Zingiber officinale: Its antibacterial activity on Pseudomonas aeruginosa and mode of action evaluated by flow cytometry.

Biofilm formation, low membrane permeability and efflux activity developed by Pseudomonas aeruginosa, play an important role in the mechanism of infection and antimicrobial resistance. In the present study we evaluate the antibacterial effect of Zingiber officinale against multi-drug resistant strain of P. aeruginosa. The study explores antibacterial efficacy and time-kill study concomitantly the effect of herbal extract on bacterial cell physiology with the use of flow cytometry and inhibition of biofilm formation. Z. officinale was found to inhibit the growth of P. aeruginosa, significantly. A major decline in the Colony Forming Units was observed with 3 log10 at 12 h of treatment. Also it is found to affect the membrane integrity of the pathogen, as 70.06 ± 2.009% cells were found to stain with Propidium iodide. In case of efflux activity 86.9 ± 2.08% cells were found in Ethidium bromide positive region. Biofilm formation inhibition ability was found in the range of 68.13 ± 4.11% to 84.86 ± 2.02%. Z.officinale is effective for killing Multi-Drug Resistant P. aeruginosa clinical isolate by affecting the cellular physiology and inhibiting the biofilm formation.

Effect of aquo-alchoholic extract of Glycyrrhiza glabra against Pseudomonas aeruginosa in Mice Lung Infection Model.

The prevalence of lung infection caused by Pseudomonas aeruginosa strains that are classified as multi-drug resistant has increased considerably and is mainly attributed to relative insufficiency of potent chemotherapeutic modalities. The present study was conducted to evaluate the antimicrobial activity of aquo-alcoholic extract of Glycyrrhiza glabra against the P. aeruginosa causing lung infection in Swiss albino mice. The study involves evaluation of lethal dose of P. aeruginosa in Swiss albino mice and analysis of disease manifestation that includes bacteremia, hypothermia, reduction in body weight and other parameters for 48h of infection. Physical manifestations of infected mice showed a significant decline in body temperature that is 29±0.57°C (at 48th h) from 38.81±0.33°C (0h) and 30% weight loss was observed at the end of the study. Further the efficacy of G. glabra extract against lung infection induced with the calculated lethal dose was evaluated by employing bacteremia, histopathology and radiological analysis. Bacterial burden showed that 2.30±0.02 Log10CFU/mL at day 7, a significant decline in the bacterial load as compared to day 1 when the bacterial burden was found to be 3.32±0.1 Log10CFU/mL. Histopathological results showed more diffuse and patchy accumulation of inflammatory cells within the alveolar space also the infiltrates were noted in all the lung section of infected mice. In treated animal group improved lung histology was seen with the exudates were less seen in D1 dose (20mg/kg) and disappeared in D2 dose (80mg/kg). The study clearly declares that the G. glabra extract is effective against lung infection caused by P. aeruginosa at dose of 80mg/kg. The LCMS results revealed that the extract contains Glycyrrhizin, Stigmasterol and Ergosterol, Licochalcone and Glabridin. The current study expected to further exploit the biomedical properties of this extract in the preparation of a potent regimen against such threatening pathogen.

In vivo anti-arthritic efficacy of Camellia sinensis (L.) in collagen induced arthritis model.

BACKGROUND: Rheumatoid arthritis (RA), an autoimmune inflammatory disorder with synovial hyperplasia, destruction of cartilage, bone damage is often associated with risk of infections. Such risk could be attributed towards usage of immunosuppressive agents. Thus, the present study was undertaken to evaluate the anti-arthritic efficacy of aquo-alcoholic extract of Camellia sinensis (L.). MATERIAL AND METHODS: Dried leaves of Camellia sinensis (L.) or Cs were filtered and extracted in 1:1 aqueous: ethanol by Soxhlet apparatus followed by lyophilization and spray drying to develop amorphous powder. Four different oral doses (50, 100, 200, 400mg/kg/body wt.) of aquo-alcoholic extract were evaluated for anti-edematogenic effect in collagen induced arthritis model. The selected anti-arthritic doses of Cs were evaluated for the oxidative stress markers like Glutathione [5-5'dithio-bis-2-nitrobenzoicacid (DTNB)], Superoxide dismutase [Epinephrine], Catalase [Hydrogen peroxide], Lipid peroxidation [Thiobarbituric acid reactive substance (TBARS)], Nitric oxide [Griess reagents:Nitrobluetetrazolium], Articular elastase [N-methoxysuccinyl-Ala-Ala-Pro- Val p-nitroanilide] in joints followed by haematological evaluation including RBC, WBC, Haemoglobin, platelets and haematocrit. To validate these biochemical changes, the radiological and histopathological (Haematoxylin & Eosin) evaluation was also conducted. RESULTS: The selected anti-arthritic dose of Cs i.e. 400mg/kg/body wt. (∼60% anti-arthritic efficacy on 35th day) could be attributed towards significant (p<0.05) increase in the levels of enzymatic (Superoxide dismutase and Catalase) and non-enzymatic (Glutathione) antioxidants by 34%, 59% and 50% respectively. Simultaneously, the significant (p<0.05) reduction of lipid peroxides, nitrite radical and elastase activity by 32%, 45% & 32% respectively as compare to control indicated overall decrease in oxidative stress. Haematological evaluation revealed restoration of RBC, WBC and platelets level in treatment group. The confirmatory analysis utilizing radiological and histological assessment showed alleviation of joint deformity, tissue swelling, pannus formation and neutrophils infiltration in treatment group as compared to collagen induced arthritis. CONCLUSION: The analysis showed that Cs can play an effective role in reduction of oxidative stress by modulating levels of antioxidants, reducing levels of free radicals while restoring normal haematopoietic cascade as observed in collagen induced arthritis model. Thus, the cumulative dose impact of 400mg/kg body wt., over a period of 14days also found extremely effective in terms of safeguarding their structural conformity against such auto-immune disorder.

Effect of Holarrhena antidysentrica (Ha) and Andrographis paniculata (Ap) on the biofilm formation and cell membrane integrity of opportunistic pathogen Salmonella typhimurium.

Increasing occurrence of gastroenteritis outbreaks caused by food borne opportunistic microorganisms has become a major problem in food industry as well as in immunocompromised host. Antimicrobial agents are losing their efficacy due to increase in the microbial resistance. For such reasons, conventional treatment has become limited to manage the infections state. Need of the hour is to instigate the search for safer holistic alternatives. The present study was hence conducted to assess the antibiofilm effect and mode of action of aquo alcoholic extracts of Holarrhena antidysentrica (Ha) and Andrographis paniculata (Ap) against the Salmonella enterica serovar typhimurium. Both the extracts were screened for the presence of phytocompounds followed by the characterization using Attenuated Total Reflection (ATR) infrared spectroscopy and bioactivity finger print analysis. Anti-biofilm assays were determined to test the potential of both extracts to inhibit the biofilm formation, while Propidium Iodide (PI) uptake analysis revealed that cell membrane was damaged by the exposure of nutraceuticals for 1 h. This study has demonstrated that both nutraceuticals have anti-biofilm and antimicrobial activity perturbing the membrane integrity of food-borne S. typhimurium and could be used as curative remedy to control the food borne microbial infection.

Alternative to antibiotics against Pseudomonas aeruginosa: Effects of Glycyrrhiza glabra on membrane permeability and inhibition of efflux activity and biofilm formation in Pseudomonas aeruginosa and its in vitro time-kill activity.

The multi-drug resistance offered by Pseudomonas aeruginosa to antibiotics can be attributed towards its propensity to develop biofilm, modification in cell membrane and to efflux antibacterial drugs. The present study explored the activity of Glycyrrhiza glabra and one of its pure compounds, glycyrrhizic acid against P. aeruginosa and their mechanism of action in terms of the effect on membrane permeability, efflux activity, and biofilm formation were determined. Minimum inhibitory concentrations were determined by using broth dilution technique. The minimum bactericidal concentrations were assessed on agar plate. The MIC of the extract and glycyrrhizic acid was found to be 200 and 100 µg ml(-1), respectively. The MBC was found to be 800 and 400 µg ml(-1) in the case of extract and glycyrrhizic acid, respectively. Time -dependent killing efficacy was also estimated. Flowcytometric analysis with staining methods was used to determine the effect of extract and glycyrrhizic acid at 2 × MIC on different physiological parameters and compared it with the standard (antibiotic). The growth of P. aeruginosa was significantly inhibited by extract and the pure compound. The herbal extract and the glycyrrhic acid were also found to effective in targeting the physiological parameters of the bacteria that involve cell membrane permeabilization, efflux activity, and biofilm formation. This study reports the antipseudomonal action of Glycyrrhiza glabra and one of its compound and provides insight into their mode of action.

In vitro bactericidal activity of promising nutraceuticals for targeting multidrug resistant Pseudomonas aeruginosa.

OBJECTIVE: We evaluated the bactericidal activity of nutraceuticals against multidrug resistant Pseudomonas aeruginosa. The nutritionally valued herbs were screened on the basis of a matrix modeling approach and molecular docking based validation analysis. METHODS: The database of 38 herbs developed earlier using fuzzy logic based scoring analysis was subjected to molecular docking based validation. The molecular docking (Hex 6.12) analyses of predominant phytoligands (∼10 per herb) against exoenzyme S of P. aeruginosa filtered potent herbs were selected. The preauthenticated bacterial inoculum (10(8) CFU/mL) was added to the sterile nutrient broth impregnated with standardized aqueous-alcoholic herbal extracts (1-1600 µg/mL). After overnight incubation at 37°C, antibacterial activity was evaluated in terms of minimum inhibitory and minimum bactericidal concentrations. RESULTS: Five herbs were selected on the basis of fuzzy set scoring, an herbal informatics model, and validation analysis based on energy of docking (i.e., Evalue of 380) phytoligands with maximum scoring obtained by Glycyrrhiza glabra. Among the 5 nutraceuticals, G. glabra showed maximum bactericidal activity significantly (P < 0.05) higher than Amikacin, a standard antibiotic, which was in consonance with in silico bioprospection. Zingiber officinale, despite a low Evalue, showed considerably higher inhibition attributed to its higher flavonoid content as compared to other herbs. CONCLUSION: G. glabra (licorice), a flavoring agent; Z. officinale (ginger), a condiment; and Mentha piperita (mint), a fragrance component, showed significant therapeutic potential against multidrug resistant strains of P. aeruginosa.

Attenuation of adhesion, quorum sensing and biofilm mediated virulence of carbapenem resistant Escherichia coli by selected natural plant products.

The multi-drug resistance offered by Carbapenem Resistant Escherichia coli (Family: Enterobacteriaceae; Class: Gammaproteobacteria) against third line antibiotics can be attributed towards its ability to develop biofilm. Such process involves adhesion and quorum-sensing induced colonization leading to biomass development. The present study explored the anti-adhesion, anti-quorum sensing and anti-biofilm potential of 05 pre-standardized potent herbals. Berberis aristata (PTRC-2111-A) exhibited maximum potential in all these activities i.e. 91.3% ± 0.05% (Anti-adhesion), 96.06% ± 0.05% (Anti-Quorum sensing) and 51.3% ± 0.07% (Anti-Biofilm formation) respectively. Camellia sinensis (PTRC-31911-A) showed both anti-adhesion (84.1% ± 0.03%) and anti-quorum sensing (90.0%) potential while Holarrhena antidysenterica (PTRC-8111-A) showed only anti-quorum sensing potential as compared to standards/antibiotics. These findings were in line with the molecular docking analysis of phytoligands against Lux S and Pilin receptors. Furthermore, the pairwise correlation analysis of the tested activities with qualitative, quantitative and bioactivity functional descriptors revealed that an increased content of alkaloid, moderate content of flavonoids and decreased content of tannins supported all the three activities. In addition, nitric oxide and superoxide scavenging activity were found to be correlated with anti-quorum sensing activity. The findings indicated clearly that B. aristata (Family: Berberidaceae) and C. sinensis (Family: Theaceae) were potent herbal leads with significant therapeutic potential which further needs to be explored at pre-clinical level in the future.

Camellia sinensis Ameliorates the Efficacy of Last Line Antibiotics Against Carbapenem Resistant Escherichia coli.

Aquo-ethanolic extract of Camellia sinensis (PTRC-31911-A), standardized using Fourier transform infrared analysis, was found to have seven common functional groups in comparison with pre-identified marker compound 'quercetin'. Phyto-chemical quantitation analysis revealed the presence of 10.65 µg/mg of flavonoids. The bioactivity fingerprint profile of PTRC-31911-A includes IC50 (Hydroxyl radical site specific scavenging) = 11.36 ± 0.5 µg/mL, IC80 (Hydroxyl radical non-site specific scavenging) = 26.44 ± 0.5 µg/mL and IC50 (Superoxide ion scavenging) = 10.141 ± 0.5 µg/mL. The drug combination analysis of PTRC-31911-A with five third-line antibiotics was carried out against carbapenem-resistant Escherichia coli. The analysis of combination of PTRC-31911-A (6.25-1000 µg/mL) and antibiotics (6.25-1000 µg/mL) revealed synergistic behaviour (fractional inhibitory concentration indices < 1) with tigecycline, ertapenem, meropenem, colistin and augmentin. The lead combination of PTRC-31911-A + ertapenem or meropenem showed maximum augmentative potential at 50 and 100 µg/mL, respectively, with nearly five-fold decrease in minimum inhibitory concentrations as compared with respective antibiotics alone. The synergistic effects implied that the antibacterial combinations of PTRC-31911-A and ertapenem, meropenem, colistin, tigecycline or augmentin would be more effective than a single monotherapy with either of the antibacterial agent.