Assessing the synergistic potential of bacteriophage endolysins and antimicrobial peptides for eradicating bacterial biofilms.

Phage-encoded endolysins have emerged as a potential substitute to conventional antibiotics due to their exceptional benefits including host specificity, rapid host killing, least risk of resistance. In addition to their antibacterial potency and biofilm eradication properties, endolysins are reported to exhibit synergism with other antimicrobial agents. In this study, the synergistic potency of endolysins was dissected with antimicrobial peptides to enhance their therapeutic effectiveness. Recombinantly expressed and purified bacteriophage endolysin [T7 endolysin (T7L); and T4 endolysin (T4L)] proteins have been used to evaluate the broad-spectrum antibacterial efficacy using different bacterial strains. Antibacterial/biofilm eradication studies were performed in combination with different antimicrobial peptides (AMPs) such as colistin, nisin, and polymyxin B (PMB) to assess the endolysin's antimicrobial efficacy and their synergy with AMPs. In combination with T7L, polymyxin B and colistin effectively eradicated the biofilm of Pseudomonas aeruginosa and exhibited a synergistic effect. Further, a combination of T4L and nisin displayed a synergistic effect against Staphylococcus aureus biofilms. In summary, the obtained results endorse the theme of combinational therapy consisting of endolysins and AMPs as an effective remedy against the drug-resistant bacterial biofilms that are a serious concern in healthcare settings.

Mechanism of Virus Inactivation by Cold Atmospheric-Pressure Plasma and Plasma-Activated Water.

Viruses cause serious pathogenic contamination that severely affects the environment and human health. Cold atmospheric-pressure plasma efficiently inactivates pathogenic bacteria; however, the mechanism of virus inactivation by plasma is not fully understood. In this study, surface plasma in argon mixed with 1% air and plasma-activated water was used to treat water containing bacteriophages. Both agents efficiently inactivated bacteriophages T4, Φ174, and MS2 in a time-dependent manner. Prolonged storage had marginal effects on the antiviral activity of plasma-activated water. DNA and protein analysis revealed that the reactive species generated by plasma damaged both nucleic acids and proteins, consistent with the morphological examination showing that plasma treatment caused the aggregation of bacteriophages. The inactivation of bacteriophages was alleviated by the singlet oxygen scavengers, demonstrating that singlet oxygen played a primary role in this process. Our findings provide a potentially effective disinfecting strategy to combat the environmental viruses using cold atmospheric-pressure plasma and plasma-activated water.IMPORTANCE Contamination with pathogenic and infectious viruses severely threatens human health and animal husbandry. Current methods for disinfection have different disadvantages, such as inconvenience and contamination of disinfection by-products (e.g., chlorine disinfection). In this study, atmospheric surface plasma in argon mixed with air and plasma-activated water was found to efficiently inactivate bacteriophages, and plasma-activated water still had strong antiviral activity after prolonged storage. Furthermore, it was shown that bacteriophage inactivation was associated with damage to nucleic acids and proteins by singlet oxygen. An understanding of the biological effects of plasma-based treatment is useful to inform the development of plasma into a novel disinfecting strategy with convenience and no by-product.

New Insights into DNA Polymerase Function Revealed by Phosphonoacetic Acid-Sensitive T4 DNA Polymerases.

The bacteriophage T4 DNA polymerase (pol) and the closely related RB69 DNA pol have been developed into model enzymes to study family B DNA pols. While all family B DNA pols have similar structures and share conserved protein motifs, the molecular mechanism underlying natural drug resistance of nonherpes family B DNA pols and drug sensitivity of herpes DNA pols remains unknown. In the present study, we constructed T4 phages containing G466S, Y460F, G466S/Y460F, P469S, and V475W mutations in DNA pol. These amino acid substitutions replace the residues in drug-resistant T4 DNA pol with residues found in drug-sensitive herpes family DNA pols. We investigated whether the T4 phages expressing the engineered mutant DNA pols were sensitive to the antiviral drug phosphonoacetic acid (PAA) and characterized the in vivo replication fidelity of the phage DNA pols. We found that G466S substitution marginally increased PAA sensitivity, whereas Y460F substitution conferred resistance. The phage expressing a double mutant G466S/Y460F DNA pol was more PAA-sensitive. V475W T4 DNA pol was highly sensitive to PAA, as was the case with V478W RB69 DNA pol. However, DNA replication was severely compromised, which resulted in the selection of phages expressing more robust DNA pols that have strong ability to replicate DNA and contain additional amino acid substitutions that suppress PAA sensitivity. Reduced replication fidelity was observed in all mutant phages expressing PAA-sensitive DNA pols. These observations indicate that PAA sensitivity and fidelity are balanced in DNA pols that can replicate DNA in different environments.

Reversible inactivation and desiccation tolerance of silicified viruses.

Long-distance host-independent virus dispersal is poorly understood, especially for viruses found in isolated ecosystems. To demonstrate a possible dispersal mechanism, we show that bacteriophage T4, archaeal virus Sulfolobus spindle-shaped virus Kamchatka, and vaccinia virus are reversibly inactivated by mineralization in silica under conditions similar to volcanic hot springs. In contrast, bacteriophage PRD1 is not silicified. Moreover, silicification provides viruses with remarkable desiccation resistance, which could allow extensive aerial dispersal.

Photocatalytic inactivation of bacteriophages by TiO2-coated glass plates under low-intensity, long-wavelength UV irradiation.

The International Organization for Standardization (ISO) was used to evaluate antibacterial activity by titanium dioxide (TiO(2)) photocatalysis since 2006. We evaluated photocatalytic inactivation of Qß and T4 bacteriophages induced by low-intensity, long-wavelength ultraviolet A (UVA; 0.1 mW cm(-2) and 0.001 mW cm(-2)) irradiation on a TiO(2)-coated glass plate using the ISO methodology. The results indicated that both bacteriophages were inactivated at 0.001 mW cm(-2) UVA. The intensity of UV light, including long-wavelength light (UVA), is very low in an actual indoor environment. Thus, TiO(2) photocatalysis can be beneficial for inactivating viruses in an indoor environment. Experiments using qPCR and bovine serum albumin degradation assume that viral inactivation is caused by outer viral protein disorder and not by viral RNA reduction by reactive oxygen species produced during TiO(2) photocatalysis. Furthermore, we showed that the ISO methodology for standard testing of antibacterial activity by TiO(2) photocatalysis can be applied to assess antiviral activity.

Highly efficient DNA compaction mediated by an in vivo antitumor-active tetrazolato-bridged dinuclear platinum(II) complex.

We investigated the effects of antitumor-active tetrazolato-bridged dinuclear platinum(II) complexes [{cis-Pt(NH(3))(2)}(2)(µ-OH)(µ-tetrazolato-N(1),N(2))](2+) (1) and [{cis-Pt(NH(3))(2)}(2)(µ-OH)(µ-tetrazolato-N(2),N(3))](2+) (2) on the higher-order structure of a large DNA molecule (T4 phage DNA, 166 kbp) in aqueous solution through single-molecule observation by fluorescence microscopy. Complexes 1 and 2 cause irreversible compaction of DNA through an intermediate state in which coil and compact parts coexist in a single DNA molecule. The potency of compaction is in the order 2 > 1 ≫ cisplatin. Transmission electron microscopic observation showed that both complexes collapsed DNA into an irregularly packed structure. Circular dichroism measurements revealed that the dinuclear platinum(II) complexes change the secondary structure of DNA from the B to C form. These characteristics of platinum(II) complexes are markedly different from those of the usual condensing agents such as spermidine(3+) and [Co(III)(NH(3))(6)](3+). The ability to cause DNA compaction by the platinum(II) complexes is discussed in relation to their potent antitumor activity.

Inactivation of Escherichia coli and bacteriophage T4 by high levels of dissolved CO2.

Little information is available regarding the effectiveness of water disinfection by CO(2) at low pressure. The aim of this study was to evaluate the use of high levels of dissolved CO(2) at 0.3-0.6 MPa for the inactivation of microorganisms. Bacteriophage T4 was chosen as the model virus and Escherichia coli was selected as the representative bacterium. The results of the study showed a highly effective log inactivation of E. coli and bacteriophage T4 at low and medium initial concentrations by high levels of dissolved CO(2) at 0.3 MPa with a treatment time of 20 min. When the pressure was increased to 0.6 MPa, inactivation of both microorganisms at high initial concentrations was improved to different extents. Neither pressurized air nor O(2) effectively inactivated both E. coli and bacteriophage T4. The pH was not a key factor affecting the inactivation process by this method. The results of scanning electron microscopy of E. coli and transmission electron microscopy of bacteriophage T4 suggested that "CO(2) uptake at high pressure and bursting of cells by depressurization" were the main reasons for lethal effect on microorganisms. This technology has potential for application in the disinfection of water, wastewater, and liquid food in the future.

Regression supports two mechanisms of fork processing in phage T4.

Replication forks routinely encounter damaged DNA and tightly bound proteins, leading to fork stalling and inactivation. To complete DNA synthesis, it is necessary to remove fork-blocking lesions and reactivate stalled fork structures, which can occur by multiple mechanisms. To study the mechanisms of stalled fork reactivation, we used a model fork intermediate, the origin fork, which is formed during replication from the bacteriophage T4 origin, ori(34). The origin fork accumulates within the T4 chromosome in a site-specific manner without the need for replication inhibitors or DNA damage. We report here that the origin fork is processed in vivo to generate a regressed fork structure. Furthermore, origin fork regression supports two mechanisms of fork resolution that can potentially lead to fork reactivation. Fork regression generates both a site-specific double-stranded end (DSE) and a Holliday junction. Each of these DNA elements serves as a target for processing by the T4 ATPase/exonuclease complex [gene product (gp) 46/47] and Holliday junction-cleaving enzyme (EndoVII), respectively. In the absence of both gp46 and EndoVII, regressed origin forks are stabilized and persist throughout infection. In the presence of EndoVII, but not gp46, there is significantly less regressed origin fork accumulation apparently due to cleavage of the regressed fork Holliday junction. In the presence of gp46, but not EndoVII, regressed origin fork DSEs are processed by degradation of the DSE and a pathway that includes recombination proteins. Although both mechanisms can occur independently, they may normally function together as a single fork reactivation pathway.

Low-Voltage Bacterial and Viral Killing Using Laser-Induced Graphene-Coated Non-woven Air Filters.

Laser-induced graphene (LIG) is uniquely positioned to advance applications in which electrically conductive carbon coatings are required. Recently, the antifouling, antiviral, and antibacterial properties of LIG have been proven in both air and water filtration applications. For example, an unsupported LIG based filter (pore size: ∼0.3 µm) demonstrated exceptional air filtration properties, while its joule heating effects successfully sterilized and removed unwanted biological components in air despite persisting challenges such as pressure drop, energy consumption, and lack of mechanical robustness. Here, we developed a polyimide (PI) non-woven supported LIG air filter with negligible pressure drop changes compared to the non-woven support material and showed that low electrical current density inactivates aerosolized bacteria. A current density of 4.5 mA/cm2 did not cause significant joule heating, and 97.2% bacterial removal was obtained. The low-voltage antibacterial mechanism was elucidated using bacterial inhibition experiments on a titanium surface and on an LIG surface fabricated on dense PI films. Complete sterilization was obtained using current densities of ∼8 mA/cm2 applied for 2 min or ∼ 6 mA/cm2 for 10 min upon the dense PI-LIG surface. Lastly, >98% bacterial removal was observed using a low-resistance LIG-coated non-woven polyimide air filter at 5 V. However, only very low voltages (∼0.3 V) were needed to remove ∼99% Pseudomonas aeruginosa bacteria and 100% of T4 virus when the LIG-coated filters were hybridized with a stainless steel mesh. Our results show that low current density levels at very low voltages are sufficient for substantial bacterial and viral inactivation, and that these principles might be effectively used in a wide number of air filtration applications such as air conditioners or other ventilation systems, which might limit the spread of infectious particles in hospitals, homes, workplaces, and the transportation industry.

Purification and concentration of bacteriophage T4 using monolithic chromatographic supports.

Phages are gaining importance due to their wide usage. In this work strong anion exchange monolithic chromatographic column was used for single step phage purification. Most of the proteins and DNA were removed and recovery of approximately 70% of infective virus was reproducibly achieved. 30 ml of phage sample was purified in around 10 min.

Biomolecular Simulations of Halogen Bonds with a GROMOS Force Field.

Halogen bonds (XBs) are non-covalent interactions in which halogens (X), acting as electrophiles, interact with Lewis bases. XBs are able to mediate protein-ligand recognition and therefore play an important role in rational drug design. In this context, the development of molecular modeling tools that can tackle XBs is paramount. XBs are predominantly explained by the existence of a positive region on the electrostatic potential of X named the σ-hole. Typically, with molecular mechanics force fields, this region is modeled using a charged extra point (EP) linked to X along the R-X covalent bond axis. In this work, we developed the first EP-based strategy for GROMOS force fields (specifically GROMOS 54A7) using bacteriophage T4 lysozyme in complex with both iodobenzene and iodopentafluorobenzene as a prototype system. Several EP parametrization schemes were tested by adding a virtual interaction site to ligand topologies retrieved from the Automated Topology Builder (ATB) and Repository. Contrary to previous approaches using other force fields, our analysis is based on the capability of each parametrization scheme to sample XBs during MD simulations. Our results indicate that the implementation of an EP at a distance from iodine corresponding to Rmin provides a good qualitative description of XBs in MD simulations, supporting the compatibility of our approach with the GROMOS 54A7 force field.

Sensitive fluorescence sensing of T4 polynucleotide kinase activity and inhibition based on DNA/polydopamine nanospheres platform.

5'-Polynucleotide kinase (PNK) is a crucial enzyme that catalyzes the phosphorylation of nucleic acid with 5'-OH termini and this phosphorylation reaction has been involved in many important cellular activities. The evaluation of PNK activity has received an increasing attention due to the significance of PNK. Here, the polydopamine nanospheres (PDANS) could adsorb single-stranded DNA (ssDNA) through π-π stacking or hydrogen bonding between nucleobases and aromatic groups of PDANS, while the interaction between double-stranded DNA (dsDNA) with PDANS was weakened due to the changed conformation. Hence, a novel DNA/PDANS platform was constructed for the sensitive and selective determination of T4 PNK activity based on the preferential binding properties of PDANS for ssDNA over dsDNA and the excellent fluorescence quenching property of PDANS. The dye-labeled dsDNA was phosphorylated by T4 PNK and then digested by λ exonuclease, yielding dye-labeled ssDNA, which would be adsorbed on the surface of the PDANS and the fluorescence was greatly quenched by PDANS. Because of the preferential binding properties of PDANS for ssDNA over dsDNA and the high quenching property of PDANS, the developed DNA/PDANS platform exhibited good analytical performance for T4 PNK sensing in complex biological matrix and applied to screening inhibitors. The proposed DNA/PDANS based platform is promising in developing high-throughput assays for drug screening and clinical diagnostics.

An antitumor drug-induced topoisomerase cleavage complex blocks a bacteriophage T4 replication fork in vivo.

Many antitumor and antibacterial drugs inhibit DNA topoisomerases by trapping covalent enzyme-DNA cleavage complexes. Formation of cleavage complexes is important for cytotoxicity, but evidence suggests that cleavage complexes themselves are not sufficient to cause cell death. Rather, active cellular processes such as transcription and/or replication are probably necessary to transform cleavage complexes into cytotoxic lesions. Using defined plasmid substrates and two-dimensional agarose gel analysis, we examined the collision of an active replication fork with an antitumor drug-trapped cleavage complex. Discrete DNA molecules accumulated on the simple Y arc, with branch points very close to the topoisomerase cleavage site. Accumulation of the Y-form DNA required the presence of a topoisomerase cleavage site, the antitumor drug, the type II topoisomerase, and a T4 replication origin on the plasmid. Furthermore, all three arms of the Y-form DNA were replicated, arguing strongly that these are trapped replication intermediates. The Y-form DNA appeared even in the absence of two important phage recombination proteins, implying that Y-form DNA is the result of replication rather than recombination. This is the first direct evidence that a drug-induced topoisomerase cleavage complex blocks the replication fork in vivo. Surprisingly, these blocked replication forks do not contain DNA breaks at the topoisomerase cleavage site, implying that the replication complex was inactivated (at least temporarily) and that topoisomerase resealed the drug-induced DNA breaks. The replication fork may behave similarly at other types of DNA lesions, and thus cleavage complexes could represent a useful (site-specific) model for chemical- and radiation-induced DNA damage.

[Comparison on resistance of bacteriophages to sodium dichloroisocyanurate in laboratory].

OBJECTIVE: To scan the most resistable bacteriophage as an indicator in disinfection tests, and to study the resistance of bacteriophage T4, Phichi 174D, and f2 to the sodium dichloroisocyanurate (NaDCC) in laboratory. METHODS: The virucidal activity of NaDCC against bacteriophage T4, Phichi 174D, and f2 were assessed by suspension test. The neutralizer was selected and be appraised by test of neutralizer. Bacteriophage T4, Phichi 174D, and f2 were detected and enumerated by the double-agar-layer plaque technique. RESULTS: (1) With 150 mg/L of available chlorine of NaDCC solution, within a contact time of 40 minutes, or 300 mg/L, 5 minutes, the reductions of bacteriophage T4 achieved the "disinfection" level [log(10) inactivation value or log(10) reduction value of bacteriophage T4 (log(10)No-log(10)Nt) > or = 4.00 log(10)]. (2) With 300 mg/L of available chlorine of NaDCC solution, within a contact time of 5 minutes, or 400 mg/L, 3 minutes, the reductions of bacteriophage Phichi 174D achieved the "disinfection" level. (3) With 2000 mg/L of available chlorine of NaDCC solution, within a contact time of 20 minutes, or 4000 mg/L, 5 minutes, the reductions of bacteriophage f2 might achieve the "disinfection" level. CONCLUSION: The order of resistance of the above three bacteriophages to NaDCC from greatest to smallest is as follows: bacteriophage f2 > bacteriophage T4 > bacteriophage Phichi 174D.

[A comparative study on the resistance of bacteriophage T4, phiXM174D, and f2 to glutaraldehyde in laboratory].

OBJECTIVE: To identify the best indicative bacteriophage in disinfection tests through comparing the resistance of bacteriophage T4, phiX174D, and f2 to glutaraldehyde. METHODS: The virucidal activities of glutaraldehyde against bacteriophage T4, phiX174D, and f2 were assessed with suspension tests along with neutralizer tests. The double-agar-layer plaque technique was used to detect the bacteriophage T4, phiX174D, and f2. RESULTS: (1)In a condition of 3000 mg/L of glutaraldehyde and 20 minutes of contact or 6000 mg/L of glutaraldehyde and 5 minutes of contact, "disinfection" level for bacteriophage T4 was achieved, with the log10 inactivation value (LIV) or log10o reduction value (LRV) (=log10No-log10Nt) > or = 4. 00 log10. (2) In a condition of 2500 mg/L of glutaraldehyde and 20 minutes of contact or 5000 mg/L of glutaraldehyde and 5 minutes of contact, the LIV for bacteriophage phiX174D reached "disinfection" level; (3) In a condition of 4000 mg/L of glutaraldehyde and 40 minutes of contact or 8000 mg/L of glutaraldehyde and 10 minutes of contact, the LIV for bacteriophage f2 reached "disinfection" level. CONCLUSION: Bacteriophage f2 and bacteriophage phiX174D has the strongest and weakest resistance to glutaraldehyde respectively.

Dual role of the sequence-specific bacteriophage T4 endoribonuclease RegB. mRNA inactivation and mRNA destabilization.

Gene regB of bacteriophage T4 encodes a sequence-specific endoribonuclease that introduces cuts in early phage messenger RNAs. Cutting takes place specifically in the middle of the tetranucleotide GGAG, as soon as the first minute of infection. Out of the 20 processing sites so far identified, seven are in Shine-Dalgarno sequences. The others are localized in intercistronic regions or within coding sequences. In the latter case, cutting efficiency is much lower. regB-dependent cleavages can occur within AU-rich sequences downstream of processed GGAG motifs that are not in effective translation initiation sites. We looked for possible consequences of regB-dependent cuts on gene expression in two early regions of the T4 chromosome. In the comC alpha region, none of the three major RegB cleavage sites is in a Shine-Dalgarno sequence, and in the motA region the unique regB-dependent processing site is found within the Shine-Dalgarno sequence of the gene. We find that in the region of gene comC alpha, RegB decreases two- to threefold the chemical half-life of early transcripts, but does not change the functional half-life of mRNAs coding for protein ComC alpha. The amount of MotA protein synthesized by the wild-type is half that obtained in a regB mutant infection. We show that this is a direct consequence of mRNA processing by RegB at the Shine-Dalgarno sequence of motA. This regB-mediated translation inhibition is not accompanied by an important modification in motA mRNA chemical half-life. We show that rapid shut-off of MotA protein synthesis that occurs soon after infection results both from RegB processing within the translation initiation region of motA and from early transcription inhibition followed by regB-independent breakdown of the motA mRNA.

The spectrum of acridine resistant mutants of bacteriophage T4 reveals cryptic effects of the tsL141 DNA polymerase allele on spontaneous mutagenesis.

Mutations in the ac gene of bacteriophage T4 confer resistance to acridine-inhibition of phage development. Previous studies had localized the ac gene region; we show that inactivation of T4 Open Reading Frame 52.2 confers the Acr phenotype. Thus, 52.2 is ac. The resistance mechanism is unknown. The ac gene provides a convenient forward mutagenesis assay. Its compact size (156 bp) simplifies mutant sequencing and diverse mutant types are found: base substitutions leading to missense or nonsense codons, in-frame deletions or duplications within the coding sequence, deletion or duplication frameshifts, insertions, complex mutations, and large deletions extending into neighboring sequences. Comparisons of spontaneous mutagenesis between phages bearing the wild-type or tsL141 alleles of DNA polymerase demonstrate that the impact of the mutant polymerase is cryptic when total spontaneous mutant frequencies are compared, but the DNA sequences of the ac mutants reveal a substantial alteration of fidelity by the mutant polymerase. The patterns of base substitution mutagenesis suggest that some site-specific mutation rate effects may reflect hotspots for mutagenesis arising by different mechanisms. A new class of spontaneous duplication mutations, having sequences inconsistent with misaligned pairing models, but consistent with nick-processing errors, has been identified at a hotspot in ac.

Bacteriophage T4 mutants hypersensitive to an antitumor agent that induces topoisomerase-DNA cleavage complexes.

Many antitumor agents and antibiotics affect cells by interacting with type II topoisomerases, stabilizing a covalent enzyme-DNA complex. A pathway of recombination can apparently repair this DNA damage. In this study, transposon mutagenesis was used to identify possible components of the repair pathway in bacteriophage T4. Substantial increases in sensitivity to the antitumor agent m-AMSA [4'-(9-acridinylamino)methanesulfon-m-anisidide] were found with transposon insertion mutations that inactivate any of six T4-encoded proteins: UvsY (DNA synaptase accessory protein), UvsW (unknown function), Rnh (RNase H and 5' to 3' DNA exonuclease), alpha-gt (alpha-glucosyl transferase), gp47.1 (uncharacterized), and NrdB (beta subunit of ribonucleotide reductase). The role of the rnh gene in drug sensitivity was further characterized. First, an in-frame rnh deletion mutation was constructed and analyzed, providing evidence that the absence of Rnh protein causes hypersensitivity to m-AMSA. Second, the m-AMSA sensitivity of the rnh-deletion mutant was shown to require a drug-sensitive T4 topoisomerase. Third, analysis of double mutants suggested that uvsW and rnh mutations impair a common step in the recombinational repair pathway for m-AMSA-induced damage. Finally, the rnh-deletion mutant was found to be hypersensitive to UV, implicating Rnh in recombinational repair of UV-induced damage.

Antiviral effects of polyphenols: development of bio-based cleaning wipes and filters.

Polyphenol molecules play multiple essential roles in plant physiology such as defences against plant-pathogens and micro-organisms. The present study reports a chemical modification of the surface of non-woven cellulosic fibre filters (Kimwipes(®)) by fixing polyphenol in order to confer them antiviral properties. The grafting of the non-woven fibres by the antiviral entity was performed using laccase. T4D bacteriophage virus of Escherichia coli B was used as virus model. Catechin polyphenol was tested as antiviral entity. Proteomic experiments were performed to quantify the potential protein target of catechin on viruses. When the modified filter was in contact with the viral suspension a large improvement in the reduction of the viral concentration was observed (5-log after 1h). Thus, we propose that this material could be used as virucidal wipes for the virus elimination from contaminated surfaces. Virus filtration experiments were performed by spraying an aerial suspension of T4D bacteriophage virus through the designed filter. The best virus capture factor f (ratio of upstream to downstream virus contents) was obtained when using 2 functionalized filters (f=2.9×10(3)). When these 2 layers were placed inside a commercial medical mask in place of its cellulose layer (Kolmi M24001 mask) (f=3.5×10(4)), the f ratio then reached 2.6×10(5) for 2h of filtration. Based on these results, this novel bio-based antiviral mask represents a significant improvement over conventional medical masks.

Selection of bacteriophage T4 antimutator DNA polymerases: a link between proofreading and sensitivity to phosphonoacetic acid.

During DNA replication, DNA polymerases alternate between DNA synthesis and proofreading the newly synthesized DNA. In order to understand the molecular details of how DNA polymerases determine the balance between polymerase and proofreading activities, it would be useful to have mutants which switch between the two activities either more or less frequently. Antimutator DNA polymerases switch more frequently and thus have more opportunity for proofreading. We have observed that mutant DNA polymerases which proofread less frequently have a mutator phenotype and are inhibited by the pyrophosphate analogue phosphonoacetic acid. Sensitivity to phosphonoacetic acid can be used to isolate second-site suppressor mutations. These suppressor mutations encode amino acid substitutions which produce antimutator DNA polymerases.