The Phenolic Contents and Antimicrobial Activities of Some Marine Algae from the Mediterranean Sea (Algeria)

In this study, three marine algae collected from western coast of algerian mediteranean sea (Ulva lactuca, Dictyota dichotoma, and Corallina elongata) were tested using the agar-well diffusion method for their production of antibacterial and antifungal agents on various organisms that cause diseases of humans and plants (Eschirichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Salmonella sp, Candida albicans, and Penicillium sp.). The total phenol content and antimicrobial activity were determined using different crude seaweeds extracts (methanol, diethylether, and chloroform). The results show that the chloroform extracts of (Ulva lactuca and Corallina elongata) had the highest activity against E. coli and Salmonella sp. The methanol extract obtained from (Ulva lactuca, Dictyota dichotoma, and Corallina elongata) showed antifungal activity for Candida albicans. The results of the study revealed that the seaweeds from Algeria appear to have immense potential as a source of antibacterial and antifungal compounds; they can be used in treating diseases caused by these organisms.     

Growth of Ag-nanoparticles in an aqueous solution and their antimicrobial activities against Gram positive,Gram negative bacterial strains and Candida fungus

Silver nanoparticles (AgNPs) were synthesized using Ocimum sanctum (Tulsi) leaves aqueous extract as reducing as well as a capping agent in absence and presence of cetyltrimethylammonium bromide (CTAB). The resulting nanomaterials were characterized by UV–visible spectrophotometer, and transmission electron microscope. The UV–Vis spectroscopy revealed the formation of AgNPs at 400–450 nm. TEM photographs indicate that the truncated triangular silver nanoplates and/or spherical morphology of the AgNPs with an average diameter of 25 nm have been distorted markedly in presence of CTAB. The AgNPs were almost mono disperse in nature. Antimicrobial activities of AgNPs were determined by using two bacteria (Gram positive Staphylococcus aureus MTCC-3160), Gram negative Escherichia coli MTCC-450) and one species of Candida fungus (Candida albicans ATCC 90030) with Kirby-Bauer or disc diffusion method. The zone of inhibition seems extremely good showing a relatively large zone of inhibition in both Staphylococcus aureus, Escherichia coli, and Candida albicans strains.     

Encapsulation in Polymeric Microparticles Improves Daptomycin Activity Against Mature Staphylococci Biofilms—a Thermal and Imaging Study

Eradication of Gram-positive biofilms is a critical aspect in implant-associated infection treatment. Although antibiotic-containing particulate carriers may be a promising strategy for overcoming biofilm tolerance, the assessment of their interaction with biofilms has not been fully explored. In the present work, the antibiofilm activity of daptomycin- and vancomycin-loaded poly(methyl methacrylate) (PMMA) and PMMA-Eudragit RL 100 (EUD) microparticles against methicillin-resistant Staphylococcus aureus (MRSA) and polysaccharide intercellular adhesin-positive S. epidermidis biofilms was investigated using isothermal microcalorimetry (IMC) and fluorescence in situ hybridization (FISH). The minimal biofilm inhibitory concentrations (MBIC) of MRSA biofilms, as determined by IMC, were 5 and 20 mg/mL for daptomycin- and vancomycin-loaded PMMA microparticles, respectively. S. epidermidis biofilms were less susceptible, with a MBIC of 20 mg/mL for daptomycin-loaded PMMA microparticles. Vancomycin-loaded microparticles were ineffective. Adding EUD to the formulation caused a 4- and 16-fold reduction of the MBIC values of daptomycin-loaded microparticles for S. aureus and S. epidermidis, respectively. FISH corroborated the IMC results and provided additional insights on the antibiofilm effect of these particles. According to microscopic analysis, only daptomycin-loaded PMMA-EUD microparticles were causing a pronounced reduction in biofilm mass for both strains. Taken together, although IMC indicated that a biofilm inhibition was achieved, microscopy showed that the biofilm was not eradicated and still contained FISH-positive, presumably viable bacteria, thus indicating that combining the two techniques is essential to fully assess the effect of microparticles on staphylococcal biofilms.     

Isolation Purification and Characterization of Antimicrobial Peptides from <Emphasis Type="Italic">Cuminum cyminum</Emphasis> L. Seeds

In this work, antimicrobial peptides from Cuminum cyminum L. seeds were isolated and purified for the first time by 50% ethanol extraction, C18 reverse phase column chromatography and ion exchange chromatography for separation different peptides fraction. Then isolated fractions were characterized by Gel electrophoresis (SDS-PAGE), high-pressure liquid chromatography and the peptides components and molecular weights were determined by liquid chromatography and mass spectrometry. The extracts were tested against some strains of bacteria (E. coli and Staphylococcus aureus) and one strain of fungi (Candida albicans) using well diffusion and broth dilution assays. The extracts from C. cyminum L. seeds demonstrated a high degree of activity (some antibacterial effect) against the bacteria strains and аntifungal activity against the Candida albicans. However, the study indicates that the crude peptide extracts from C. cyminum L. seeds have promising antimicrobial and antioxidant activities that can be harnessed as leads for potential bioactive compounds.     

The Mycotoxin Zearalenone Hinders <Emphasis Type="Italic">Candida albicans</Emphasis> Biofilm Formation and Hyphal Morphogenesis

Yeast–mold mycobiota inhabit several natural ecosystems, in which symbiotic relationships drive strategic pathoadaptation. Mycotoxins are metabolites produced by diverse mycotoxigenic fungi as a defense against yeasts, though at times yeasts secrete enzymes that degrade, detoxify, or bio-transform mycotoxins. The present study is focused on the in vitro inhibitory effects of zearalenone (ZEN), a F2 mycotoxin produced by several Fusarium and Gibberella species, on different microbial strains. ZEN exhibited no effect on the planktonic growth or biofilms of several Gram positive and negative bacteria at the tested concentrations. Remarkably, Candida albicans biofilm formation and hyphal morphogenesis were significantly inhibited when treated with 100 µg/mL of ZEN. Likewise, ZEN proficiently disrupted pre-formed C. albicans biofilms without disturbing planktonic cells. Furthermore, these inhibitions were confirmed by crystal violet staining and XTT reduction assays and by confocal and scanning electron microscopy. In an in vivo model, ZEN significantly suppressed C. albicans infection in the nematode Caenorhabditis elegans. The study reports the in vitro antibiofilm efficacy of ZEN against C. albicans strains, and suggests mycotoxigenic fungi participate in asymmetric competitive interactions, such as, amensalism or antibiosis, rather than commensal interactions with C. albicans, whereby mycotoxins secreted by fungi destroy C. albicans biofilms.     

Coating polypropylene surfaces with protease weakens the adhesion and increases the dispersion of <Emphasis Type="Italic">Candida albicans</Emphasis> cells

Objectives

To investigate the ability of the proteases, subtilisin and α-chymotrypsin (aCT), to inhibit the adhesion of Candida albicans biofilm to a polypropylene surface.

Results

The proteases were immobilized on plasma-treated polypropylene by covalently linking them with either glutaraldehyde (GA) or N′-diisopropylcarbodiimide (DIC) and N-hydroxysuccinimide (NHS). The immobilization did not negatively affect the enzyme activity and in the case of subtilisin, the activity was up to 640% higher than that of the free enzyme when using N-acetyl phenylalanine ethyl ester as the substrate. The efficacies against biofilm dispersal for the GA-linked SubC and aCT coatings were 41 and 55% higher than the control (polypropylene coated with only GA), respectively, whereas no effect was observed with enzymes immobilized with DIC and NHS. The higher dispersion efficacy observed for the proteases immobilized with GA could be both steric (proper orientation of the active site) and dynamic (higher protein mobility/flexibility).

Conclusions

Proteases immobilized on a polypropylene surface reduced the adhesion of C. albicans biofilms and therefore may be useful in developing anti-biofilm surfaces based on non-toxic molecules and sustainable strategies.

     

Usefulness of the Non-conventional <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis> Model to Assess <Emphasis Type="Italic">Candida</Emphasis> Virulence

Invasive candidiasis is caused mainly by Candida albicans, but other Candida species have increasing etiologies. These species show different virulence and susceptibility levels to antifungal drugs. The aims of this study were to evaluate the usefulness of the non-conventional model Caenorhabditis elegans to assess the in vivo virulence of seven different Candida species and to compare the virulence in vivo with the in vitro production of proteinases and phospholipases, hemolytic activity and biofilm development capacity. One culture collection strain of each of seven Candida species (C. albicans, Candida dubliniensis, Candida glabrata, Candida krusei, Candida metapsilosis, Candida orthopsilosis and Candida parapsilosis) was studied. A double mutant C. elegans AU37 strain (glp-4;sek-1) was infected with Candida by ingestion, and the analysis of nematode survival was performed in liquid medium every 24 h until 120 h. Candida establishes a persistent lethal infection in the C. elegans intestinal tract. C. albicans and C. krusei were the most pathogenic species, whereas C. dubliniensis infection showed the lowest mortality. C. albicans was the only species with phospholipase activity, was the greatest producer of aspartyl proteinase and had a higher hemolytic activity. C. albicans and C. krusei caused higher mortality than the rest of the Candida species studied in the C. elegans model of candidiasis.     

The antibacterial agent,moxifloxacin inhibits virulence factors of <Emphasis Type="Italic">Candida albicans</Emphasis> through multitargeting

Fluoroquinolines are broad spectrum fourth generation antibiotics. Some of the Fluoroquinolines exhibit antifungal activity. We are reporting the potential mechanism of action of a fluoroquinoline antibiotic, moxifloxacin on the growth, morphogenesis and biofilm formation of the human pathogen Candida albicans. Moxifloxacin was found to be Candidacidal in nature. Moxifloxacin seems to inhibit the yeast to Hyphal morphogenesis by affecting signaling pathways. It arrested the cell cycle of C. albicans at S phase. Docking of moxifloxacin with predicted structure of C. albicans DNA Topoisomerase II suggests that moxifloxacin may bind and inhibit the activity of DNA Topoisomerase II in C. albicans. Moxifloxacin could be used as a dual purpose antibiotic for treating mixed infections caused by bacteria as well as C. albicans. In addition chances of developing moxifloxacin resistance in C. albicans are less considering the fact that moxifloxacin may target multiple steps in yeast to hyphal transition in C. albicans.     

Tolerance towards gentamicin is a function of nutrient concentration in biofilms of patient-isolated <Emphasis Type="Italic">Staphylococcus epidermidis</Emphasis>

Staphylococcus epidermidis is a biofilm-forming bacterial strain that can cause major problems as an agent of nosocomial infections. Bacteria in biofilms are shielded from the environment and can survive high doses of antibiotics. We here test the antibiotic susceptibility of Staphylococcus epidermidis to rising gentamicin concentrations in optimal growth conditions as used in routine bacteriology laboratories with low nutrient situations as suggested to be found in clinical situations. We found that gentamicin-resistant Staphylococcus epidermidis biofilms survived in the absence of external nutrient supply in PBS. While addition of gentamicin sulfate significantly reduced the pH value of all used media and solutions, this acidification did not alter survival of bacteria in the biofilm. We found a statistically significant and dose-dependent reduction of survival in low nutrient situations using gentamicin sulfate in three out of four patient isolates of Staphylococcus epidermidis which have been tested to be gentamicin-resistant under optimal growth conditions. Supporting the original profiling, survival in full media under the same antibiotic dosages was not significantly reduced. Our data here show that antibiotic resistance is a function of the provided nutrient concentration. Antibiotic resistance profiling should consider variations in nutrient availability.     

Virulence Factors in <Emphasis Type="Italic">Candida albicans</Emphasis> and <Emphasis Type="Italic">Streptococcus mutans</Emphasis> Biofilms Mediated by Farnesol

The aim of this study was to evaluate the effect of farnesol on the production of acids and hydrolytic enzymes by biofilms of Streptococcus mutans and Candida albicans. The present study also evaluated the time-kill curve and the effect of farnesol on matrix composition and structure of single-species and dual-species biofilms. Farnesol, at subinhibitory concentrations, showed a significant reduction in S. mutans biofilm acid production, but did not alter C. albicans hydrolytic enzyme production. The number of cultivable cells of both microorganisms was significantly reduced after 8 h of contact with farnesol. Extracellular matrix protein content was reduced for biofilms formed in the presence of farnesol. In addition, confocal laser scanning and scanning electron microscopy displayed structural alterations in all biofilms treated with farnesol, which included reduction in viable cells and extracellular matrix. In conclusion, farnesol showed favorable properties controlling some virulence factors of S. mutans and C. albicans biofilms. These findings should stimulate further studies using this quorum-sensing molecule, combined with other drugs, to prevent or treat biofilm-associated oral diseases.     

In vitro activity of vancomycin, quinupristin/dalfopristin, and linezolid against intact and disrupted biofilms of staphylococci

 

Shed cells or disrupted parts of the biofilm may enter the circulation causing serious and very hard to treat biofilm-associated infections. The activity of antimicrobial agents against the shed cells/disrupted biofilms is largely unknown.

Methods

We studied the in vitro susceptibility of intact and disrupted biofilms of thirty clinical isolates of methicillin-resistant and methicillin–susceptible Staphylococcus aureus (MRSA and MSSA) and Staphylococcus epidermidis to vancomycin, quinupristin/dalfopristin, and linezolid and compared it to that of the suspended (planktonic) cells.

Results

Bacteria in the disrupted biofilms were as resistant as those in the intact biofilms at the minimum inhibitory concentrations of the antibiotics. At higher concentrations, bacteria in the disrupted biofilms were significantly (P < 0.001) less resistant than those in the intact biofilms but more resistant than the planktonic cells. Quinupristin/dalfopristin showed the best activity against cells of the disrupted biofilms at concentrations above MICs and vancomycin, at 500 and 1,000 μg/ml, was significantly more active against the biofilms of MRSA and S. epidermidis

Conclusion

The difficulty of treating biofilm-associated infections may be attributed not only to the difficulty of eradicating the biofilm focus but also to the lack of susceptibility of cells disrupted from the biofilm to antimicrobial agents.     

Identification and Characterization of Novel Antimicrobial Peptide from <Emphasis Type="Italic">Hippocampus comes</Emphasis> by In Silico and Experimental Studies

Antimicrobial peptides (AMPs) have attracted attentions as a novel antimicrobial agent because of their unique activity against microbes. In the present study, we described a new, previously unreported AMP, moronecidin-like peptide, from Hippocampus comes and compared its antimicrobial activity with moronecidin from hybrid striped bass. Antibacterial assay indicated that gram-positive bacteria were more sensitive to moronecidin and moronecidin-like compared with gram-negative bacteria. Furthermore, both AMPs were found to exhibit effective antifungal activity. Comparative analysis of the antimicrobial activity revealed that moronecidin-like peptide has higher activity against Acinetobacter baumannii and Staphylococcus epidermidis relative to moronecidin. Both moronecidin-like and moronecidin peptides retained their antibacterial activity in physiological pH and salt concentration. The time-killing assay showed that the AMPs completely killed A. baumannii and S. epidermidis isolates after 1 and 5 h at five- and tenfold above their corresponding MICs, respectively. Anti-biofilm assay demonstrated that peptides were able to inhibit 50% of biofilm formation at sub-MIC of 1/8 MIC. Furthermore, moronecidin-like significantly inhibited biofilm formation more than moronecidin at 1/16 MIC. Collectively, our results revealed that antimicrobial and anti-biofilm activities of moronecidin-like are comparable to moronecidin. In addition, the hemolytic and cytotoxic activities of moronecidin-like were lower than those of moronecidin, suggesting it as a potential novel therapeutic agent, and a template to design new therapeutic AMPs.     

The <Emphasis Type="Italic">in vitro</Emphasis> and <Emphasis Type="Italic">in vivo</Emphasis> efficacy of fluconazole in combination with farnesol against <Emphasis Type="Italic">Candida albicans</Emphasis> isolates using a murine vulvovaginitis model

Farnesol is a quorum-sensing molecule that inhibits biofilm formation in Candida albicans. Previous in vitro data suggest that, in combination with certain antifungals, farnesol may have an adjuvant anti-biofilm agent. However, the in vivo efficacy of farnesol is very questionable. Therefore, the in vitro and in vivo activity of fluconazole combined with farnesol was evaluated against C. albicans biofilms using fractional inhibitory concentration index (FICI) determination, time-kill experiments and a murine vulvovaginitis model. The median biofilm MICs of fluconazole-sensitive C. albicans isolates ranged between 4 -> 512 mg/L and 150–300 μM for fluconazole and farnesol, respectively. These values were 512 -> 512 mg/L and > 300 μM for fluconazole-resistant clinical isolates. Farnesol decreased the median MICs of fluconazole by 2-64-fold for biofilms. Based on FICI, synergistic interaction was observed only in the case of the sessile SC5314 reference strain (FICIs: 0.16–0.27). In time-kill studies, only the 512 mg/L fluconazole and 512 mg/L fluconazole + 75 μM farnesol reduced biofilm mass significantly at each time point in the case of all isolates. The combination reduced the metabolic activity of biofilms for all isolates in a concentration- and time-dependent manner. Our findings revealed that farnesol alone was not protective in a murine vulvovaginitis model. Farnesol was not beneficial in combination with fluconazole for fluconazole-susceptible isolates, but partially increased fluconazole activity against one fluconazole-resistant isolate, but not the other one.     

<Emphasis Type="Italic">Lactobacillus plantarum</Emphasis> as a Probiotic Potential from Kouzeh Cheese (Traditional Iranian Cheese) and Its Antimicrobial Activity

The aim of this study is to isolate and identify Lactobacillus plantarum isolates from traditional cheese, Kouzeh, and evaluate their antimicrobial activity against some food pathogens. In total, 56 lactic acid bacteria were isolated by morphological and biochemical methods, 12 of which were identified as Lactobacillus plantarum by biochemical method and 11 were confirmed by molecular method. For analyzing the antimicrobial activity of these isolates properly, diffusion method was performed. The isolates were identified by 318 bp band dedicated for L. plantarum. The isolated L. plantarum represented an inhibitory activity against four of the pathogenic bacteria and showed different inhibition halos against each other. The larger halos were observed against Staphylococcus aureus and Staphylococcus epidermidis (15 ± 0.3 and 14.8 ± 0.7 mm, respectively). The inhibition halo of Escherichia coli was smaller than that of other pathogen and some L. plantarum did not show any inhibitory activity against E. coli, which were resistant to antimicrobial compounds produced by L. plantarum. The isolated L. plantarum isolates with the antimicrobial activity in this study had strong probiotic properties. These results indicated the nutritional value of Kouzeh cheese and usage of the isolated isolates as probiotic strains.     

Synthetic Antimicrobial Peptides: I. Antimicrobial Activity of Amphiphilic and Nonamphiphilic Cationic Peptides

Comparative antimicrobial properties of three artificial cationic synthetic antimicrobial peptides (SAMP): (RAhaR)₄AhaβA (where R is Arg, Aha is 6-aminohexanoic acid, βA is beta-alanine), (KFF)₃K and R₉F₂ with various amphiphilic properties have been studied relative to pathogenic strains of microorganisms: Gram-negative bacteria Pseudomonas aeruginosa, Escherichia coli, Proteus mirabilis, and Salmonella enterica, Gram-positive bacteria Staphylococcus aureus, and pathogenic yeast fungus Candida albicans. The selectivity index (SI) values of the peptide preparations were calculated as the ratio of the 50% cytotoxic concentration (TC₅₀) towards eukaryotic host cells to the MIC₅₀ values of the testing antimicrobial peptides. The studied SAMPs appeared to be the most active against the pathogenic yeast fungus C. albicans and the bacterial strains St. aureus and P. aeruginosa. The SI values in these cases exceed 40. Some assumed molecular interactions of the studied SAMPs on the microbial cells have been considered, and possible pathways to increase their antimicrobial activity have been suggested. The proposed SAMPs can serve as a basis for the design and synthesis of new promising synthetic antimicrobial agents.     

Large-scale biofilm cultivation of Antarctic bacterium <Emphasis Type="Italic">Pseudoalteromonas haloplanktis</Emphasis> TAC125 for physiologic studies and drug discovery

Microbial biofilms are mainly studied due to detrimental effects on human health but they are also well established in industrial biotechnology for the production of chemicals. Moreover, biofilm can be considered as a source of novel drugs since the conditions prevailing within biofilm can allow the production of specific metabolites. Antarctic bacterium Pseudoalteromonas haloplanktis TAC125 when grown in biofilm condition produces an anti-biofilm molecule able to inhibit the biofilm of the opportunistic pathogen Staphylococcus epidermidis. In this paper we set up a P. haloplanktis TAC125 biofilm cultivation methodology in automatic bioreactor. The biofilm cultivation was designated to obtain two goals: (1) the scale up of cell-free supernatant production in an amount necessary for the anti-biofilm molecule/s purification; (2) the recovery of P. haloplanktis TAC125 cells grown in biofilm for physiological studies. We set up a fluidized-bed reactor fermentation in which floating polystyrene supports were homogeneously mixed, exposing an optimal air–liquid interface to let bacterium biofilm formation. The proposed methodology allowed a large-scale production of anti-biofilm molecule and paved the way to study differences between P. haloplanktis TAC125 cells grown in biofilm and in planktonic conditions. In particular, the modifications occurring in the lipopolysaccharide of cells grown in biofilm were investigated.     

Synthesis,characterization and evaluation of antimicrobial and cytotoxic activities of biogenic silver nanoparticles synthesized from <Emphasis Type="Italic">Streptomyces xinghaiensis</Emphasis> OF1 strain

We report synthesis of silver nanoparticles (AgNPs) from Streptomyces xinghaiensis OF1 strain, which were characterised by UV–Vis and Fourier transform infrared spectroscopy, Zeta sizer, Nano tracking analyser, and Transmission electron microscopy. The antimicrobial activity of AgNPs alone, and in combination with antibiotics was evaluated against bacteria, namely Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis, and yeasts viz., Candida albicans and Malassezia furfur by using micro-dilution method. The minimum inhibitory concentration (MIC) and minimum biocidal concentration of AgNPs against bacterial and yeast strains were determined. Synergistic effect of AgNPs in combination with antibacterial and antifungal antibiotics was determined by FIC index. In addition, MTT assay was performed to study cytotoxicity of AgNPs alone and in combination with antibiotics against mouse fibroblasts and HeLa cell line. Biogenic AgNPs were stable, spherical, small, polydispersed and capped with organic compounds. The variable antimicrobial activity of AgNPs was observed against tested bacteria and yeasts. The lowest MIC (16 µg ml^?1) of AgNPs was found against P. aeruginosa, followed by C. albicans and M. furfur (both 32 µg ml^?1), B. subtilis and E. coli (both 64 µg ml^?1), and then S. aureus and Klebsiella pneumoniae (256 µg ml^?1). The high synergistic effect of antibiotics in combination with AgNPs against tested strains was found. The in vitro cytotoxicity of AgNPs against mouse fibroblasts and cancer HeLa cell lines revealed a dose dependent potential. The IC₅₀ value of AgNPs was found in concentrations of 4 and 3.8 µg ml^?1, respectively. Combination of AgNPs and antibiotics significantly decreased concentrations of both antimicrobials used and retained their high antibacterial and antifungal activity. The synthesis of AgNPs using S. xinghaiensis OF1 strain is an eco-friendly, cheap and nontoxic method. The antimicrobial activity of AgNPs could result from their small size. Remarkable synergistic effect of antibiotics and AgNPs offer their valuable potential in nanomedicine for clinical application as a combined therapy in the future.     

A novel targeted/untargeted GC-Orbitrap metabolomics methodology applied to <Emphasis Type="Italic">Candida albicans</Emphasis> and <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> biofilms

Introduction

Combined infections from Candida albicans and Staphylococcus aureus are a leading cause of death in the developed world. Evidence suggests that Candida enhances the virulence of Staphylococcus—hyphae penetrate through tissue barriers, while S. aureus tightly associates with the hyphae to obtain entry to the host organism. Indeed, in a biofilm state, C. albicans enhances the antimicrobial resistance characteristics of S. aureus. The association of these microorganisms is also associated with significantly increased morbidity and mortality. Due to this tight association we hypothesised that metabolic effects were also in evidence.

Objectives

To explore the interaction, we used a novel GC-Orbitrap-based mass spectrometer, the Q Exactive GC, which combines the high peak capacity and chromatographic resolution of gas chromatography with the sub-ppm mass accuracy of an Orbitrap system. This allows the capability to leverage the widely available electron ionisation libraries for untargeted applications, along with expanding accurate mass libraries and targeted matches based around authentic standards.

Methods

Optimised C. albicans and S. aureus mono- and co-cultured biofilms were analysed using the new instrument in addition to the fresh and spent bacterial growth media.

Results

The targeted analysis experiment was based around 36 sugars and sugar phosphates, 22 amino acids and five organic acids. Untargeted analysis resulted in the detection of 465 features from fresh and spent medium and 405 from biofilm samples. Three significantly changing compounds that matched to high scoring library fragment patterns were chosen for validation.

Conclusion

Evaluation of the results demonstrates that the Q Exactive GC is suitable for metabolomics analysis using a targeted/untargeted methodology. Many of the results were as expected: e.g. rapid consumption of glucose and fructose from the medium regardless of the cell type. Modulation of sugar-phosphate levels also suggest that the pentose phosphate pathway could be enhanced in the cells from co-cultured biofilms. Untargeted metabolomics results suggested significant production of cell-wall biosynthesis components and the consumption of non-proteinaceous amino-acids.