Carbamoylase-based impedimetric electronic tongue for rapid detection of paralytic shellfish toxins.

Phytotoxins produced by marine microalgae, such as paralytic shellfish toxins (PSTs), can accumulate in bivalve molluscs, representing a human health concern due to the life-threatening symptoms they cause. To avoid the commercialization of contaminated bivalves, monitoring programs were established in the EU. The purpose of this work is the implementation of a PST transforming enzyme-carbamoylase-in an impedimetric test for rapid simultaneous detection of several carbamate and N-sulfocarbamoyl PSTs. Carbamoylase hydrolyses carbamate and sulfocarbamoyl toxins, which may account for up to 90% of bivalve toxicity related to PSTs. Conformational changes of carbamoylase accompanying enzymatic reactions were probed by Fourier transform mid-infrared spectroscopy (FT-MIR) and electrochemical impedance spectroscopy (EIS). Furthermore, a combination of EIS with a metal electrode and a carbamoylase-based assay was employed to harness changes in the enzyme conformation and adsorption on the electrode surface during the enzymatic reaction as an analytical signal. After optimization of the working conditions, the developed impedimetric e-tongue could quantify N-sulfocarbamoyl toxins with a detection limit of 0.1 µM. The developed e-tongue allows the detection of these toxins at concentration levels observed in bivalves with PST toxicity close to the regulatory limit. The quantification of a sum of N-sulfocarbamoyl PSTs in naturally contaminated mussel extracts using the developed impedimetric e-tongue has been demonstrated.

Microplastics in marine mussels, biological effects and human risk of intake: A case study in a multi-stressor environment.

This study documented seasonal levels of microplastics (MPs) and biomarkers (condition index, neurotoxicity, energy, oxidative stress) in mussels (Mytilus galloprovincialis), and water physico-chemical parameters in the Douro estuary (NE Atlantic coast), and estimated the human risk of MP intake (HRI) through mussels. Mussel stress was determined through the Integrated Biomarker Response (IBR). HRI was estimated from mussel MP concentrations and consumer habits. MPs were mainly micro-fibres (72 %) with varied chemical composition. Seasonal MP means (±SEM) in mussels ranged from 0.111 ± 0.044 (spring) to 0.312 ± 0.092 MPs/g (summer). Seasonal variations of mussel stress (IBR: 1.4 spring to 9.7 summer) and MP concentrations were not related. MeO-BDEs, PBDEs, temperature, salinity and other factors likely contributed to mussel stress variation. HRI ranged from 2438 to 2650 MPs/year. Compared to the literature, MP contamination in mussels is low, as well as the human risk of MP intake through their consumption.

Towards the Understanding of the Function of Lanthipeptide and TOMM-Related Genes in Haloferax mediterranei.

Research on secondary metabolites produced by Archaea such as ribosomally synthesized and post-translationally modified peptides (RiPPs) is limited. The genome of Haloferax mediterranei ATCC 33500 encodes lanthipeptide synthetases (medM1, medM2, and medM3) and a thiazole-forming cyclodehydratase (ycaO), possibly involved in the biosynthesis of lanthipeptides and the TOMMs haloazolisins, respectively. Lanthipeptides and TOMMs often have antimicrobial activity, and H. mediterranei has antagonistic activity towards haloarchaea shown to be independent of medM genes. This study investigated (i) the transcription of ycaO and medM genes, (ii) the involvement of YcaO in bioactivity, and (iii) the impact of YcaO and MedM-encoding genes' absence in the biomolecular profile of H. mediterranei. The assays were performed with biomass grown in agar and included RT-qPCR, the generation of knockout mutants, bioassays, and FTIR analysis. Results suggest that ycaO and medM genes are transcriptionally active, with the highest number of transcripts observed for medM2. The deletion of ycaO gene had no effect on H. mediterranei antihaloarchaea activity. FTIR analysis of medM and ycaO knockout mutants suggest that MedMs and YcaO activity might be directly or indirectly related t lipids, a novel perspective that deserves further investigation.

Major characteristics of microplastics in mussels from the Portuguese coast.

The present study reports the quantity, shape, colour and chemical properties of microplastics (MP) and MP-like in whole soft tissues of the mussel Mytilus spp. collected in January and February 2019 from four natural banks in the Portuguese coast. Three sites are located in estuarine areas influenced by anthropogenic pressures and freshwater discharges, and one in the coast far from urbanised areas. An alkaline digestion (KOH) of biological tissues was used and a polymeric identification of 20% of the visually sorted particles was achieved using the Fourier-transform mid-infrared spectroscopy (FT-MIR). MP and MP-like concentrations ranged from 0.54 to 3.0 items g-1 without significant differences among the sites. Particle size varied from 36 to 4439 µm, being fibers the most abundant shape (50%) followed by films (22%) and spherules (18%). FT-MIR revealed that 69% of the analysed particles were plastic, being identified six polymers and two polymeric blends, and 32% were cellulose-based materials. Fibers identified in mussel tissues were mainly composed of cotton and viscose (77%). This study emphasizes the importance of the polymer's spectroscopic identification after microscopic observation to recognise MP.

Antibacterial Multi-Layered Nanocellulose-Based Patches Loaded with Dexpanthenol for Wound Healing Applications.

Antibacterial multi-layered patches composed of an oxidized bacterial cellulose (OBC) membrane loaded with dexpanthenol (DEX) and coated with several chitosan (CH) and alginate (ALG) layers were fabricated by spin-assisted layer-by-layer (LbL) assembly. Four patches with a distinct number of layers (5, 11, 17, and 21) were prepared. These nanostructured multi-layered patches reveal a thermal stability up to 200 °C, high mechanical performance (Young's modulus ≥ 4 GPa), and good moisture-uptake capacity (240-250%). Moreover, they inhibited the growth of the skin pathogen Staphylococcus aureus (3.2-log CFU mL-1 reduction) and were non-cytotoxic to human keratinocytes (HaCaT cells). The in vitro release profile of DEX was prolonged with the increasing number of layers, and the time-dependent data imply a diffusion/swelling-controlled drug release mechanism. In addition, the in vitro wound healing assay demonstrated a good cell migration capacity, headed to a complete gap closure after 24 h. These results certify the potential of these multi-layered polysaccharides-based patches toward their application in wound healing.

Multifunctional nanofibrous patches composed of nanocellulose and lysozyme nanofibers for cutaneous wound healing.

Cutaneous wounds frequently require the use of patches to promote healing, nevertheless, most commercial products are fabricated with non-biodegradable synthetic substrates that pose environmental problems upon disposal. Herein, the partnership between two biobased nanofibrous polymers, namely a polysaccharide (nanofibrillated cellulose (NFC)) and a protein (lysozyme nanofibers (LNFs)), is explored to design sustainable fibrous patches with good mechanical performance and biological functionalities for wound healing applications. Two patches with different morphologies were prepared by vacuum filtration of a water-based suspension of both nanofibers and by sequential filtration of the separated suspensions (layered patch). The resultant freestanding patches exhibited high thermal stability (up to 250 °C), mechanical performance (Young's modulus ≥3.7 GPa), and UV-barrier properties. The combination of the bioactive LNFs with the mechanically robust NFC conveyed antioxidant activity (76-79% DPPH scavenging) and antimicrobial activity against Staphylococcus aureus (3.5-log CFU mL-1 reduction), which is a major benefit to prevent microbial wound infections. Moreover, these patches are biocompatible towards L929 fibroblast cells, and the in vitro wound healing assay evidenced a good migration capacity leading to an almost complete wound occlusion. Therefore, the partnership between the two naturally derived nanofibrous polymers represents a potential blueprint to engineer sustainable multifunctional patches for cutaneous wound healing.

The Health-Promoting Potential of Salix spp. Bark Polar Extracts: Key Insights on Phenolic Composition and In Vitro Bioactivity and Biocompatibility.

Salix spp. have been exploited for energy generation, along with folk medicine use of bark extracts for antipyretic and analgesic benefits. Bark phenolic components, rather than salicin, have demonstrated interesting bioactivities, which may ensure the sustainable bioprospection of Salix bark. Therefore, this study highlights the detailed phenolic characterization, as well as the in vitro antioxidant, anti-hypertensive, Staphylococcus aureus growth inhibitory effects, and biocompatibility of Salix atrocinerea Brot., Salix fragilis L., and Salix viminalis L. bark polar extracts. Fifteen phenolic compounds were characterized by ultra-high-performance liquid chromatography-ultraviolet detection-mass spectrometry analysis, from which two flavan-3-ols, an acetophenone, five flavanones, and a flavonol were detected, for the first time, as their bark components. Salix bark extracts demonstrated strong free radical scavenging activity (5.58-23.62 µg mL-1 IC50 range), effective inhibition on angiotensin-I converting enzyme (58-84%), and S. aureus bactericidal action at 1250-2500 µg mL-1 (6-8 log CFU mL-1 reduction range). All tested Salix bark extracts did not show cytotoxic potential against Caco-2 cells, as well as S. atrocinerea Brot. and S. fragilis L. extracts at 625 and 1250 µg mL-1 against HaCaT and L929 cells. These valuable findings can pave innovative and safer food, nutraceutical, and/or cosmetic applications of Salix bark phenolic-containing fractions.

Antimicrobial and Conductive Nanocellulose-Based Films for Active and Intelligent Food Packaging.

Bacterial nanocellulose (BNC) is becoming an important substrate for engineering multifunctional nanomaterials with singular and tunable properties for application in several domains. Here, antimicrobial conductive nanocomposites composed of poly(sulfobetaine methacrylate) (PSBMA) and BNC were fabricated as freestanding films for application in food packaging. The nanocomposite films were prepared through the one-pot polymerization of sulfobetaine methacrylate (SBMA) inside the BNC nanofibrous network and in the presence of poly(ethylene glycol) diacrylate as cross-linking agent. The ensuing films are macroscopically homogeneous, more transparent than pristine BNC, and present thermal stability up to 265 °C in a nitrogen atmosphere. Furthermore, the films have good mechanical performance (Young's modulus ≥ 3.1 GPa), high water-uptake capacity (450-559%) and UV-blocking properties. The zwitterion film with 62 wt.% cross-linked PSBMA showed bactericidal activity against Staphylococcus aureus (4.3-log CFU mL-1 reduction) and Escherichia coli (1.1-log CFU mL-1 reduction), and proton conductivity ranging between 1.5 × 10-4 mS cm-1 (40 °C, 60% relative humidity (RH)) and 1.5 mS cm-1 (94 °C, 98% RH). Considering the current set of properties, PSBMA/BNC nanocomposites disclose potential as films for active food packaging, due to their UV-barrier properties, moisture scavenging ability, and antimicrobial activity towards pathogenic microorganisms responsible for food spoilage and foodborne illness; and also for intelligent food packaging, due to the proton motion relevant for protonic-conduction humidity sensors that monitor food humidity levels.

Zwitterionic Nanocellulose-Based Membranes for Organic Dye Removal.

The development of efficient and environmentally-friendly nanomaterials to remove contaminants and pollutants (including harmful organic dyes) ravaging water sources is of major importance. Herein, zwitterionic nanocomposite membranes consisting of cross-linked poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC) and bacterial nanocellulose (BNC) were prepared and tested as tools for water remediation. These nanocomposite membranes fabricated via the one-pot polymerization of the zwitterionic monomer, 2-methacryloyloxyethyl phosphorylcholine, within the BNC three-dimensional porous network, exhibit thermal stability up to 250 °C, good mechanical performance (Young's modulus ≥ 430 MPa) and high water-uptake capacity (627%-912%) in different pH media. Moreover, these zwitterionic membranes reduced the bacterial concentration of both gram-positive (Staphylococcus aureus) and gram-negative (Escherichia coli) pathogenic bacteria with maxima of 4.3- and 1.8-log CFU reduction, respectively, which might be a major advantage in reducing or avoiding bacterial growth in contaminated water. The removal of two water-soluble model dyes, namely methylene blue (MB, cationic) and methyl orange (MO, anionic), from water was also assessed and the results demonstrated that both dyes were successfully removed under the studied conditions, reaching a maximum of ionic dye adsorption of ca. 4.4-4.5 mg g-1. This combination of properties provides these PMPC/BNC nanocomposites with potential for application as antibacterial bio-based adsorbent membranes for water remediation of anionic and cationic dyes.

MIR spectroscopy as alternative method for further confirmation of foodborne pathogens Salmonella spp. and Listeria monocytogenes.

Listeriosis and Salmonellosis are two of the most common foodborne diseases. Consequently, an early and accurate detection of Listeria monocytogenes and Salmonella spp. in food products is a critical concern of public health policies. Therefore, it is of great interest to develop rapid, simple, and inexpensive alternatives for pathogen detection in food products. In this study, mid-infrared spectroscopy has been successfully used to confirm Listeria species and the presence of Salmonella isolated from food samples. This methodology showed to be very sensitive and could be a rapid alternative to detect these important pathogens, allowing to obtain results in a few minutes after previous growth in selective media, avoiding the confirmation procedures that delay the achievement of the results for up to 2 days.

Protein Expression Modifications in Phage-Resistant Mutants of Aeromonas salmonicida after AS-A Phage Treatment.

The occurrence of infections by pathogenic bacteria is one of the main sources of financial loss for the aquaculture industry. This problem often cannot be solved with antibiotic treatment or vaccination. Phage therapy seems to be an alternative environmentally-friendly strategy to control infections. Recognizing the cellular modifications that bacteriophage therapy may cause to the host is essential in order to confirm microbial inactivation, while understanding the mechanisms that drive the development of phage-resistant strains. The aim of this work was to detect cellular modifications that occur after phage AS-A treatment in A. salmonicida, an important fish pathogen. Phage-resistant and susceptible cells were subjected to five successive streak-plating steps and analysed with infrared spectroscopy, a fast and powerful tool for cell study. The spectral differences of both populations were investigated and compared with a phage sensitivity profile, obtained through the spot test and efficiency of plating. Changes in protein associated peaks were found, and these results were corroborated by 1-D electrophoresis of intracellular proteins analysis and by phage sensitivity profiles. Phage AS-A treatment before the first streaking-plate step clearly affected the intracellular proteins expression levels of phage-resistant clones, altering the expression of distinct proteins during the subsequent five successive streak-plating steps, making these clones recover and be phenotypically more similar to the sensitive cells.

Effects of single and combined use of bacteriophages and antibiotics to inactivate Escherichia coli.

A major concern of phage therapy is the emergency of phage-resistant mutants. This limitation can be overcome by the combined use of phages and antibiotics. It has been shown that the combination of antibiotics and phages is an alternative that cannot only be effective at reducing bacterial numbers, but also to contribute to the management of resistance levels. However, this view has only been discussed with regard to antibiotic resistance and not to control phage-mutant emergence. In our study we compared not only the resistance of the bacteria to the four antibiotics tested with and without phages addition, but also the resistance to the phages in the presence and absence of antibiotics. The aim of this study was to evaluate the potential synergistic effect of phages and antibiotics in the inactivation of Escherichia coli in order to control infections, namely urinary tract infection (UTI), and to reduce the development of bacterial resistance to phages. Phage therapy combined with antibiotics (ampicillin, piperacillin, kamanycin, tetracycline, chloramphenicol and ciprofloxacin) was evaluated in the inactivation of E. coli, both in saline solution and urine samples. Phage and antibiotic combinations could result in high synergistic effects in the inactivation of bacteria. The combination of phage and ciprofloxacin at sublethal concentration decreased the bacterial counts in urine samples by 7.8±0.1 log CFU/ml after 8h, but when phages or the antibiotic were tested alone, the decrease was of 3.9±0.3 log CFU/mL and 1.2±0.1 log CFU/mL, respectively, after the same time. The efficacy of the combination of the two therapies depends on the antibiotic resistance status of the targeted bacteria to the employed antibiotic and of the antibiotic type (bactericide or bacteriostatic), causing the same or less bacterial resistance than phages and antibiotics applied alone (1.2±1.0×10-5 to 2.4±1.5×10-7 CFU/mL for the combined treatment, 2.7±0.2×10-4 CFU/mL for the antibiotics and 5.0±1.5×10-6 CFU/mL for the phages). The addition of antibiotics, at subinhibitory concentration, during phage treatment can control the phage-mutant. The high bacterial inactivation efficiency of these combined techniques and the long periods of phage survival in urine, pave the way for depth studies to control UTI and to overcome the development of resistances by bacteria.

Efficient Catalytic Oxidation of 3-Arylthio- and 3-Cyclohexylthio-lapachone Derivatives to New Sulfonyl Derivatives and Evaluation of Their Antibacterial Activities.

New sulfonyl-lapachones were efficiently obtained through the catalytic oxidation of arylthio- and cyclohexylthio-lapachone derivatives with hydrogen peroxide in the presence of a Mn(III) porphyrin complex. The antibacterial activities of the non-oxidized and oxidized lapachone derivatives against the Gram-negative bacteria Escherichia coli and the Gram-positive bacteria Staphylococcus aureus were evaluated after their incorporation into polyvinylpyrrolidone (PVP) micelles. The obtained results show that the PVP-formulations of the lapachones 4b-g and of the sulfonyl-lapachones 7e and 7g reduced the growth of S. aureus.

Characterization and in vitro evaluation of new bacteriophages for the biocontrol of Escherichia coli.

In the present study two new phages (phT4A and ECA2) were characterized and their efficacy was evaluated separately and in cocktail (phT4A/ECA2) to control Escherichia coli. The isolated phages, phT4A and ECA2, belonged to the Myoviridae and Podoviridae family, respectively and both are safe (no integrase and toxin codifying genes) to be used in bacterial control. In general, the increase of multiplicity of infection (MOI) from 1 to 100 promoted a significant increase in the efficiency of phage phT4A and phage cocktail phT4A/ECA2. Both phages were effective against E. coli, but phage phT4A (reduction of 5.8 log CFU/mL after 8h treatment) was more effective than phage ECA2 phage (reduction of 4.7 log CFU/mL after 8h treatment). The use of a cocktail phT4A/ECA2 was significantly more effective (reductions of 6.2 log CFU/mL after 6h treatment) than the use single phage suspensions of phT4A and ECA2 (reductions 5.3 log CFU/mL and 4.9 log CFU/mL, respectively, after 6h treatment). The rate of emergence of phage-resistant mutants was lower for phage phT4A when compared with phage ECA2 and phage cocktail phT4A/ECA2.The results indicate that in addition to the efficacy, the potential development of phage-resistant mutants must also be considered in the design of phage cocktails.

Application of phage therapy during bivalve depuration improves Escherichia coli decontamination.

The present study investigated the potential application of the bacteriophage (or phage) phT4A, ECA2 and the phage cocktail phT4A/ECA2 to decrease the concentration of Escherichia coli during the depuration of natural and artificially contaminated cockles. Depuration in static seawater at multiplicity of infection (MOI) of 1 with single phage suspensions of phT4A and ECA2 was the best condition, as it decreased by ∼2.0 log CFU/g the concentration of E. coli in artificially contaminated cockles after a 4 h of treatment. When naturally contaminated cockles were treated in static seawater with single phage suspensions and the phage cocktail, similar decreases in the concentration of E. coli (∼0.7 log CFU/g) were achieved. However, when employing the phage cocktail, a longer treatment time was required to obtain comparable results to those achieved when using single phage suspensions. When naturally contaminated cockles were depurated with phage phT4A in a recirculated seawater system (mimicking industrial depuration conditions), a 0.6 log CFU/g reduction of E. coli was achieved after a 2 h of treatment. When the depuration process was performed without phage addition, a 4 h treatment was necessary to obtain a similar decrease. By combining phage therapy and depuration procedures, a reduction in bivalves depuration period can be achieved for, thus decreasing the cost associated with this procedure and even enhance the quality and safety of depurated bivalves destined for human consumption.

Overall biochemical changes in bacteria photosensitized with cationic porphyrins monitored by infrared spectroscopy.

BACKGROUND: Photodynamic inactivation of micro-organisms is a promising nonantibiotic multitarget approach to treat localized and superficial infections through oxidative stress. Herein, the changes occurring on major cellular components of Escherichia coli and Staphylococcus warneri, induced by photosensitization with cationic porphyrins (Tri-Py(+)-Me-PF and Tetra-Py(+)-Me) and white light, were monitored by infrared spectroscopy. RESULTS: In E. coli, most of the changes occurred on proteins and lipids, suggesting a key effect on lipopolysaccharides in the first irradiation times. In S. warneri, proteins were the major molecular targets of oxidative damage but phospholipids and polysaccharides were also affected. CONCLUSION: Infrared spectroscopy is a very interesting tool to monitor biochemical changes induced by photosensitization in bacteria and also to infer on its mechanism of action.

Bacteriophages with potential to inactivate Salmonella Typhimurium: Use of single phage suspensions and phage cocktails.

The aim of this study was to compare the dynamics of three previously isolated bacteriophages (or phages) individually (phSE-1, phSE-2 and phSE-5) or combined in cocktails of two or three phages (phSE-1/phSE-2, phSE-1/phSE-5, phSE-2/phSE-5 and phSE-1/phSE-2/phSE-5) to control Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) in order to evaluate their potential application during depuration. Phages were assigned to the family Siphoviridae and revealed identical restriction digest profiles, although they showed a different phage adsorption, host range, burst size, explosion time and survival in seawater. The three phages were effective against S. Typhimurium (reduction of ∼2.0 log CFU/mL after 4h treatment). The use of cocktails was not significantly more effective than the use of single phages. A big fraction of the remained bacteria are phage-resistant mutants (frequency of phage-resistant mutants 9.19×10(-5)-5.11×10(-4)) but phage- resistant bacterial mutants was lower for the cocktail phages than for the single phage suspensions and the phage phSE-1 presented the highest rate of resistance and phage phSE-5 the lowest one. The spectral changes of S. Typhimurium resistant and phage-sensitive cells were compared and revealed relevant differences for peaks associated to amide I (1620cm(-1)) and amide II (1515cm(-1)) from proteins and from carbohydrates and phosphates region (1080-1000cm(-1)). Despite the similar efficiency of individual phages, the development of lower resistance indicates that phage cocktails might be the most promising choice to be used during the bivalve depuration to control the transmission of salmonellosis.

Application of bacteriophages during depuration reduces the load of Salmonella Typhimurium in cockles.

As bivalve molluscs are filter feeder, often consumed raw or lightly cooked and are frequently cultivated in contaminated waters, they are implicated in food-borne disease transmission to human. The present study investigated the potential application of bacteriophage (or phage) phSE-2, phage phSE-5 and phage cocktail phSE-2/phSE-5 to decrease the concentration of Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) during the depuration of natural and artificially contaminated cockles (Cerastoderma edule). Cockles were artificially infected with 105 and 106colony-forming units (CFU)/mL of S. Typhimurium in static seawater and infected group were treated with phages at four different MOI values: 0.1, 1, 10 and 100. Depuration in static seawater at multiplicity of infection (MOI) of 0.1 with single phage suspensions of phSE-2 and phSE-5 provided the best results, as it decreased by ~1.3 and 1.7 logCFU/g, respectively, the concentration of Salmonella spp. after a 4h treatment. At a MOI of 0.1, the rate of inactivation with single phage suspensions was higher when compared with the results obtained using the phage cocktail. However, in naturally contaminated cockles treated in static seawater with single phage suspensions and phage cocktail phSE-2/phSE-5, similar decreases in cultivable bacteria concentration (~0.7-0.9 logCFU/g) were achieved after 6h of treatment. When artificially contaminated cockles were depurated with phage phSE-5 in a recirculated seawater system (mimicking industrial depuration conditions), a 0.9 and 2.0 logCFU/g reduction of Salmonella spp. was reached after 4 and 6h treatment. Once the depuration process was performed without phage, a 6h treatment was needed to obtain a 1.1 logCFU/g reduction of Salmonella spp. Results indicated that combining phage biocontrol with depuration procedures enhance bivalve microbial safety for human consumption by improving decontamination efficiency, proving that this technology can be transposed to the bivalves industry. Moreover, this approach also displays the advantage of reducing the time required for depuration and consequently its associated costs.

SDS-PAGE and IR spectroscopy to evaluate modifications in the viral protein profile induced by a cationic porphyrinic photosensitizer.

Reactive oxygen species can be responsible for microbial photodynamic inactivation due to its toxic effects, which include severe damage to proteins, lipids and nucleic acids. In this study, the photo-oxidative modifications of the proteins of a non-enveloped T4-like bacteriophage, induced by the cationic porphyrin 5,10,15-tris(1-methylpyridinium-4-yl)-20-(pentafluorophenyl)porphyrin tri-iodide were evaluated. Two methods were used: sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and infrared spectroscopy. SDS-PAGE analysis showed that the phage protein profile was considerably altered after photodynamic treatment. Seven protein bands putatively corresponding to capsid and tail tube proteins were attenuated and two other were enhanced. Infrared spectroscopy confirmed the time-dependent alteration on the phage protein profile detected by SDS-PAGE, indicative of a response to oxidative damage. Infrared analysis showed to be a promising and rapid screening approach for the analysis of the modifications induced on viral proteins by photosensitization. In fact, one single infrared spectrum can highlight the changes induced to all viral molecular structures, overcoming the delays and complex protocols of the conventional methods, in a much simple and cost effective way.

Effects of UV radiation on the lipids and proteins of bacteria studied by mid-infrared spectroscopy.

Knowledge of the molecular effects of UV radiation (UVR) on bacteria can contribute to a better understanding of the environmental consequences of enhanced UV levels associated with global climate changes and will help to optimize UV-based disinfection strategies. In the present work, the effects of exposure to UVR in different spectral regions (UVC, 100-280 nm; UVB, 280-320 nm; and UVA, 320-400 nm) on the lipids and proteins of two bacterial strains ( Acinetobacter sp. strain PT5I1.2G and Pseudomonas sp. strain NT5I1.2B) with distinct UV sensitivities were studied by mid-infrared spectroscopy. Exposure to UVR caused an increase in methyl groups associated with lipids, lipid oxidation, and also led to alterations in lipid composition, which were confirmed by gas chromatography. Additionally, mid-infrared spectroscopy revealed the effects of UVR on protein conformation and protein composition, which were confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), oxidative damage to amino acids, and changes in the propionylation, glycosylation and/or phosphorylation status of cell proteins. Differences in the targets of UVR in the two strains tested were identified and may explain their discrepant UV sensitivities. The significance of the results is discussed from an ecological standpoint and with respect to potential improvements in UV-based disinfection technologies.