In vivo tropism of attenuated and pathogenic measles virus expressing green fluorescent protein in macaques

The global increase in measles vaccination has resulted in a significant reduction of measles mortality. The standard route of administration for the live-attenuated measles virus (MV) vaccine is subcutaneous injection, although alternative needle-free routes, including aerosol delivery, are under investigation. In vitro, attenuated MV has a much wider tropism than clinical isolates, as it can use both CD46 and CD150 as cellular receptors. To compare the in vivo tropism of attenuated and pathogenic MV, we infected cynomolgus macaques with pathogenic or attenuated recombinant MV expressing enhanced green fluorescent protein (GFP) (strains IC323 and Edmonston, respectively) via the intratracheal or aerosol route. Surprisingly, viral loads and cellular tropism in the lungs were similar for the two viruses regardless of the route of administration, and CD11c-positive cells were identified as the major target population. However, only the pathogenic MV caused significant viremia, which resulted in massive virus replication in B and T lymphocytes in lymphoid tissues and viral dissemination to the skin and the submucosa of respiratory epithelia. Attenuated MV was rarely detected in lymphoid tissues, and when it was, only in isolated infected cells. Following aerosol inhalation, attenuated MV was detected at early time points in the upper respiratory tract, suggesting local virus replication. This contrasts with pathogenic MV, which invaded the upper respiratory tract only after the onset of viremia. This study shows that despite in vitro differences, attenuated and pathogenic MV show highly similar in vivo tropism in the lungs. However, systemic spread of attenuated MV is restricted.

Molecular differences between two Jeryl Lynn mumps virus vaccine component strains, JL5 and JL2

The Jeryl Lynn (JL) vaccine against mumps virus (MuV) contains two components, MuV(JL5) and MuV(JL2), which differ by over 400 nt. Due to the occurrence of bias in the direction of mutation, these differences and those found in nucleotide sequences of different isolates of the minor component in the vaccine (MuV(JL2)) might be due to the effect of ADAR-like deaminases on MuV grown in tissue-cultured cells. A molecular clone Of MuV(JL2) (pMuV(JL2)) and MuV(JL2) -specific helper plasmids were constructed in order to investigate molecular interactions between MuV(JL5) and MuV(JL2), to augment the existing molecular clone Of MuV(JL)5 (pMuV(JL5)) and MuV(JL5) -specific helper plasmids. Genome and mRNA termini Of MuV(JL2) were characterized, and an unusual oligo-G insertion transcriptional editing event was detected near the F mRNA polyadenylation site of MuV(JL2), but not Of MuV(JL5). Genes encoding glycoproteins of rMuV(JL2) and rMuV(JL5) have been exchanged to characterize the oligo-G insertion, which associated with the specific sequence of the IF gene of MuV(JL2) and not with any other genes or the RNA-dependent RNA polymerase of strain MuV(JL2). The results indicate that a single G-to-A sequence change obliterates the co-transcriptional editing of the F mRNA and that this oligo-G insertion does not affect the growth of the virus.

Molecular investigations into vaginal immunization with HIV gp41 antigenic construct H4A in a quick release solid dosage form

A robust vaginal immune response is considered essential for an effective prophylactic vaccine that prevents transmission of HIV and other sexually acquired diseases. Considerable attention has recently focused on the potential of vaginally administered vaccines as a means to induce such local immunity. However, the potential for vaccination at this site remains in doubt as the vaginal mucosa is generally considered to have low immune inductive potential. In the current study, we explored for the first time the use of a quick release, freeze-dried, solid dosage system for practical vaginal administration of a protein antigen. These solid dosage forms overcome the common problem associated with leakage and poor retention of vaginally administered antigen solutions. Mice were immunized vaginally with H4A, an HIV gp41 envelope based recombinant protein, using quick release, freeze-dried solid rods, and the immune responses compared to a control group immunized via subcutaneous H4A injection. Vaginally immunized mice failed to elicit robust immune responses. Our detailed investigations, involving cytokine analysis, the stability of H4A in mouse cervicovaginal lavage, and elucidation of the state of H4A protein in the immediate-release dosage form, revealed that antigen instability in vaginal fluid, the state of the antigen in the dosage form, and the cytokine profile induced are all likely to have contributed to the observed lack of immunogenicity. These are important factors affecting vaginal immunization and provide a rational basis for explaining the typically poor and variable elicitation of immunity at this site, despite the presence of immune responsive cells within the vaginal mucosae. In future mucosal vaccine studies, a more explicit focus on antigen stability in the dosage form and the immune potential of available antigen-responsive cells is recommended. © 2012 Elsevier Ltd. All rights reserved.

Development of liposome-based freeze-dried rods for vaginal vaccine delivery against HIV-1

The present investigation deals with development and characteriza- tion of the liposomes-based freeze-dried rods for the vaginal delivery of gp140 antigen in mice. Positively charged, negatively charged and neutral liposomes were prepared and characterized for various parameters e.g. morphology, size, polydispersity index, zeta potential and antigen encapsulation efficiency. To further improve the efficacy of vaccine delivery, antigen encapsulated liposomes were formulated as polymer gel-based freeze-dried rods, which were then characterized for moisture content. The redispersibility of the liposomes-based freeze- dried rods was determined in simulated vaginal fluid and liposome gel was investigated for mucoadhesion. The developed liposome-based freeze-dried rods systems could offer potential as stable and practical dosage form for the mucosal immunization against HIV-1 infection.

Innate immunogenicity and in vitro protective potential of Schistosoma mansoni lung schistosomula excretory-secretory candidate vaccine antigens

Excretory secretory products (ESP) of Schistosoma mansoni developing larvae are ideal potential vaccines as such molecules may readily induce host primary immune responses, and local memory immune response effectors that would target, surround, and pursue the larvae while negotiating the lung blood capillaries. We herein characterized the cytokines response ESP, e.g., SG3PDH, 14-3-3-like protein, TPX, and calpain induce in the natural context of infection, and defined the global cytokine profile conducive to effective schistosome larvae killing. Accordingly, spleen cells (SC) taken from naive, and 7-, or 9-day S. mansoni-infected mice were stimulated in vitro with the selected ESP, in a recombinant or multiple antigen peptide (MAP) form, and examined for production of T helper type (Th) 1, Th2, and Th17 cytokines, and the ability to mediate in vitro attrition of lung-stage schistosomula. The study indicated that larval ESP principally elicit Th1 and Th17 type cytokines. Recombinant SG3PDH was the only test ESP to additionally activate SC from S. mansoni-infected BALB/c mice to release higher IL-4 levels than unstimulated SC and mediate significant (P

Immune responses and vaccine-induced immunity against Porcine circovirus type 2

Porcine circovirus type 2 (PCV2) is essential but not sufficient for postweaning multi-systemic wasting syndrome (PMWS) occurrence in pigs. The outcome of PCV2 infection depends on the specific immune responses that are developing during the infection. Diseased pigs are immunosupressed and unable to mount effective immune responses to clear the virus from circulation. In the final stage, PMWS-affected pigs suffer from extensive lymphoid lesions and altered cytokine expression patterns in peripheral blood mononuclear cells (PBMCs) and lymphoid organs. PCV2 infection can also be asymptomatic, demonstrating that not every infection will guarantee the occurrence of severe immunopathological disturbances. Asymptomatic animals have higher virus specific and neutralising antibody titres than PMWS-affected animals. Recent results have pointed out that the mechanisms by which PCV2 can affect the immune responses involve the induction of IL-10, virus accumulation into and modulation of plasmacytoid dendritic cells and the role of viral DNA in regulation of immune cell functions. Fourteen years after the first description of PMWS in Canada, efficient commercial vaccines against PCV2 are available. The vaccine success is based on activated humoral and cellular immune responses against PCV2. This review focuses on the recent research on immunological aspects during PCV2 infections and summarizes what is currently known about the vaccine-induced immunity. (C) 2010 Elsevier B.V. All rights reserved.

An anthrax subunit vaccine candidate based on protective regions of Bacillus anthracis protective antigen and lethal factor

Studies have confirmed the key role of Bacillus anthracis protective antigen (PA) in the US and UK human anthrax vaccines. However, given the tripartite nature of the toxin, other components, including lethal factor (LF), are also likely to contribute to protection. We examined the antibody and T cell responses to PA and LF in human volunteers immunized with the UK anthrax vaccine (AVP). Individual LF domains were assessed for immunogenicity in mice when given alone or with PA. Based on the results obtained, a novel fusion protein comprising D1 of LF and the host cell-binding domain of PA (D4) was assessed for protective efficacy. Murine protection studies demonstrated that both full-length LF and D1 of LF conferred complete protection against a lethal intraperitoneal challenge with B. anthracis STI spores. Subsequent studies with the LFD1-PAD4 fusion protein showed a similar level of protection. LF is immunogenic in humans and is likely to contribute to the protection stimulated by AVP. A single vaccine comprising protective regions from LF and PA would simplify production and confer a broader spectrum of protection than that seen with PA alone.

Nasal immunization with Burkholderia multivorans outer membrane proteins and the mucosal adjuvant adamantylamide dipeptide confers efficient protection against experimental lung infections with B. multivorans and B. cenocepacia

Chronic lung infection by opportunistic pathogens, such as Pseudomonas aeruginosa and members of the Burkholderia cepacia complex, is a major cause of morbidity and mortality in patients with cystic fibrosis. Outer membrane proteins (OMPs) of gram-negative bacteria are promising vaccine antigen candidates. In this study, we evaluated the immunogenicity, protection, and cross-protection conferred by intranasal vaccination of mice with OMPs from B. multivorans plus the mucosal adjuvant adamantylamide dipeptide (AdDP). Robust mucosal and systemic immune responses were stimulated by vaccination of naive animals with OMPs from B. multivorans and B. cenocepacia plus AdDP. Using a mouse model of chronic pulmonary infection, we observed enhanced clearance of B. multivorans from the lungs of vaccinated animals, which correlated with OMP-specific secretory immunoglobulin A responses. Furthermore, OMP-immunized mice showed rapid resolution of the pulmonary infection with virtually no lung pathology after bacterial challenge with B. multivorans. In addition, we demonstrated that administration of B. multivorans OMP vaccine conferred protection against B. cenocepacia challenge in this mouse infection model, suggesting that OMPs provide cross-protection against the B. cepacia complex. Therefore, we concluded that mucosal immunity to B. multivorans elicited by intranasal vaccination with OMPs plus AdDP could prevent early steps of colonization and infection with B. multivorans and also ameliorate lung tissue damage, while eliciting cross-protection against B. cenocepacia. These results support the notion that therapies leading to increased mucosal immunity in the airways may help patients with cystic fibrosis.