Combinatory effects of Dipterocarpus alatus twig emulgel: Wound-restoring, antibacterial, and anti-inflammatory activities against methicillin-resistant Staphylococcus aureus-infected mouse superficial wounds.

Dipterocarpus alatus has been used for the treatment of infectious skin diseases and ulcerative wounds in Thai traditional medicine. A major pathogen in human superficial skin infections is methicillin-resistant Staphylococcus aureus (MRSA). This study determined the wound healing, antibacterial, and anti-inflammatory activities of D. alatus twig emulgel against MRSA-infected mouse superficial skin wounds. Ethyl acetate-methanol crude extract of D. alatus twig was incorporated into emulgel at concentrations of 20 and 40 mg/g (D20 and D40) and its activity was compared to tetracycline emulgel (160 µg/g, Tetra). MRSA-infected superficial wounds demonstrated decreased skin barrier strength, increased transepidermal water loss (TEWL), and mast cell accumulation. Expression of toll-like receptor 2 (TLR-2), NF-κß, TNFα, IL-1ß, IL-6 and IL-10 genes were induced after MRSA infection. Daily application of 100 µL of D20 or D40 for 9 days restored skin barrier strength and TEWL while reducing mast cell and MRSA numbers compared to the non-treated group (MRSA-NT). The wounds treated with D20 and D40 were entirely healed on day 9. Expression of TLR-2 and cytokine-related genes NF-κß, TNFα, IL-1ß, IL-6 and IL-10 were normalized by treatment with either D20 or D40. Therefore, emulgel containing 20 to 40 mg/g ethyl acetate-methanol crude D. alatus twig extract is a good candidate for development as a topical formulation for MRSA-infected ulcerated wounds.

Reactogenicity and immunogenicity of the intradermal administration of BNT162b2 mRNA vaccine in healthy adults who were primed with an inactivated SARS-CoV-2 vaccine.

Because of the outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), safe and effective vaccines are urgently required. The shortage of effective vaccines is a major challenge in many developing countries. We studied intradermal (ID) fractional dose BNT162b2 mRNA (Comirnaty®, Pfizer-BioNTech) as a booster dose in healthy adults who were previously immunized with an inactivated SARS-CoV-2 vaccine. This is a retrospective cohort study that included healthy adults who were immunized with two doses of inactivated SARS-CoV-2 vaccine and received a booster dose with ID fractional dose or intramuscular (IM) full-dose BNT162b2 mRNA between August 1 to August 15, 2021. The primary endpoint was safety that included local and systemic adverse reactions. The secondary endpoints were levels of SARS-CoV-2 spike protein receptor-binding domain IgG antibody (anti-S-RBD IgG) and neutralizing antibody activity against the Delta variant (B.1.617.2) using surrogate viral neutralization test (sVNT) 3 weeks after the booster dose. A total of 43 healthy adults (median age of 31 years) were included in the study; among them, 23 participants received ID fractional dose (6 µg) BNT162b2 mRNA, and 20 participants received IM full-dose (30 µg) BNT162b2 mRNA. No serious adverse reactions were observed. Local adverse reactions occurred more frequently in the ID group. No differences were observed in the baseline level of anti-S-RBD IgG (289 vs 286 AU/mL, p > 0.9, in the ID and IM groups, respectively). After booster, anti-S-RBD IgG titer increased to 13294 (9255-19573) AU/mL in the ID group and 23456 (16943-38539) AU/mL in the IM group. All participants in the IM group and 95.6 % of participants in the ID group had seroconversion evaluated by sVNT (≥68 % inhibition to the Delta variant). ID administration of BNT162b2 mRNA was safe and well-tolerated and generated a robust immune response. Therefore, ID delivery of the BNT162b2 mRNA vaccine has the potential for a dose-sparing strategy.

Bioprospecting for Antibacterial Drugs: a Multidisciplinary Perspective on Natural Product Source Material, Bioassay Selection and Avoidable Pitfalls.

Bioprospecting is the exploration, extraction and screening of biological material and sometimes indigenous knowledge to discover and develop new drugs and other products. Most antibiotics in current clinical use (eg. ß-lactams, aminoglycosides, tetracyclines, macrolides) were discovered using this approach, and there are strong arguments to reprioritize bioprospecting over other strategies in the search for new antibacterial drugs. Academic institutions should be well positioned to lead the early stages of these efforts given their many thousands of locations globally and because they are not constrained by the same commercial considerations as industry. University groups can lack the full complement of knowledge and skills needed though (eg. how to tailor screening strategy to biological source material). In this article, we review three key aspects of the bioprospecting literature (source material and in vitro antibacterial and toxicity testing) and present an integrated multidisciplinary perspective on (a) source material selection, (b) legal, taxonomic and other issues related to source material, (c) cultivation methods, (d) bioassay selection, (e) technical standards available, (f) extract/compound dissolution, (g) use of minimum inhibitory concentration and selectivity index values to identify progressible extracts and compounds, and (h) avoidable pitfalls. The review closes with recommendations for future study design and information on subsequent steps in the bioprospecting process.

Effect of sialidase fusion protein (DAS 181) on human metapneumovirus infection of Hep-2 cells.

METHODS: Hep-2 cells were preincubated with DAS181 or control DAS185 (a mutated sialidase) prior to inoculation with human metapneumovirus strains. Infectivity was assessed by a cell-based ELISA quantitating human metapneumovirus matrix protein. The effect of DAS181 on binding of recombinant G attachment protein was also determined. RESULTS: DAS181 blocked infection of human metapneumovirus strains A2, B1, and B2 at low concentrations. No effect of DAS185 was observed. Binding of MPV G protein to Hep-2 cells was also markedly inhibited by preincubation of cells with DAS181. CONCLUSIONS: These results suggest that human metapneumovirus may utilize sialic acids as an entry cofactor. DAS181 may thus represent a new therapeutic agent useful for the treatment of human metapneumovirus.

hMPV lineage nomenclature and heparin binding.

Human metapneumovirus (hMPV), first described in 2001 [1], is responsible for causing serious respiratory illness in young children, the elderly and immunocompromised patients. Four distinct lineages of hMPV have been identified with the original nomenclature for these subgroups (A1, A2, B1 and B2), reported by van den Hoogen et al. [2], utilised by many. An alternate terminology (1A, 1B, 2A and 2B) was also published by Ishiguro et al. in 2004 [3] which has been adopted by others. However, this has caused some confusion in the interpretation of publication results as the terminology is similar yet describes different subtypes. As a result, a number of investigators have made a submission to the International Committee on Taxonomy of Viruses (ICTV, ICTV taxonomic proposal 2012.012V) for the official adoption of the original terminology as an approved nomenclature for hMPV [4]. We welcome this officially approved nomenclature which should provide clarification of these subtypes in future. Therefore to assist with the interpretation of our recently published research in the 2012 special issue of Viruses: Pneumoviruses and Metapneumoviruses entitled "Diversity in Glycosaminoglycan Binding Amongst hMPV G Protein Lineages" [5] we have updated the Figure 3 in this letter (see Figure 1), showing the proposed ICTV terminology compared to the Ishiguro classification (used in our publication). Note that in the original publication the alphanumeric order for the Ishiguro classification was transposed (e.g., 1A was referred to as A1).

Diversity in glycosaminoglycan binding amongst hMPV G protein lineages.

We have previously shown that hMPV G protein (B2 lineage) interacts with cellular glycosaminoglycans (GAGs). In this study we examined subtypes A1, A2 and B1 for this interaction. GAG-dependent infectivity of available hMPV strains was demonstrated using GAG-deficient cells and heparin competition. We expressed the G protein ectodomains from all strains and analysed these by heparin affinity chromatography. In contrast to the B2 lineage, neither the A2 or B1 G proteins bound to heparin. Sequence analysis of these strains indicated that although there was some homology with the B2 heparin-binding domains, there were less positively charged residues, providing a likely explanation for the lack of binding. Although sequence analysis did not demonstrate well defined positively charged domains in G protein of the A1 strain, this protein was able to bind heparin, albeit with a lower affinity than G protein of the B2 strain. These results indicate diversity in GAG interactions between G proteins of different lineages and suggest that the GAG-dependency of all strains may be mediated by interaction with an alternative surface protein, most probably the conserved fusion (F) protein. Analysis of both native and recombinant F protein confirmed that F protein binds heparin, supporting this conclusion.

Role of cellular glycosaminoglycans and charged regions of viral G protein in human metapneumovirus infection.

Human metapneumovirus (hMPV) is an important cause of lower respiratory tract disease, particularly in infants and young children. hMPV has two major glycoproteins, G and F, which are responsible for virus attachment and membrane fusion, respectively. We investigated the role of cellular glycosaminoglycans (GAGs) and G protein in hMPV infection. The pretreatment of hMPV with soluble heparin markedly inhibited the infection of HEp-2 cells. Recombinant G protein, comprising the extracellular domain of G, bound to heparin-agarose columns and also to HEp-2 cells. hMPV infection and G protein binding to HEp-2 cells was inhibited by other soluble GAGs, including chondroitin sulfates, by the enzymatic removal of cell surface GAGs with GAG lyases or by the pretreatment of cells with basic fibroblast growth factor. The role of cellular GAGs was confirmed by the binding of G protein to wild-type CHO cells but not to GAG-deficient CHO-pgsA745 cells. An analysis of the G protein sequence revealed two adjacent clusters of positively charged amino acids ((149)EKKKTRA(155) and (159)QRRGKGKE(166)). Truncated G fragments were expressed, and only the fragment containing these putative heparin binding domains retained heparin binding. The alanine mutagenesis of charged residues in either of these regions resulted in the loss of binding to heparin and to HEp-2 cells, suggesting that both sites are likely to be required for hMPV attachment. These results, taken together with the inhibition of hMPV infection by soluble G protein, indicate an important role for G protein and cellular GAGs in hMPV infection.