multidrug-Resistant/drug therapy , Bacterial Typing Techniques , Cohort Studies , Extensively Drug-Resistant Tuberculosis/microbiology , Genotype , Humans , Kanamycin/pharmacology , Levofloxacin/pharmacology , Microbial Sensitivity Tests , Moxifloxacin/pharmacology , Mycobacterium tuberculosis/drug effects , Mycobacterium tuberculosis/growth & development , Phenotype , Rifampin/pharmacology , Tuberculosis, Multidrug-Resistant/microbiology , Whole Genome Sequencing
9.
Biosci Biotechnol Biochem ; 81(8): 1461-1465, 2017 Aug.
Article in English | MEDLINE | ID: mdl-28475419

ABSTRACT

Various antimicrobial constituents of camu-camu fruit were isolated. Acylphloroglucinol (compound 1) and rhodomyrtone (compound 2) were isolated from the peel of camu-camu (Myrciaria dubia) fruit, while two other acylphloroglucinols (compounds 3 and 4) were obtained from camu-camu seeds. The structures of the isolated compounds were characterized by spectrophotometric methods. Compounds 1 and 4 were confirmed to be new acylphloroglucinols with different substituents at the C7 or C9 position of 2, and were named myrciarone A and B, respectively. Compound 3 was determined to be isomyrtucommulone B. This is the first report of the isolation of 3 from a natural resource. The antimicrobial activities of compounds 1, 3, and 4 were similar to those of 2, and the minimum inhibitory concentrations were either similar to or lower than that of kanamycin. These results suggest that the peel and seeds of camu-camu fruit could be utilized for therapeutic applications.


Subject(s)
Anti-Infective Agents/chemistry , Fruit/chemistry , Myrtaceae/chemistry , Phloroglucinol/isolation & purification , Seeds/chemistry , Xanthones/chemistry , Anti-Infective Agents/isolation & purification , Anti-Infective Agents/pharmacology , Candida albicans/drug effects , Candida albicans/growth & development , Gram-Negative Bacteria/drug effects , Gram-Negative Bacteria/growth & development , Gram-Positive Bacteria/drug effects , Gram-Positive Bacteria/growth & development , Kanamycin/pharmacology , Microbial Sensitivity Tests , Molecular Structure , Phloroglucinol/analogs & derivatives , Phloroglucinol/pharmacology , Plant Extracts/chemistry , Saccharomyces cerevisiae/drug effects , Saccharomyces cerevisiae/growth & development , Xanthones/isolation & purification , Xanthones/pharmacology
10.
BMC Complement Altern Med ; 17(1): 168, 2017 Mar 24.
Article in English | MEDLINE | ID: mdl-28340621

ABSTRACT

BACKGROUND: Recinodindron heudelotii (Euphorbiaceae) is a plant used in Africa, particularly in Cameroon to treat various ailments including bacterial infections. In this study, we evaluated the extracts of the leaves (RHL) and bark (RHB) of R. heudelotii for their antibacterial and antibiotic resistance modulating activities against 29 Gram-negative bacteria, including multidrug-resistant (MDR) phenotypes. METHODS: The broth micro-dilution assay was used to evaluate the antibacterial activity, and the antibiotic resistance modulating effects of the plant extracts. RESULTS: RHL displayed the most important spectrum of activity with minimal inhibitory concentrations (MICs) values ranging from 256 to 1024 µg/mL against 75.86% of the 29 tested bacteria strains while RHB was not active. RHL also showed killing effects with minimal bactericidal concentrations (MBCs) ranging from 256 to 1024 µg/mL. The activities of tetracycline and kanamycin associated with RHL were improved on 88.89% and 77.78% of the tested MDR bacteria, at MIC/2 at MIC/4 respectively, with 2 to 16-folds decreasing of MIC. This suggests the antibiotic resistance modulating effects of these antibiotics. CONCLUSION: The present study provides data indicating a possible use of the leaves extract of Recinodindron heudelotii alone or in association with common antibiotics in the fight against bacterial infections including those involving MDR bacteria.


Subject(s)
Anti-Bacterial Agents/pharmacology , Euphorbiaceae/chemistry , Gram-Negative Bacteria/drug effects , Plant Extracts/pharmacology , Drug Resistance, Multiple, Bacterial , Gram-Negative Bacteria/physiology , Humans , Kanamycin/pharmacology , Microbial Sensitivity Tests , Plant Bark/chemistry , Plant Leaves/chemistry , Tetracycline/pharmacology
11.
Nanotechnology ; 27(12): 125601, 2016 Mar 29.
Article in English | MEDLINE | ID: mdl-26883410

ABSTRACT

Recently, antibiotic drug-resistant therapies have become very important due to the emergence of antibiotic-resistant bacterial strains. The development of novel antibacterial materials has received significant attention. Here, quaternized chitosan hydrogels incorporated with NaYF4:Er/Yb/Mn@photosensitizer-doped silica (UCNPs/MB) were synthesized for effective killing of both gram-positive oxacillin-resistant S. aureus (DR-S. aureus) and gram-negative kanamyclin-resistant E. coli (DR-E. coli) bacteria upon near-infrared (NIR) laser irradiation. In this system, the cationic macroporous nature of the hydrogel acts as a molecular 'anion sponge', which sucks the outer part of the anionic microbe membrane into the gel interior voids and causes microbe membrane disruption. By incorporating UCNPs/MB-doped silica into the hydrogel, we have combined photodynamic therapy (PDT) with quaternized chitosan to obtain a high therapeutic index via a synergistic effect. In vitro experiments have demonstrated that our system had excellent antibacterial efficiency to both DR-S. aureus and DR-E. coli bacteria. More importantly, our new synergistic treatment modality provided an excellent therapy platform for drug-resistant bacteria, which could improve antimicrobial efficiency.


Subject(s)
Drug Resistance, Bacterial/drug effects , Hydrogels/chemical synthesis , Hydrogels/pharmacology , Nanoparticles/chemistry , Photosensitizing Agents/chemical synthesis , Photosensitizing Agents/pharmacology , Anti-Bacterial Agents/chemical synthesis , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/pharmacology , Chitosan/chemistry , Escherichia coli/drug effects , Hydrogels/chemistry , Kanamycin/pharmacology , Microbial Sensitivity Tests , Nanoparticles/ultrastructure , Oxacillin/pharmacology , Photosensitizing Agents/chemistry , Staphylococcus aureus/drug effects
12.
BMC Complement Altern Med ; 15: 412, 2015 Nov 22.
Article in English | MEDLINE | ID: mdl-26590804

ABSTRACT

BACKGROUND: The present study was designed to investigate the antibacterial activities of the methanol extracts from different parts of Beilschmedia acuta Kosterm (Lauraceae), Clausena anisata (Willd) Hook (Rutaceae), Newbouldia laevis Seem (Bignoniaceae) and Polyscias fulva (Hiern) Harms (Araliaceae) as well as their synergistic effects with antibiotics against a panel of Gram-negative bacteria, including multi-drug resistant (MDR) phenotypes expressing active efflux pumps. METHODS: Broth microdilution method was used to determine the minimum inhibitory concentrations (MICs) and the minimum bactericidal concentrations (MBCs) of the extracts, as well as those of antibiotics in association with the most active ones, B. acuta, N. laevis and P. fulva. RESULTS: MIC values obtained indicate that extracts from the bark of B. acuta were active on all the 26 tested Gram-negative bacteria, with MICs ranging from values below 8 to 256 µg/mL. Other samples displayed selective activities, their inhibitory effects being observed on 9 (34.62 %) of the 26 bacterial strains for N. laevis leaves extract, 6 (23.10 %) for both C. anisata leaves and roots extracts, 7 (26.9 %) and 4 (15.4 %) for leaves and roots extracts of P. fulva respectively. Extract from B. actua bark displayed the best antibacterial activity with MIC values below 100 µg/mL against 16 (61.5 %) of the 26 tested microorganisms. The lowest MIC values (below 8 µg/mL) were obtained with this extract against Escherichia coli W3110 and Klebsiella pneumoniae ATCC11296. The MIC values of this extract were lower than those of ciprofloxacin against E. coli W3110, Enterobacter aerogenes ATCC13048, CM64 and Providencia stuartii NAE16. At MIC/2, the best percentages of synergistic effects (100 %), were obtained with B. acuta bark extract and tetracycline (TET) as well as with P. fulva leaves extract and TET and kanamycin (KAN). CONCLUSION: The overall results of the present study provide information for the possible use of the studied plants and mostly Beilschmedia acuta in the control of bacterial infections including MDR phenotypes.


Subject(s)
Anti-Bacterial Agents/pharmacology , Araliaceae/chemistry , Bignoniaceae/chemistry , Clausena/chemistry , Gram-Negative Bacteria/drug effects , Lauraceae/chemistry , Plant Extracts/pharmacology , Rutaceae/chemistry , Anti-Bacterial Agents/chemistry , Ciprofloxacin/pharmacology , Drug Resistance, Multiple, Bacterial/drug effects , Drug Synergism , Kanamycin/pharmacology , Methanol , Microbial Sensitivity Tests , Plant Extracts/chemistry , Plant Leaves/chemistry , Tetracycline/pharmacology
14.
Zhong Yao Cai ; 36(12): 1904-7, 2013 Dec.
Article in Chinese | MEDLINE | ID: mdl-25090668

ABSTRACT

OBJECTIVE: Based on the system of regeneration,the genetic transformation system of Lonicera macranthoides was established. METHODS: Tissue culture method of seedlings, Agrobacterium tumefaciens mediated transformation method of explants, report gene was detected by gus staining and PCR. RESULTS: The efficient transformation time was 8 minutes of infection. The good transformation rate was gained with the kanamycin 35 mg/L and cefotaxime 600 mg/L. The concentration of kanamycin had a leading effect on bud differentiation between two antibiotics, and bud induction rate reached extremely significant difference. Results of gus staining and PCR proved that the gus gene was integrated into Lonicera macranthoides genome. CONCLUSION: The genetic transformation system of Lonicera macranthoides leaves mediated by Agrobacterium tumefaciens EHAlO5 was established for the first time.


Subject(s)
Agrobacterium tumefaciens/genetics , Lonicera/genetics , Plants, Medicinal/genetics , Transformation, Genetic , Cefotaxime/pharmacology , Genetic Vectors , Kanamycin/pharmacology , Lonicera/growth & development , Plant Leaves/genetics , Plant Leaves/growth & development , Plants, Genetically Modified/genetics , Plants, Genetically Modified/growth & development , Plants, Medicinal/growth & development , Polymerase Chain Reaction , Tissue Culture Techniques/methods
15.
Mol Syst Biol ; 8: 617, 2012.
Article in English | MEDLINE | ID: mdl-23047527

ABSTRACT

The inoculum effect (IE) refers to the decreasing efficacy of an antibiotic with increasing bacterial density. It represents a unique strategy of antibiotic tolerance and it can complicate design of effective antibiotic treatment of bacterial infections. To gain insight into this phenomenon, we have analyzed responses of a lab strain of Escherichia coli to antibiotics that target the ribosome. We show that the IE can be explained by bistable inhibition of bacterial growth. A critical requirement for this bistability is sufficiently fast degradation of ribosomes, which can result from antibiotic-induced heat-shock response. Furthermore, antibiotics that elicit the IE can lead to 'band-pass' response of bacterial growth to periodic antibiotic treatment: the treatment efficacy drastically diminishes at intermediate frequencies of treatment. Our proposed mechanism for the IE may be generally applicable to other bacterial species treated with antibiotics targeting the ribosomes.


Subject(s)
Anti-Bacterial Agents/pharmacology , Bacteria/drug effects , Bacteria/growth & development , Chloramphenicol/pharmacology , Colony Count, Microbial , Escherichia coli/drug effects , Escherichia coli/growth & development , Heat-Shock Response/drug effects , Kanamycin/pharmacology , Kinetics , Microbial Sensitivity Tests , Models, Biological , Proteolysis/drug effects , Ribosomes/drug effects , Ribosomes/metabolism , Salmonella typhimurium/drug effects , Salmonella typhimurium/growth & development
16.
Plant Cell Rep ; 31(10): 1899-909, 2012 Oct.
Article in English | MEDLINE | ID: mdl-22733208

ABSTRACT

Genetic transformation of the Indian medicinal plant, Bacopa monnieri, using a gene encoding cryptogein, a proteinaceous elicitor, via Ri and Ti plasmids, were established and induced bioproduction of bacopa saponins in crypt-transgenic plants were obtained. Transformed roots obtained with A. rhizogenes strain LBA 9402 crypt on selection medium containing kanamycin (100 mg l(-1)) dedifferentiated forming callus and redifferentiated to roots which, spontaneously showed shoot bud induction. Ri crypt-transformed plants thus obtained showed integration and expression of rol genes as well as crypt gene. Ti crypt-transformed B. monnieri plants were established following transformation with disarmed A. tumefaciens strain harboring crypt. Transgenic plants showed significant enhancement in growth and bacopa saponin content. Bacopasaponin D (1.4-1.69 %) was maximally enhanced in transgenic plants containing crypt. In comparison to Ri-transformed plants, Ri crypt-transformed plants showed significantly (p ≤ 0.05) enhanced accumulation of bacoside A(3), bacopasaponin D, bacopaside II, bacopaside III and bacopaside V. Produced transgenic lines can be used for further research on elicitation in crypt-transgenic plants as well as for large scale production of saponins. Key message The cryptogein gene, which encodes a proteinaceous elicitor is associated with increase in secondary metabolite accumulation-either alone or in addition to the increases associated with transformation by A. rhizogenes.


Subject(s)
Bacopa/genetics , Fungal Proteins/metabolism , Glycosides/biosynthesis , Saponins/biosynthesis , Agrobacterium/genetics , Agrobacterium/metabolism , Bacopa/drug effects , Bacopa/metabolism , Cell Dedifferentiation , Cell Differentiation , Culture Media/metabolism , Fungal Proteins/genetics , Gene Expression Regulation, Plant , Kanamycin/pharmacology , Phytophthora/chemistry , Plant Roots/drug effects , Plant Roots/genetics , Plant Roots/metabolism , Plants, Genetically Modified/genetics , Plants, Genetically Modified/metabolism , Plasmids/genetics , Plasmids/metabolism , Transformation, Genetic , Triterpenes
17.
Plant Cell Rep ; 31(8): 1517-26, 2012 Aug.
Article in English | MEDLINE | ID: mdl-22534683

ABSTRACT

UNLABELLED: A method has been developed to genetically transform the medicinal plant Maesa lanceolata. Initially, we tested conditions for transient expression of GFP-bearing constructs in agroinfiltrated leaves. Leaf tissues of M. lanceolata were infiltrated with Agrobacterium tumefaciens carrying a nuclear-targeted GFP construct to allow the quantification of the transformation efficiency. The number of transfected cells was depended on the bacterial density, bacterial strains, the co-cultivation time, and presence of acetosyringone. The transient transformation assay generated the highest ratio of transfected cells over non-transfected cells upon 5 days post-infiltration using A. tumefaciens strain LBA4404 at an OD600 = 1.0 in the presence of 100 µM acetosyringone and in the absence of a viral suppressor construct. In a second series of experiments we set up a stable transformation protocol that resulted in the regeneration of kanamycin-resistant plants expressing nuclear GFP. This transformation protocol will be used to introduce overexpression and RNAi constructs into M. lanceolata plants that may interfere with triterpenoid saponin biosynthesis. KEY MESSAGE: We have developed a transformation protocol for saponin producing Maesa lanceolata. Using the protocol reported here, now we are able to generate the tools for the modification of saponin production.


Subject(s)
Agrobacterium tumefaciens/physiology , Genetic Techniques , Plant Leaves/microbiology , Primulaceae/genetics , Primulaceae/microbiology , Saponins/biosynthesis , Transformation, Genetic , Acetophenones/metabolism , Agrobacterium tumefaciens/cytology , Agrobacterium tumefaciens/drug effects , Agrobacterium tumefaciens/growth & development , Cefotaxime/pharmacology , Cell Nucleus/drug effects , Cell Nucleus/metabolism , Colony Count, Microbial , Gene Expression/drug effects , Gene Silencing/drug effects , Genes, Suppressor , Green Fluorescent Proteins/metabolism , Kanamycin/pharmacology , Plant Leaves/cytology , Plant Leaves/drug effects , Plant Shoots/drug effects , Plant Shoots/physiology , Polymerase Chain Reaction , Primulaceae/drug effects , Regeneration/drug effects , Time Factors , Transfection , Transformation, Genetic/drug effects
18.
J Appl Microbiol ; 113(1): 192-9, 2012 Jul.
Article in English | MEDLINE | ID: mdl-22533708

ABSTRACT

AIMS: To investigate the influence of polyphenols and plant extracts on the susceptibility of Escherichia coli to antibiotics. METHODS AND RESULTS: Susceptibility of E. coli to antibiotics in the presence of extracts and polyphenols was estimated by the determination of the minimum inhibitory concentrations (MICs). To study gene expression, we used strains of E. coli carrying fusions between promoters of genes katG, sodA, iucC and structural ß-galactosidase gene. Treatment with polyphenols and some plant extracts significantly decreased the antibacterial effects of antibiotics, to a larger extent, ciprofloxacin. The most remarkable protective effect was observed for the extracts of Chamerion (Epilobium) angustifolium, Filipendula vulgaris, Tanacetum vulgare and Serratula coronata. These extracts increased the MICs of ciprofloxacin by four and more times. In case of kanamycin, extracts of Artemisia austriaca and Artemisia pontica increased MICs by four and eight times, respectively. Polyphenol quercetin also caused protective effect against ciprofloxacin, increasing the MIC by four times. A positive correlation was found between protective effects of polyphenols and extracts and their antioxidant activity. CONCLUSION: Medicinal plant extracts and polyphenols may protect cells of E. coli against antibiotic toxicity. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study may be used to enhance the efficiency of antibacterial therapies.


Subject(s)
Escherichia coli/drug effects , Herb-Drug Interactions , Plant Extracts/pharmacology , Plants, Medicinal/chemistry , Polyphenols/pharmacology , Anti-Bacterial Agents/pharmacology , Ciprofloxacin/pharmacology , Drug Resistance, Bacterial , Escherichia coli/enzymology , Escherichia coli/genetics , Kanamycin/pharmacology , Microbial Sensitivity Tests , Plant Extracts/chemistry , Polyphenols/chemistry , Quercetin/chemistry , Quercetin/pharmacology , beta-Galactosidase/metabolism
19.
Appl Biochem Biotechnol ; 166(8): 1871-95, 2012 Apr.
Article in English | MEDLINE | ID: mdl-22434351

ABSTRACT

An optimized protocol for Agrobacterium tumefaciens-mediated transformation of patchouli using leaf disk explants is reported. In vitro antibacterial activity of leaf extracts of the plants revealed Agrobacterium sensitivity to the extracts. Fluorometric assay of bacterial cell viability indicated dose-dependent cytotoxic activity of callus extract against Agrobacterium cells. Addition of 0.1% Tween 20 and 2 g/l L-glutamine to Agrobacterium infection medium counteracted the bactericidal effect and significantly increased the T-DNA delivery to explants. A short preculture of explants for 2 days followed by infection with Agrobacterium in medium containing 150 µM of acetosyringone were found essential for efficient T-DNA delivery. Cocultivation for 3 days at 22 °C in conjunction with other optimized factors resulted in maximum T-DNA delivery. The Agrobacterium-mediated transformation of leaf disk explants were found significantly related to physiological age of the explants, age and origin of the of the donor plant. Leaf explants from second node of the 3-month-old in vivo plants showed highest transformation efficiency (94.3%) revealed by transient GUS expression assay. Plants selected on medium containing 20 mg/l kanamycin showed stable GUS expression in leaves and stem. The elongated shoots readily developed roots on kanamycin-free rooting medium and on transfer to soil, plants were successfully established. Polymerase chain reaction (PCR) and reverse-transcriptase PCR analysis in putative plants confirmed their transgenic nature. The established transformation method should provide new opportunities for the genetic improvement of patchouli for desirable trait.


Subject(s)
Agrobacterium/genetics , Lamiaceae/cytology , Lamiaceae/genetics , Plant Extracts/pharmacology , Plant Leaves/cytology , Plant Leaves/genetics , Transformation, Genetic , Acetophenones/pharmacology , Agrobacterium/cytology , Agrobacterium/drug effects , Agrobacterium/growth & development , Anti-Bacterial Agents/pharmacology , Biological Transport , Cell Survival/drug effects , Coculture Techniques , DNA, Bacterial/genetics , DNA, Bacterial/metabolism , Glucuronidase/genetics , Glutamine/pharmacology , Kanamycin/pharmacology , Lamiaceae/chemistry , Lamiaceae/growth & development , Plant Leaves/chemistry , Plant Leaves/growth & development , Plant Shoots/drug effects , Plant Shoots/physiology , Polysorbates/pharmacology , Regeneration/drug effects , Temperature , Time Factors , Transformation, Genetic/drug effects
20.
Planta Med ; 78(2): 128-31, 2012 Jan.
Article in English | MEDLINE | ID: mdl-22002849

ABSTRACT

Twenty-six microbiologically inactive (MIC > 512 µg/mL) convolvulaceous resin glycosides ( 1- 26) were tested for resistance modulatory activity in vitro against Escherichia coli Rosetta-gami and two nosocomial pathogens, Salmonella typhi and Shigella flexneri. These compounds exerted a potentiation effect of the clinically useful antibiotics tetracycline, kanamycin, and chloramphenicol against the tested gram-negative bacteria by increasing antibiotic susceptibility up to 32-fold at concentrations of 25 µg/mL. Therefore, the oligosaccharides from the morning glory family (Convolvulaceae) represent metabolites that reverse microbial resistance mechanisms, favoring an increase in the strength and effectiveness of current antibiotics that are not effective in the treatment of refractive infections caused by multidrug-resistant strains.


Subject(s)
Anti-Bacterial Agents/pharmacology , Drug Resistance/drug effects , Glycosides/pharmacology , Gram-Negative Bacteria/drug effects , Ipomoea/chemistry , Plant Extracts/pharmacology , Resins, Plant/pharmacology , Chloramphenicol/pharmacology , Drug Synergism , Escherichia coli/drug effects , Kanamycin/pharmacology , Microbial Sensitivity Tests , Oligosaccharides/pharmacology , Salmonella/drug effects , Shigella/drug effects , Tetracycline/pharmacology
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