Base Sequence

Example 2

As discussed herein above, the disclosed methods improve the antiseptic properties of a dental implant without using charged metallic ions via conversion of the nitrogen moieties in titanium nitride surface to a positively charged quaternary ammonium via a Menschutkin reaction.

To prepare the antibacterial quaternized TiN surface, an implant which has been coated with TiN was used. The implant was cleaned to improve yield. The implant was washed with two solvents in sequence, acetone and isopropanol, to remove any dust particulate and other residue. The native oxide layer was removed by sonicating in 1:10 HCl:deionized water for 1 minute. This treatment additionally removes any residue that may not have been removed by the solvents. Acetonitrile was used as the solvent; however, any solvent may be used with preference for polar solvents giving improved reaction times (Stanger K., et al. J Org Chem. 2007 72(25):9663-8; Harfenist M., et al. J Am Chem Soc 1957 79(16):4356-4358). An excess of allyl bromide was added to the solvent and continuously stirred. The sample was then submerged in the solution, and full reaction of the surface occurred within about 60 minutes, as confirmed by contact angle measurement. A reference was also measured by submerging in solvent for the duration with no reactant to ensure any changes in surface properties was due to the quaternization.

Without wishing to be bound by a particular theory, the increased hydrophobicity of the treated surfaces can be due to the presence of the allyl groups on the surface which will impart some hydrophobicity. The contact angle measurements provide information on whether or not a reaction has occurred and whether it has saturated.

The biocidal activity was tested using live bacteria cultures from a patient's mouth, which provides the full flora to act against rather than targeting an individual strain of bacteria. The bacteria was incubated on the sample surface using several bacteria film thicknesses. The thickness is defined by keeping the same interaction surface area while varying the volume of bacteria solution added. Across two separate patients and several separate growths, within 4 hours 40-50% reduction in bacteria unit counts were observed for quaternized TiN as compared to traditional Titanium implants, outperforming traditional TiN coatings. FIG. 4 shows for two separate patients a set of typical bacteria growth result of the quaternized samples. The exact efficiency varies, as each patient has different flora which varies depending on environmental factors such as hygiene, diet, and familial history.

Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of skill in the art to which the disclosed invention belongs. Publications cited herein and the materials for which they are cited are specifically incorporated by reference.

It will be apparent to those skilled in the art that various modifications and variations can be made in the present disclosure without departing from the scope or spirit of the disclosure. Other aspects of the disclosure will be apparent to those skilled in the art from consideration of the specification and practice of the disclosure disclosed herein. It is intended that the specification and examples be considered as exemplary only, with a true scope and spirit of the disclosure being indicated by the following claims.

Example 3

Amino acids sequences alignment shows 67% identity between OR5A1 and OR5A2, 58% with OR5AN1 and 41% with OR11A1 (FIGS. 2A-B). To further address the question of how well paralogy predicts functionality and selectivity, we compared the response of these ORs to beta-ionone and 2-ethyl fenchol, the two well-known agonists of OR5A1 and OR11A1 respectively (Jaeger et al., 2013; Adipietro et al., 2012). These compounds were tested in concentration-response analysis in luciferase assays, as described previously. In each experiment, an empty vector was used as negative control (pEFIBRHO). Representative concentration-response curves are given in FIGS. 3A-B.

It was observed that OR5A1, the closest paralog of OR5A2, and OR11A1 are both activated by their own cognate agonist. On the contrary, in these experimental conditions, OR of the invention (namely OR5A2) as well as OR5AN1 are stimulated neither by beta-ionone nor by 2 ethyl-fenchol, both showing concentration-response curves similar to the empty vector.

Altogether, these results indicate that OR5A1 and OR5A2, although members of the same subfamily, show different agonist specificity (beta-ionone vs musk) indicating that amino acids similarity doesn't robustly predict OR selectivity and functionality among paralogs.

Example 16

The instant study was designed to test the efficacy in cotton rats of hMPV vaccines against a lethal challenge. mRNA vaccines encoding hMPV fusion protein were used. The mRNA polynucleotide encodes a full-length fusion protein and comprises the wild-type nucleotide sequence obtained from the hMPV A2a strain.

Cotton rats were immunized intramuscularly (IM) at week 0 and week 3 with the mRNA vaccines encoding hMPV fusion protein with either 2 μg or 10 μg doses for each immunization. The animals were then challenged with a lethal dose of hMPV in week 7 post initial immunization via IV, IM or ID. The endpoint was day 13 post infection, death or euthanasia. Viral titers in the noses and lungs of the cotton rats were measured. The results (FIGS. 9A and 9B) show that a 10 μg dose of mRNA vaccine protected the cotton mice 100% in the lung and drastically reduced the viral titer in the nose after challenge (˜2 log reduction). Moreover, a 2 μg dose of mRNA vaccine showed a 1 log reduction in lung viral titer in the cotton mice challenged.

Further, the histopathology of the lungs of the cotton mice immunized and challenged showed no pathology associated with vaccine-enhanced disease (FIG. 10).