Vitamin C Enhances the Antibacterial Activity of Honey against Planktonic and Biofilm-Embedded Bacteria.

Multifactorial antibacterial action is an important feature of honey; however, its bactericidal efficacy against biofilm-embedded bacteria is limited. The aim of this study was to investigate the impact of vitamin C (Vit C) on the antibacterial activity of natural honeys against planktonic as well as biofilm-embedded bacterial pathogens. The antibacterial activity of four honey samples supplemented with Vit C was expressed as the minimum inhibitory concentration (MIC). At sub-MICs, Vit C significantly increased the antibacterial activity of the tested honeys against Pseudomonas aeruginosa in planktonic cultures. However, after supplementation, honeydew honey, the most active honey, was ineffective against Staphylococcus aureus. On the other hand, when 100% honeydew honey was supplemented with Vit C (100 mg/g of honey) in a multispecies wound biofilm model, complete eradication of almost all bacterial isolates, including S. aureus, was observed. Furthermore, a mixture of honey and Vit C was partially effective against Enterococcus faecalis, whereas honey alone exhibited no antibacterial activity against this bacterium. Vit C counteracted hydrogen peroxide in honey solution and, thus, eliminated the major antibacterial compound present in honey. It is likely that a combination of honey with Vit C may trigger the intracellular production of reactive oxygen species in bacterial cells, but the exact cellular mechanisms warrant further investigations.

Quinine Enhances Photo-Inactivation of Gram-Negative Bacteria.

BACKGROUND: Antimicrobial resistance is a significant concern to public health, and there is a pressing need to develop novel antimicrobial therapeutic modalities. METHODS: In this study, we investigated the capacity for quinine hydrochloride (Q-HCL) to enhance the antimicrobial effects of antimicrobial blue light ([aBL] 405 nm wavelength) against multidrug-resistant (MDR) Gram-negative bacteria in vitro and in vivo. RESULTS: Our findings demonstrated the significant improvement in the inactivation of MDR Pseudomonas aeruginosa and Acinetobacter baumannii (planktonic cells and biofilms) when aBL was illuminated during Q-HCL exposure. Furthermore, the addition of Q-HCL significantly potentiated the antimicrobial effects of aBL in a mouse skin abrasion infection model. In addition, combined exposure of aBL and Q-HCL did not result in any significant apoptosis when exposed to uninfected mouse skin. CONCLUSIONS: In conclusion, aBL in combination with Q-HCL may offer a novel approach for the treatment of infections caused by MDR bacteria.

Natural and non-toxic products from Fabaceae Brazilian plants as a replacement for traditional antifouling biocides: an inhibition potential against initial biofouling.

In this study, we screened for the antifouling activity of 15 species plant extracts from Brazilian the Brazilian Caatinga Fabaceae against the initial colonization of natural marine bacterial biofilm. We also investigated the potential toxicity of extracts against planktonic and benthic non-target organisms. Aqueous extracts of plants collected in the Caatinga biome (PE, Brazil) were prepared and tested at different concentration levels (0, 0.5, 1, 2, 4, and 8 mg mL-1). Natural marine bacterial consortium was inoculated in multi-well plates and incubated with the different treatments for 48 h. The biofilm and planktonic bacterial density and biomass inhibition were evaluated along with biofilm biomass eradication. The extracts that showed the highest bacterial biofilm inhibition were evaluated for toxicity against microalgae and crustaceans. The biofilm and planktonic bacterial inhibition potential were evaluated through flow cytometry and spectrophotometry. The selected treatments were evaluated for their toxicity using the microalgae Chaetoceros calcitrans, the copepod Nitokra sp., and the brine shrimp Artemia salina as bioindicators. Our work demonstrates the biotechnological potential of Fabaceae plant compounds as a safe antifouling alternative. Anadenanthera colubrina var. cebil fruits and Apuleia leiocarpa leaf extracts showed antibiofilm activity (≥ 80%), while Myroxylon peruiferum and Dioclea grandiflora leaf extracts showed antibiotic activity. These extracts were safe to planktonic and benthic non-target organisms. The results of this study point to potential substitutes to highly toxic antifouling paints and shed light on the prospect of a yet to be explored biome for more sustainable alternatives in biofouling research.

Exploring different photosensitizers to optimize elimination of planktonic and biofilm forms of Enterococcus faecalis from infected root canal during antimicrobial photodynamic therapy.

BACKGROUND: Despite the high success rate of endodontic treatment, failure may occur in some cases. In this case, Enterococcus faecalis is the most common species in endodontic treatment failure and post-treatment apical periodontitis. Therefore, a new adjunctive strategy is needed for the prevention of endodontic infections due to E. faecalis. The aim of the present study was to compare the antimicrobial and anti-biofilm activities of different common photosensitizers (PSs) for use in antimicrobial photodynamic therapy (aPDT) against E. faecalis. MATERIALS AND METHODS: E. faecalis strain ATCC 29212 was used as the tested strain and methylene blue (MB), toluidine blue O (TBO), indocyanine green (ICG), and curcumin (CUR) were used as PSs. Irradiation was carried out using diode laser and light emitting diode (LED) at wavelengths related to the above PSs. Then, antimicrobial and anti-biofilm activities were measured using the microbial viability assay and crystal violet test, respectively. RESULTS: aPDT with using the above PSs significantly decreased the CFU/mL count of E. faecalis compared to the control group (P < 0.05). The killing percentage of E. faecalis via PS mediated aPDT was 99.6%, 98.2%, 85.1%, and 65.0% for CUR, ICG, TBO, and MB, respectively. aPDT using the above PSs significantly decreased the biofilm formation ability of E. faecalis compared to the control group (P < 0.05). The biofilm reduction percentage of the PSs was 68.4%, 62.9%, 59.0%, and 47.6% for CUR, ICG, TBO, and MB, respectively. CONCLUSION: CUR and ICG mediated aPDT exhibited considerably more antimicrobial activity than other PSs, while TBO and MB demonstrated weaker anti-biofilm effects against E. faecalis compared to other PSs.

Differences in the accumulation of phosphorus between vegetative cells and heterocysts in the cyanobacterium Nodularia spumigena.

The cyanobacterium Nodularia spumigena is a species that frequently forms blooms in the Baltic Sea. Accumulation of the vital nutrient phosphorus (P) apparently plays an important role in the ability of this and other cyanobacteria to grow even when dissolved inorganic phosphorus is depleted. However, until now, this has not been studied in N. spumigena at the cellular level. Therefore, in this study, phosphorus incorporation and distribution in cyanobacterial filaments over time was examined by scanning electron microscopy in combination with energy dispersive X-ray analysis (SEM/EDX) and nanoscale secondary ion mass spectrometry (NanoSIMS). Immediately after phosphate addition to a phosphorus-depleted population, the phosphate concentration decreased in the water while intracellular polyphosphate accumulated. Microscopically, phosphorus in form of polyphosphate granules was stored preferentially in vegetative cells, whereas heterocysts remained low in intracellular phosphorus. This information is an essential step towards understanding the phosphorus dynamics of this species and demonstrates that the division of tasks between vegetative cells and heterocysts is not restricted to nitrogen fixation.

The effect of emodin on Staphylococcus aureus strains in planktonic form and biofilm formation in vitro.

Staphylococcus aureus (S. aureus) is a Gram-positive pathogen and forms biofilm easily. Bacteria inside biofilms display an increased resistance to antibiotics and disinfectants. The objective of the current study was to assess the antimicrobial activities of emodin, 1,2,8-trihydroxy-6-methylanthraquinone, an anthraquinone derivative isolated from Polygonum cuspidatum and Rheum palmatum, against S. aureus CMCC26003 grown in planktonic and biofilm cultures in vitro. In addition, a possible synergistic effect between emodin and berberine chloride was evaluated. As quantified by crystal violet method, emodin significantly decreased S. aureus biofilm growth in a dose-dependent manner. The above findings were further supported by scanning electron microscopy. Moreover, the present study demonstrated that sub-MICs emodin obviously intervened the release of extracellular DNA and inhibited expression of the biofilm-related genes (cidA, icaA, dltB, agrA, sortaseA and sarA) by real-time RT-PCR. These results revealed a promising application for emodin as a therapeutic agent and an effective strategy to prevent S. aureus biofilm-related infections.

Effects of grazing, phosphorus and light on the growth rates of major bacterioplankton taxa in the coastal NW Mediterranean.

Estimation of growth rates is crucial to understand the ecological role of prokaryotes and their contribution to marine biogeochemical cycling. However, there are only a few estimates for individual taxa. Two top-down (grazing) and bottom-up (phosphorus (P) availability) manipulation experiments were conducted under different light regimes in the NW Mediterranean Sea. Growth rate of different phylogenetic groups, including the Bacteroidetes, Rhodobacteraceae, SAR11, Gammaproteobacteria and its subgroups Alteromonadaceae and the NOR5/OM60 clade, were estimated from changes in cell numbers. Maximal growth rates were achieved in the P-amended treatments but when comparing values between treatments (response ratios), the response to predation removal was in general larger than to P-amendment. The Alteromonadaceae displayed the highest rates in both experiments followed by the Rhodobacteraceae, but all groups largely responded to filtration and P-amendment, even the SAR11 which presented low growth rates. Comparing light and dark treatments, growth rates were on average equal or higher in the dark than in the light for all groups, except for the Rhodobacteraceae and particularly the NOR5 clade, groups that contain photoheterotrophic species. These results are useful to evaluate the potential contributions of different bacterial types to biogeochemical processes under changing environmental conditions.

The Antimicrobial Peptides P-113Du and P-113Tri Function against Candida albicans.

Two antimicrobial P-113 peptide derivatives, P-113Du and P-113Tri, were investigated in this study. Notably, P-113Du and P-113Tri contained significant fractions of α-helix conformation and were less sensitive to high salt and low pH than P-113. Moreover, compared to P-113, these peptides exhibited increased antifungal activity against planktonic cells, biofilm cells, and clinical isolates of Candida albicans and non-albicans Candida spp. These results suggest that P-113Du and P-113Tri are promising candidates for development as novel antifungal agents.

In vitro activity of Aloe vera inner gel against microorganisms grown in planktonic and sessile phases.

The failure of traditional antimicrobial treatments is becoming a worldwide problem. The use of Aloe vera is of particular interest for its role as curative agent and its efficacy in complementary therapies for a variety of illnesses. This study evaluated the antimicrobial activity of A. vera inner gel against a panel of microorganisms, Gram-positive and -negative bacteria, and Candida albicans. In addition to A. vera inner gel being used in the treatment of peptic ulcers, in dermatological treatments, and wound healing, it was also tested on the sessile phase of clinical Helicobacter pylori strains (including multi-drug-resistant strains) and on planktonic and sessile phase of Staphylococcus aureus/Pseudomonas aeruginosa clinical isolates from venous leg ulcers.A. vera inner gel expresses its prevalent activity against Gram-negative bacteria and C. albicans in respect to Gram-positive bacteria. The results of the A. vera antibiofilm activity showed a decrease of the produced biomass in a concentration-dependent-way, in each analyzed microorganism. The data obtained show that A. vera inner gel has both an antimicrobial and antibiofilm activity suggesting its potential use for the treatment of microbial infections, in particular for H. pylori gastric infection, especially in case of multi-drug-resistance, as well as for an effective wound dressing.

Within-lake heterogeneity of environmental factors structuring bacterial community composition in Lake Dongting, China.

Within-lake heterogeneity of bacterial community composition (BCC) was investigated in the large and shallow regulating Lake Dongting. Samples were collected at 13 sites located in different areas of the lake. PCR-denaturing gradient gel electrophoresis (DGGE) and redundancy analysis (RDA) were used for revealing spatial distribution of BCC and the relationships between BCC and environmental variables. The DGGE banding patterns revealed a remarkable spatial heterogeneity which was closely related to their geographical positions. RDA result demonstrated that TP and TN, as well as Secchi depth, were the three most influential factors, responsible for a major part of the observed variation in BCC. Total bacterial abundances were significantly higher in Eastern Dongting due to high TP and suspended solids. In conclusion, bacterial community diversity in Lake Dongting was mainly shaped by within-lake heterogeneity of nutrient and transparency.

Heterogeneity in the nutrient limitation of different bacterioplankton groups in the Eastern Mediterranean Sea.

The heterotrophic bacterial community of the Eastern Mediterranean Sea is believed to be limited by phosphorus (P) availability. This observation assumes that all bacterial groups are equally limited, something that has not been hitherto examined. To test this hypothesis, we performed nutrient addition experiments and investigated the response of probe-identified groups using microautoradiography combined with catalyzed reporter deposition fluorescence in situ hybridization. Our results show contrasting responses between the bacterial groups, with Gammaproteobacteria being the group more affected by P availability. The Roseobacter clade was likely colimited by P and nitrogen (N), whereas Bacteroidetes by P, N and organic carbon (C). In contrast, SAR11 cells were active regardless of the nutrient concentration. These results indicate that there is high heterogeneity in the nutrient limitation of the different components of the bacterioplankton community.

Susceptibility of clinical isolates of Candida to photodynamic effects of curcumin.

BACKGROUND AND OBJECTIVE: The resistance of Candida species to antifungals represents a major challenge for therapeutic and prophylactic strategies. This study evaluated photodynamic therapy (PDT) mediated by Curcumin (CUR) against clinical isolates of C. albicans, C. tropicalis, and C. glabrata, both in planktonic and biofilm forms. STUDY DESIGN/MATERIALS AND METHODS: Suspensions of Candida were treated with three CUR concentrations and exposed to four LED fluences. The protocol that showed the best outcomes for inactivation of the planktonic phase was selected to be evaluated against Candida biofilms. In addition, two higher CUR concentrations were tested. The metabolic activity of biofilms was evaluated by means of XTT reduction assay and the biofilm biomass was evaluated using crystal violet (CV) staining assay. Data were analyzed in a mixed model nested ANOVA, Wilcoxon's nonparametric tests, and the Kruskal-Wallis test (α = 5%). RESULTS: The use of CUR in association with light was able to promote a significant antifungal effect against the planktonic form of the yeasts. When using 40 µM of CUR, the metabolic activity of C. albicans, C. glabrata, and C. tropicalis biofilms was reduced by 85%, 85%, and 73%, respectively, at 18 J/cm(2) . CUR-mediated PDT also decreased the biofilm biomass of all species evaluated. In addition, CV staining showed that C. albicans isolates were strong biofilm-forming strains, when compared with C. glabrata and C. tropicalis isolates. CONCLUSION: The results from the present investigation showed that low CUR concentrations can be highly effective for inactivating Candida isolates when associated with light excitation.

Dynamics of the hydrocarbon-degrading Cycloclasticus bacteria during mesocosm-simulated oil spills.

We used catalysed reported deposition - fluorescence in situ hybridization (CARD-FISH) to analyse changes in the abundance of the bacterial groups Alphaproteobacteria, Gammaproteobacteria and Bacteroidetes, and of hydrocarbon-degrading Cycloclasticus bacteria in mesocosms that had received polycyclic aromatic hydrocarbons (PAHs) additions. The effects of PAHs were assessed under four contrasting hydrographic conditions in the coastal upwelling system of the Rías Baixas: winter mixing, spring bloom, summer stratification and autumn upwelling. We used realistic additions of water soluble PAHs (approximately 20-30 microg l(-1) equivalent of chrysene), but during the winter period we also investigated the effect of higher PAHs concentrations (10-80 microg l(-1) chrysene) on the bacterial community using microcosms. The most significant change observed was a significant reduction (68 +/- 5%) in the relative abundance of Alphaproteobacteria. The magnitude of the response of Cycloclasticus bacteria (positive with probe CYPU829) to PAHs additions varied depending on the initial environmental conditions, and on the initial concentration of added PAHs. Our results clearly show that bacteria of the Cycloclasticus group play a major role in low molecular weight PAHs biodegradation in this planktonic ecosystem. Their response was stronger in colder waters, when their background abundance was also higher. During the warm periods, the response of Cycloclasticus was limited, possibly due to both, a lower bioavailability of PAHs caused by abiotic factors (solar radiation, temperature), and by inorganic nutrient limitation of bacterial growth.

Bacterial abundance, activity, and viability in the eutrophic River Warnow, northeast Germany.

The River Warnow is the drinking water source for the city of Rostock. Its eutrophic status is accompanied by high amounts of bacteria, which may reach up to 24 x 10(6) cells mL(-1) as recorded during a seasonal study in 2002. Because the river is eutrophic and also heavily loaded with organic matter, this burden is a problem for drinking water purification, as it must be removed completely to not trigger new bacterial growth in the pipeline network. Therefore, restoration measures in the river have to be planned, and bacteria have to be favored as decomposers. That includes the investigation of the physiological state of bacteria in situ. Viable and active cells in the lower reaches of River Warnow were estimated using a broad set of methods. Intact bacteria were investigated by the LIVE/DEAD BacLight bacterial viability kit, containing a mixture of permeant and impermeant nucleic acid stains. Cells with ribosomes were visualized by fluorescence in situ hybridization with the EUB338 oligonucleotide probe. Intact cells and ribosome-containing bacteria represented 24% of total numbers stained by 4'6,-diamidino-2-phenylindole (DAPI) or 66 and 62%, respectively, in relation to all bacteria visualized by the LIVE/DEAD kit. Both fractions were considered as viable, although the fraction of RIB + bacteria is most likely underestimated by the protocol applied. 5-Cyano-2,3-ditolyltetrazolium chloride (CTC) was applied to mark respiring bacteria. The esterase substrate CellTracker Green 5-chloromethylfluorescein diacetate showed cells with intracellular hydrolytic activity. Whereas 1.5% of DAPI-stained bacteria were observed as respiring, 3.8% exhibited intracellular hydrolytic activity on average. If these active fractions were calculated as the percentages of intact cells, much higher fractions of 5.4% were respiring and 16% hydrolytic. Temperature was a main factor influencing total and viable cell numbers simultaneously. The results confirm that there are different states of viable and active cells in natural bacterioplankton communities. However, it remains unclear why fractions of viable and active cells were rather low in this eutrophic river in comparison to similar waters. We recommend to carefully address cells as viable in contrast to nonviable, i.e., dead. As viable cells may be active or inactive with respect to many different activities, e.g., substrate uptake, respiration, hydrolysis, and cell deviation, it is necessary to choose the method to visualize active cells according to the question to be answered.

Comparative efficacies of quinupristin-dalfopristin, linezolid, vancomycin, and ciprofloxacin in treatment, using the antibiotic-lock technique, of experimental catheter-related infection due to Staphylococcus aureus.

We performed in vitro studies to elucidate the bactericidal activity of the antibiotics in an adherent-cell biofilm model. Efficacy studies were performed in a staphylococcal central venous catheter (CVC) infection rat model. Silastic catheters were implanted into the superior cava. Via the CVC the rats were challenged with 1.0 x 10(6) CFU of a live Staphylococcus aureus strain. Twenty-four hours later, the antibiotic-lock technique was started. All animals were randomized to receive daily isotonic sodium chloride solution, quinupristin-dalfopristin (Q/D), linezolid, vancomycin, or ciprofloxacin at the minimal bactericidal concentration (MBC) and at 1,024 microg/ml in a volume of 0.1 ml that filled the CVC. The main outcome measures were MICs and MBCs for both planktonic and adherent cells, quantitative culture of the catheters and surrounding venous tissues, and quantitative peripheral blood cultures. The killing activities of all antibiotics against the adherent bacteria were at least fourfold lower than those against freely growing cells, with the exception of Q/D, which showed comparable activities against both adherent and planktonic organisms. Overall, Q/D at 1,024 microg/ml produced the greatest reduction in the number of cells recovered from the catheters, while at the same concentration, Q/D and vancomycin demonstrated higher activities than ciprofloxacin or linezolid in reducing the number of organisms recovered from the blood cultures. This study points out that treatment outcome of device-related infections cannot be predicted by the results of a standard susceptibility test such as the MIC. Our findings suggest that the clinically used antibiotics cannot eradicate the CVC infection through the antibiotic-lock technique, even at a concentration of 1,024 microg/ml.

Planktonic nitrate-reducing bacteria and sulfate-reducing bacteria in some western Canadian oil field waters.

Oil fields that use water flooding to enhance oil recovery may become sour because of the production of H(2)S from the reduction of sulfate by sulfate-reducing bacteria (SRB). The addition of nitrate to produced waters can stimulate the activities of nitrate-reducing bacteria (NRB) and control sulfide production. Many previous studies have focused on chemolithotrophic bacteria that can use thiosulfate or sulfide as energy sources while reducing nitrate. Little attention has been given to heterotrophic NRB in oil field waters. Three different media were used in this study to enumerate various types of planktonic NRB present in waters from five oil fields in western Canada. The numbers of planktonic SRB and bacteria capable of growth under aerobic conditions were also determined. In general, microbial numbers in the produced waters were very low (<10 ml x (-1)) in samples taken near or at wellheads. However, the numbers increased in the aboveground facilities. No thiosulfate-oxidizing NRB were detected in the oil field waters, but other types of NRB were detected in 16 of 18 produced water samples. The numbers of heterotrophic NRB were equal to or greater than the number of sulfide-oxidizing, chemolithotrophic NRB in 12 of 15 samples. These results showed that each of the oil fields contained NRB, which might be stimulated by nitrate amendment to control H(2)S production by SRB.