Longitudinal Assessment of BNT162b2- and mRNA-1273-Induced Anti-SARS-CoV-2 Spike IgG Levels and Avidity Following Three Doses of Vaccination.

SARS-CoV-2 vaccination-induced protection against infection is likely to be affected by functional antibody features. To understand the kinetics of antibody responses in healthy individuals after primary series and third vaccine doses, sera from the recipients of the two licensed SARS-CoV-2 mRNA vaccines were assessed for circulating anti-SARS-CoV-2 spike IgG levels and avidity for up to 6 months post-primary series and 9 months after the third dose. Following primary series vaccination, anti-SARS-CoV-2 spike IgG levels declined from months 1 to 6, while avidity increased through month 6, irrespective of the vaccine received. The third dose of either vaccine increased anti-SARS-CoV-2 spike IgG levels and avidity and appeared to enhance antibody level persistence-generating a slower rate of decline in the 3 months following the third dose compared to the decline seen after the primary series alone. The third dose of both vaccines induced significant avidity increases 1 month after vaccination compared to the avidity response 6 months post-primary series vaccination (p ≤ 0.001). A significant difference in avidity responses between the two vaccines was observed 6 months post-third dose, where the BNT162b2 recipients had higher antibody avidity levels compared to the mRNA-1273 recipients (p = 0.020).

SARS-CoV-2 IgG Spike antibody levels and avidity in natural infection or following vaccination with mRNA-1273 or BNT162b2 vaccines.

Certain aspects of the immunogenicity and effectiveness of the messenger ribonucleic acid (mRNA) vaccines (mRNA-1273 and BNT162b2) developed in response to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic are still uncharacterized. Serum or plasma samples from healthy donor recipients of either vaccine (BNT162b2 n = 53, mRNA-1273 n = 49; age 23-67), and individuals naturally infected with SARS-CoV-2 (n = 106; age 18-82) were collected 0-2 months post-infection or 1- and 4 months after second dose of vaccination. Anti-Spike antibody levels and avidity were measured via an enzyme-linked immunosorbent assay (ELISA). Overall, vaccination induced higher circulating anti-Spike protein immunoglobulin G (IgG) antibody levels and avidity compared to infection at similar time intervals. Both vaccines produced similar anti-Spike IgG concentrations at 1 month, while mRNA-1273 demonstrated significantly higher circulating antibody concentrations after 4 months. mRNA-1273 induced significantly higher avidity at month 1 compared to BNT162b2 across all age groups. However, the 23-34 age group was the only group to maintain statistical significance by 4 months. Male BNT162b2 recipients were approaching statistically significant lower anti-Spike IgG avidity compared to females by month 4. These findings demonstrate enhanced anti-Spike IgG levels and avidity following vaccination compared to natural infection. In addition, the mRNA-1273 vaccine induced higher antibody levels by 4 months compared to BNT162b2.

Influenza Antigens NP and M2 Confer Cross Protection to BALB/c Mice against Lethal Challenge with H1N1, Pandemic H1N1 or H5N1 Influenza A Viruses.

Influenza hemagglutinin (HA) is considered a major protective antigen of seasonal influenza vaccine but antigenic drift of HA necessitates annual immunizations using new circulating HA versions. Low variation found within conserved non-HA influenza virus (INFV) antigens may maintain protection with less frequent immunizations. Conserved antigens of influenza A virus (INFV A) that can generate cross protection against multiple INFV strains were evaluated in BALB/c mice using modified Vaccinia virus Ankara (MVA)-vectored vaccines that expressed INFV A antigens hemagglutinin (HA), matrix protein 1 (M1), nucleoprotein (NP), matrix protein 2 (M2), repeats of the external portion of M2 (M2e) or as tandem repeats (METR), and M2e with transmembrane region and cytoplasmic loop (M2eTML). Protection by combinations of non-HA antigens was equivalent to that of subtype-matched HA. Combinations of NP and forms of M2e generated serum antibody responses and protected mice against lethal INFV A challenge using PR8, pandemic H1N1 A/Mexico/4108/2009 (pH1N1) or H5N1 A/Vietnam/1203/2004 (H5N1) viruses, as demonstrated by reduced lung viral burden and protection against weight loss. The highest levels of protection were obtained with NP and M2e antigens delivered as MVA inserts, resulting in broadly protective immunity in mice and enhancement of previous natural immunity to INFV A.

Differential effects of Co(III), Ni(II), and Ru(III) amine complexes on Sindbis virus.

Transition metal complexes [Co(cyclen)(NH(3))(2)](ClO(4))(3)H(2)O (cyclen=1,4,7,10-tetraazacyclododecane) (2), [Co(NH(3))(5)(OH(2))](CF(3)SO(3))(3) (3) [Ni(NH(3))(6)]Br(2) (4) and [Ru(NH(3))(6)]Cl(3) (5) were tested against Sindbis infected baby hamster kidney (BHK) cells and show differential effects from the previously reported anti-viral complex [Co(NH(3))(6)]Cl(3) (1). The macrocyclic complex 2 and labile aqua complex 3 show either no or little effect on the survival on Sindbis virus-infected cells as compared to that for 1, which show a monotonic increase in % BHK cell survival. Nickel and ruthenium ammine complexes 4 and 5 had a moderate influence of cell survival. While the results showed some anti-viral activity for some of the structural variations, it appears that 1, with its potential to be a broad-spectrum anti-viral compound, occupies a unique position in its ability to both significantly enhance cell survival and to decrease viral expression of infected cells. We also show that 1 also shows anti-viral activity against Adenovirus lending support to the broad-spectrum potential of this complex.

Antiviral properties of cobalt(III)-complexes.

We have investigated the potential antiviral activity of three cobalt(III) compounds. Two compounds, Co(III)-cyclen-methylbenzoic acid and its methyl ester derivative, are based on the macrocyclic chelator, cyclen, and were synthesized in our laboratory. Both compounds have been shown to bind tightly to nucleic acids and to hydrolyze phosphodiester bonds. However, neither compound exhibited any significant antiviral activity in an in vitro model of Sindbis virus replication. In contrast, a third compound, Co(III)hexammine, significantly inhibited Sindbis virus replication in baby hamster kidney (BHK) cells in a dose- and time-dependent manner. In plaque assays, the incubation of Co(III)hexammine with Sindbis virus resulted in a dose-dependent decrease in virus replication when measured at both 24 and 48-h post-infection. Over the concentration range of 0-5mM Co(III)hexammine, the IC(50) for the inhibition of viral replication was determined to be 0.10+/-0.04mM at 48h. Additionally, when BHK cell monolayers were pretreated with Co(III)hexammine for 6h prior to Sindbis infection, optimal cellular morphology and plasma membrane integrity were observed at 0.6-1.2mM Co(III)hexammine. Analysis by flow cytometry confirmed that Co(III)hexammine mediated a concomitant dose-dependent increase in BHK cell viability and a decrease in the percentage of Sindbis virus-infected cells (IC(50)=0.13+/-0.04mM). Our findings demonstrate for the first time that Co(III)hexammine possesses potent antiviral activity. We discuss our findings within the context of the ability to further functionalize Co(III)hexammine to render it a highly specific antiviral therapeutic reagent.

Binding and neutralization of lipopolysaccharides by plant proanthocyanidins.

Proanthocyanidins (PACs), polyphenolic metabolites that are widely distributed in higher plants, have been associated with potential positive health benefits including antibacterial, chemotherapeutic, and antiatherosclerotic activities. In this paper, we analyze the binding of PACs from cranberries, tea, and grapes to lipopolysaccharide (LPS), a major component of the outer membrane of Gram-negative bacteria and the cause of several human illnesses. We demonstrate that in the case of cranberries, the most potent LPS-binding activity is contained within a PAC fraction composed of polymers with an average degree of polymerization of 21. The PAC fraction modestly inhibits the binding of LPS to the surface of HEK 293 cells expressing the full complement of LPS receptors (TLR4/MD2 and CD14), while it significantly abrogates the endocytosis of LPS. This PAC fraction also inhibits LPS-induced nuclear factor-kappaB activation in a manner that is not readily overcome by excess LPS. Such an effect is mediated through the inhibition of LPS interaction with TLR4/MD2 and the partial abrogation of LPS interaction with CD14. Importantly, PAC concentrations that mediate effective LPS neutralization elicit minimal in vitro cytotoxicity. Our results identify PACs as a new class of LPS-binding compound and suggest that they have potential utility in applications that necessitate either the purification and removal of LPS or the in vivo neutralization of LPS.

Heterogeneity of a Campylobacter jejuni protein that is secreted through the flagellar filament.

Cj0859c, or FspA, is a small, acidic protein of Campylobacter jejuni that is expressed by a sigma(28) promoter. Analysis of the fspA gene in 41 isolates of C. jejuni revealed two overall variants of the predicted protein, FspA1 and FspA2. Secretion of FspA occurs in broth-grown bacteria and requires a minimum flagellar structure. The addition of recombinant FspA2, but not FspA1, to INT407 cells in vitro resulted in a rapid induction of apoptosis. These data define a novel C. jejuni virulence factor, and the observed heterogeneity among fspA alleles suggests alternate virulence potential among different strains.

Intracellular survival of Campylobacter jejuni in human monocytic cells and induction of apoptotic death by cytholethal distending toxin.

Campylobacter jejuni 81-176 is capable of extensive replication within human monocytic cell vacuoles and induces apoptotic death via cytolethal distending toxin.

Campylobacter jejuni induces secretion of proinflammatory chemokines from human intestinal epithelial cells.

Campylobacter jejuni is a common cause of diarrhea in humans. While the pathogenic mechanisms of C. jejuni are not completely understood, host inflammatory responses are thought to be contributing factors. In this report, C. jejuni 81-176 is shown to up-regulate chemokines essential to inflammatory responses. Growth-related oncogene alpha (GROalpha), GROgamma, macrophage inflammatory protein 1, monocyte chemoattractant protein 1 (MCP-1), and gamma interferon-inducible protein 10 (gammaIP-10) mRNA transcription in INT-407 cells was enhanced within 4 h of bacterial exposure. Infection with viable campylobacters was necessary for sustained chemokine transcription and was NF-kappaB dependent. GROalpha, gammaIP-10, and MCP-1 chemokine secretions were confirmed by immunological assays.

DNA sequence and mutational analyses of the pVir plasmid of Campylobacter jejuni 81-176.

The circular pVir plasmid of Campylobacter jejuni strain 81-176 was determined to be 37,468 nucleotides in length with a G+C content of 26%. A total of 83% of the plasmid represented coding information, and all but 2 of the 54 predicted open reading frames were encoded on the same DNA strand. There were seven genes on the plasmid in a continguous region of 8.9 kb that encoded orthologs of type IV secretion proteins found in Helicobacter pylori, including four that have been described previously (D. J. Bacon, R. A. Alm, D. H. Burr, L. Hu, D. J. Kopecko, C. P. Ewing, T. J. Trust, and P. Guerry, Infect. Immun. 68:4384-4390, 2000). There were seven other pVir-encoded proteins that showed significant similarities to proteins encoded by the plasticity zones of either H. pylori J99 or 26695. Mutational analyses of 19 plasmid genes identified 5 additional genes that affect in vitro invasion of intestinal epithelial cells. These included one additional gene encoding a component of a type IV secretion system, an ortholog of Cj0041 from the chromosome of C. jejuni NCTC 11168, two Campylobacter plasmid-specific genes, and an ortholog of HP0996 from the plasticity zone of H. pylori 26695.

Phase variation of Campylobacter jejuni 81-176 lipooligosaccharide affects ganglioside mimicry and invasiveness in vitro.

The outer cores of the lipooligosaccharides (LOS) of many strains of Campylobacter jejuni mimic human gangliosides in structure. A population of cells of C. jejuni strain 81-176 produced a mixture of LOS cores which consisted primarily of structures mimicking GM(2) and GM(3) gangliosides, with minor amounts of structures mimicking GD(1b) and GD(2). Genetic analyses of genes involved in the biosynthesis of the outer core of C. jejuni 81-176 revealed the presence of a homopolymeric tract of G residues within a gene encoding CgtA, an N-acetylgalactosaminyltransferase. Variation in the number of G residues within cgtA affected the length of the open reading frame, and these changes in cgtA corresponded to a change in LOS structure from GM(2) to GM(3) ganglioside mimicry. Site-specific mutation of cgtA in 81-176 resulted in a major LOS core structure that lacked GalNAc and resembled GM(3) ganglioside. Compared to wild-type 81-176, the cgtA mutant showed a significant increase in invasion of INT407 cells. In comparison, a site-specific mutation of the neuC1 gene resulted in the loss of sialic acid in the LOS core and reduced resistance to normal human serum but had no affect on invasion of INT407 cells.