Rizeng Meng

Antifungal activity of thymol against clinical isolates of fluconazole-sensitive and -resistant Candida albicans.

Thymol (THY) was found to have in vitro antifungal activity against 24 fluconazole (FLC)-resistant and 12 FLC-susceptible clinical isolates of Candida albicans, standard strain ATCC 10231 and one experimentally induced FLC-resistant C. albicans S-1. In addition, synergism was observed for clinical isolates of C. albicans with combinations of THY-FLC and THY-amphotericin B (AMB) evaluated by the chequerboard microdilution method. The interaction intensity was determined by spectrophotometry for the chequerboard assay, and the nature of the interactions was assessed using two non-parametric approaches [fractional inhibitory concentration index (FICI) and DeltaE models]. The interaction between THY-FLC or THY-AMB in FLC-resistant and -susceptible strains of C. albicans showed a high percentage of synergism by the FICI method and the DeltaE method. The DeltaE model gave results consistent with FICI, and no antagonistic action was observed in the strains tested.

In vitro and in vivo interactions between fluconazole and allicin against clinical isolates of fluconazole-resistant Candida albicans determined by alternative methods

A checkerboard microdilution method, performed according to the recommendations of the National Committee for Clinical Laboratory Standards, was used to study the in vitro interaction of fluconazole and allicin in 24 fluconazole-resistant clinical isolates of Candida albicans, one experimentally induced strain S-1, and one ATCC type strain 10231. The interaction intensity was determined by spectrophotometric methods and visual reading of the checkerboard assay, and the nature of the interactions was assessed using two nonparametric approaches [fractional inhibitory concentration index (FICI) and DeltaE models]. Synergism was observed in 23 strains using FICI, and in 22 strains using DeltaE. The DeltaE model gave results consistent with FICI, but no antagonistic action was observed. The positive interactions were also confirmed by the time-killing test and agar diffusion in the selected strains. Moreover, the in vivo experiment showed that a combination of fluconazole and allicin exhibited a good synergism against C. albicans.

Transcriptional and Functional Analysis of the Effects of Magnolol: Inhibition of Autolysis and Biofilms in Staphylococcus aureus

Background The targeting of Staphylococcus aureus biofilm structures are now gaining interest as an alternative strategy for developing new types of antimicrobial agents. Magnolol (MOL) shows inhibitory activity against S. aureus biofilms and Triton X-100-induced autolysis in vitro, although there are no data regarding the molecular mechanisms of MOL action in bacteria. Methodology/Principal Findings The molecular basis of the markedly reduced autolytic phenotype and biofilm inhibition triggered by MOL were explored using transcriptomic analysis, and the transcription of important genes were verified by real-time RT-PCR. The inhibition of autolysis by MOL was evaluated using quantitative bacteriolytic assays and zymographic analysis, and antibiofilm activity assays and confocal laser scanning microscopy were used to elucidate the inhibition of biofilm formation caused by MOL in 20 clinical isolates or standard strains. The reduction in cidA, atl, sle1, and lytN transcript levels following MOL treatment was consistent with the induced expression of their autolytic repressors lrgA, lrgB, arlR, and sarA. MOL generally inhibited or reversed the expression of most of the genes involved in biofilm production. The growth of S. aureus strain ATCC 25923 in the presence of MOL dose-dependently led to decreases in Triton X-100-induced autolysis, extracellular murein hydrolase activity, and the amount of extracellular DNA (eDNA). MOL may impede biofilm formation by reducing the expression of cidA, a murein hydrolase regulator, to inhibit autolysis and eDNA release, or MOL may directly repress biofilm formation. Conclusions/Significance MOL shows in vitro antimicrobial activity against clinical and standard S. aureus strains grown in planktonic and biofilm cultures, suggesting that the structure of MOL may potentially be used as a basis for the development of drugs targeting biofilms.

Global gene expression profile of Saccharomyces cerevisiae induced by dictamnine

Dictamnine, a natural plant product, has been reported to have antimicrobial activity against bacteria and fungi; however, the dictamnine response mechanisms of microorganisms are still poorly understood. We have shown that dictamnine has antimicrobial activities against the model fungus Saccharomyces cerevisiae, with a minimum inhibitory concentration (MIC) value of 64 µg/ml. Commercial oligonucleotide microarrays were used to determine the global transcriptional response of S. cerevisiae triggered by treatment with dictamnine. We interpreted our microarray data using the hierarchical clustering tool, T‐profiler. Several major transcriptional responses were induced by dictamnine. The first was the induced environmental stress response, mainly under the control of the Msn2p and Msn4p transcription factors, and the repressed environmental stress response in genes containing the PAC (RNA polymerase A and C box) and rRPE (ribosomal RNA processing element) motifs. The second was the Upc2p‐mediated response involved in lipid biosynthesis. The third comprised the PDR3‐ and RPN4‐mediated responses involved in multidrug resistance (MDR). Finally, the TBP‐mediated response was induced with dictamnine treatment. TBP is an essential general transcription factor involved in directing the transcription of genes. Quantitative real‐time RT–PCR was performed on selected genes to verify the microarray results. Furthermore, morphological transitions during dictamnine exposure to S. cerevisiae L1190 (MATa/α) were examined, using confocal laser microscopy. Copyright

Microarray analysis of p-anisaldehyde-induced transcriptome of Saccharomyces cerevisiae

Abstractp-Anisaldehyde (4-methoxybenzaldehyde), an extract from Pimpinella anisum L. seeds, is a potential novel preservative. To reveal the possible action mechanism of p-anisaldehyde against microorganisms, yeast-based commercial oligonucleotide microarrays were used to analyze the genome-wide transcriptional changes in response to p-anisaldehyde. Quantitative real-time RT-PCR was performed for selected genes to verify the microarray results. We interpreted our microarray data with the clustering tool, T-profiler. Analysis of microarray data revealed that p-anisaldehyde induced the expression of genes related to sulphur assimilation, aromatic aldehydes metabolism, and secondary metabolism, which demonstrated that the addition of p-anisaldehyde may influence the normal metabolism of aromatic aldehydes. This genome-wide transcriptomics approach revealed first insights into the response of Saccharomyces cerevisiae (S. cerevisiae) to p-anisaldehyde challenge.

Allicin-induced global gene expression profile of Saccharomyces cerevisiae

To understand the response mechanisms of fungus cells upon exposure to the natural fungicide allicin, we performed commercial oligonucleotide microarrays to determine the overall transcriptional response of allicin-treated Saccharomyces cerevisiae strain L1190. Compared with the transcriptional profiles of untreated cultures, 147 genes were significantly upregulated, and 145 genes were significantly downregulated in the allicin-treated cells. We interpreted the microarray data with the hierarchical clustering tool, T-profiler. Major transcriptional responses were induced by allicin and included the following: first, Rpn4p-mediated responses involved in proteasome gene expression; second, the Rsc1p-mediated response involved in iron ion transporter activity; third, the Gcn4p-mediated response, also known as general amino acid control; finally, the Yap1p-, Msn2/4p-, Crz1p-, and Cin5p-mediated multiple stress response. Interestingly, allicin treatment, similar to mycotoxin patulin and artificial fungicide thiuram treatment, was found to induce genes involved in sulfur amino acid metabolism and the defense system for oxidative stress, especially DNA repair, which suggests a potential mutagenicity for allicin. Quantitative real-time reverse transcription-polymerase chain reaction was performed for selected genes to verify the microarray results. To our knowledge, this is the first report of the global transcriptional profiling of allicin-treated S. cerevisiae by microarray.

Antioxidative and anticancer properties of Licochalcone A from licorice

ETHNOPHARMACOLOGICAL RELEVANCE Licochalcone A (LCA) is a characteristic chalcone that is found in licorice, which is a traditional medicinal plant. In traditional medicine, LCA possesses many potential biological activities, including anti-parasitic, anti-inflammatory and antitumor activities. AIM OF THE STUDY To determine the antioxidant activity of LCA and, on this basis, to investigate the role of its anticancer activity. MATERIALS AND METHODS To validate the antioxidant activity of LCA, the proteins SOD, CAT and GPx1 were analyzed using western blotting and cellular antioxidant activity (CAA) assays. Oxidative free radicals are associated with cancer cells. Therefore, the anticancer activity of LCA was also evaluated. To assess the anticancer activity, cell viability assays were performed and apoptosis was evaluated. In addition, MAPK-related proteins were analyzed using western blotting. RESULTS The experimental data showed that the EC50 of LCA is 58.79±0.05μg/mL and 46.29±0.05μg/mL under the two conditions tested, with or without PBS. In addition, LCA at a concentration of approximately 2-8μg/mL can induce the expression of SOD, CAT and GPx1 proteins. Further, LCA inhibits the growth of HepG2 cells through cell proliferation arrest and the subsequent induction of apoptosis, and LCA attenuated the p38/JNK/ERK signaling pathway in a dose-dependent manner. CONCLUSION The results showed that LCA suppresses the oxidation of cells and markedly inhibits the proliferation of cancer cells. These findings confirm the traditional use of LCA in folk medicine.

Antimicrobial, antioxidant, and antitumor activity of epsilon-poly-L-lysine and citral, alone or in combination

Background Food safety is an important worldwide public health concern, and microbial contamination in foods not only leads to food deterioration and shelf life reduction but also results in economic losses and disease. Objective The main aim of the present study was to evaluate the effect of epsilon-poly-L-lysine (ε-PL) and citral combination against Escherichia coli O157:H7 (E. coli O157:H7) strains. The preliminary antioxidant and antitumor activities were also studied. Design Synergism is a positive interaction created when two compounds combine and exert an inhibitory effect that is greater than the sum of their individual effects. The synergistic antimicrobial effect of ε-PL and citral was studied using the checkerboard method against E. coli O157:H7. The minimal inhibitory concentration, time-kill, and scanning electron microscope assays were used to determine the antimicrobial activity of ε-PL and citral alone or in combination; 2,2-diphenyl-1-picrylhydrazyl-scavenging assay and western blotting were used in antioxidant activity assays; cell viability assay was carried out to finish preliminary antitumor test. Results Minimal inhibitory concentrations of ε-PL and citral resisted to the five E. coli O157:H7 strains were 2–4 µg/mL and 0.5–1 µg/mL, and the fractional inhibitory concentration indices were 0.25–0.375. The results of time-kill assay revealed that a stronger bactericidal effect in a laboratory medium might be exerted in the combination against E. coli O157:H7 than that in a food model. The compounds alone or in combination exhibited a potential 2,2-diphenyl-1-picrylhydrazyl radical–scavenging activity, and the expression of superoxide dismutase 1 and glutathione peroxidase 1 protein increased. The preliminary antitumor activity effect of the combination was better than ε-PL or citral alone. Conclusions These findings indicated that the combination of ε-PL and citral could not only be used as a promising naturally sourced food preservative but also be used in the pharmaceutical industry.

The synergy of berberine chloride and totarol against Staphylococcus aureus grown in planktonic and biofilm cultures.

Staphylococcus aureus (S. aureus) is commonly associated with hospital-acquired infections and is known to form biofilms. Bacteria inside biofilms display an increased resistance to chemotherapeutics and host immune defences. Efficient antibiotics or combination therapy are urgently needed to treat patients with biofilm-associated MRSA infections. The objective of the current study was to evaluate the in vitro antimicrobial activities of totarol alone or in combination with berberine chloride (BBR) against S. aureus grown in planktonic and biofilm cultures. The synergistic antimicrobial effects between BBR and totarol were observed in all tested strains grown in biofilms using a chequerboard microdilution method, with the fractional inhibitory concentration index values ranging from 0.125 to 0.375. No antagonistic activity was observed in any of the strains tested in suspension or biofilm cultures. The synergistic activity against S. aureus biofilms was also corroborated by confocal laser scanning microscopy and adhesion assays. Moreover, the present study demonstrated that combination BBR and totarol treatment effectively decreased the formation of S. aureus biofilms by affecting extracellular genomic DNA release and polysaccharide intercellular adhesin expression. Subsequently, real-time reverse transcriptase PCR analysis revealed that the combination of BBR and totarol effectively inhibited the transcription of the biofilm-related genes sarA, cidA and icaA. These results suggest that the combination of totarol and BBR is momentous for the further development of a therapy protocol against S. aureus biofilms.

Genome-wide transcription analyses in Mycobacterium tuberculosis treated with lupulone

Mycobacterium tuberculosis (M. tuberculosis), the causative agent of tuberculosis, still causes higher mortality than any other bacterial pathogen until now. With the emergence and spread of multidrug-resistant (MDR) and extensively drug-resistant (XDR-TB) strains, it becomes more important to search for alternative targets to develop new antimycobacterial drugs. Lupulone is a compound extracted from Hops (Hurnulus lupulus), which exhibits a good antimicrobial activity against M. tuberculosis with minimal inhibitory concentration (MIC) value of 10 μg/mL, but the response mechanisms of lupulone against M. tuberculosis are still poorly understood. In this study, we used a commercial oligonucleotide microarray to determine the overall transcriptional response of M. tuberculosis H37Rv triggered by exposure to MIC of lupulone. A total of 540 genes were found to be differentially regulated by lupulone. Of these, 254 genes were upregulated, and 286 genes were downregulated. A number of important genes were significantly regulated which are involved in various pathways, such as surface-exposed lipids, cytochrome P450 enzymes, PE/PPE multigene families, ABC transporters, and protein synthesis. Real-time quantitative RT-PCR was performed for choosed genes to verified the microarray results. To our knowledge, this genome-wide transcriptomics approach has produced the first insights into the response of M. tuberculosis to a lupulone challenge.