Freeze-drying of ovalbumin loaded mesoporous silica nanoparticle vaccine formulation increases antigen stability under ambient conditions

Amino functionalised mesoporous silica nanoparticles (AM-41) have been identified as a promising vaccine delivery material. The capacity of AM-41 to stabilise vaccine components at ambient temperature (23–27 °C) was determined by adsorbing the model antigen ovalbumin (OVA) to AM-41 particles (OVA-41). The OVA-41 was successfully freeze-dried using the excipients 5% trehalose and 1% PEG8000. Both the immunological activity of OVA and the nanoparticle structure were maintained following two months storage at ambient temperature. The results of immunisation studies in mice with reconstituted OVA-41 demonstrated the induction of humoral and cell-meditated immune responses. The capacity of AM-41 particles to facilitate ambient storage of vaccine components without loss of immunological potency will underpin the further development of this promising vaccine delivery platform.

Evaluation of the immunogenicity of the P97R1 adhesin of Mycoplasma hyopneumoniae as a mucosal vaccine in mice.

The immunogenicity of P97 adhesin repeat region R1 (P97R1) of Mycoplasma hyopneumoniae, an important pathogenesis-associated region of P97, was evaluated in mice as a mucosal vaccine. Mice were immunized orally with attenuated Salmonella typhimurium aroA strain CS332 harbouring a eukaryotic or prokaryotic expression vector encoding IP97R1. Local and systemic immune responses were analysed by ELISA on mouse sera, lung washes and splenocyte supernatants following splenocyte stimulation with specific antigens in vitro. Although no P97R1-specific antibody responses were detected in serum and lung washes, significant gamma interferon was produced by P97R1-stimulated splenocytes from mice immunized orally with S. typhimurium aroA harbouring either expression system, indicating induction of a cell-mediated immune response. These results suggested that live bacterial vectors carrying DNA vaccines or expressing heterologous antigens preferentially induce a Th1 response. Surprisingly, however, mice immunized with the vaccine carrier S. typhimurium aroA CS332 induced serum IgG, but not mucosal IgA, against P97R1 or S. typhimurium aroA CS332 whole-cell lysate, emphasizing the importance of assessing the suitability of attenuated S. typhimurium antigen-carrier delivery vectors in the mouse model prior to their evaluation as potential vaccines in the target species, which in this instance was pigs.

Producer Demonstration Sites (PDS) coordination and delivery

Producer Demonstration Site (PDS) - Funding to accommodate the establishment of 14 PDS sites over 3 years. Implementation of PDS to increase research adoption and practice change in Queensland.

Spatially based data delivery and recording sys-Aust Sugar Cane industry

Establish an internet platform where spatially referenced data can be viewed, entered and stored.

Internet based spatial data delivery and reporting system for cotton industry

Development of an internet based spatial data delivery and reporting system for the Australian Cotton Industry.

Continued delivery of applied solutions to weed issues in central Queensland

Developing best practices in Central Queensland to (a) manage difficult to control weeds; (b) improve herbicide efficacy under adverse conditions, and (c) manage weeds in wide-row crop systems.

Efficient organogenesis of an Australian passionfruit hybrid (Passiflora edulis x Passiflora edulis var. flavicarpa) suitable for gene delivery

An efficient regeneration protocol based on organogenesis from cotyledon explants and suitable for gene delivery has been developed for an Australian passionfruit hybrid. Multiple shoots were regenerated from 30-day-old cotyledon explants on Murashige and Skoog (MS) medium containing 6-benzylvaminopurine (BAP) and coconut water. Media pulsing experiments were conducted to investigate the effect on organogenesis of exposure time of the explants to MS containing 10 mu M BAP and 10% (v/v) coconut water, i.e. passionfruit regeneration medium (PRM). Continuous exposure of these explants to PRM maximised the number of shoots produced to 12.1 per explant. However, periods on hormone-free medium improved the appearance of the shoots and increased the number of explants with shoots from 75 to 84.6%. Further, shoots exposed for 7 days to half-strength MS supplemented with 10 mu M NAA (1-napthalene acetic acid) produced twice as many plantlets than those on half-strength MS alone. Transient GUS histochemical assays indicated delivery of the uidA gene via Agrobacterium tumefaciens.

In vivo delivery of bovine viral diahorrea virus, E2 protein using hollow mesoporous silica nanoparticles

Our work focuses on the application of mesoporous silica nanoparticles as a combined delivery vehicle and adjuvant for vaccine applications. Here we present results using the viral protein, E2, from bovine viral diarrhoea virus (BVDV). BVDV infection occurs in the target species of cattle and sheep herds worldwide and is therefore of economic importance. E2 is a major immunogenic determinant of BVDV and is an ideal candidate for the development of a subunit based nanovaccine using mesoporous silica nanoparticles. Hollow type mesoporous silica nanoparticles with surface amino functionalisation (termed HMSA) were characterised and assessed for adsorption and desorption of E2. A codon-optimised version of the E2 protein (termed Opti-E2) was produced in Escherichia coli. HMSA (120 nm) had an adsorption capacity of 80 [small mu ]g Opti-E2 per mg HMSA and once bound E2 did not dissociate from the HMSA. Immunisation studies in mice with a 20 [small mu ]g dose of E2 adsorbed to 250 [small mu ]g HMSA was compared to immunisation with Opti-E2 (50 [small mu ]g) together with the traditional adjuvant Quillaja saponaria Molina tree saponins (QuilA, 10 [small mu ]g). The humoral responses with the Opti-E2/HMSA nanovaccine although slightly lower than those obtained for the Opti-E2 + QuilA group demonstrated that HMSA particles are an effective adjuvant that stimulated E2-specific antibody responses. Importantly the cell-mediated immune responses were consistently high in all mice immunised with Opti-E2/HMSA nanovaccine formulation. Therefore we have shown the Opti-E2/HMSA nanoformulation acts as an excellent adjuvant that gives both T-helper 1 and T-helper 2 mediated responses in a small animal model. This study has provided proof-of-concept towards the development of an E2 subunit nanoparticle based vaccine.

Silica vesicles as nanocarriers and adjuvants for generating both antibody and T-cell mediated immune resposes to Bovine Viral Diarrhoea Virus E2 protein

Bovine Viral Diarrhoea Virus (BVDV) is widely distributed in cattle industries and causes significant economic losses worldwide annually. A limiting factor in the development of subunit vaccines for BVDV is the need to elicit both antibody and T-cell-mediated immunity as well as addressing the toxicity of adjuvants. In this study, we have prepared novel silica vesicles (SV) as the new generation antigen carriers and adjuvants. With small particle size of 50 nm, thin wall (similar to 6 nm), large cavity (similar to 40 nm) and large entrance size (5.9 nm for SV-100 and 16 nm for SV-140), the SV showed high loading capacity (similar to 250 mu g/mg) and controlled release of codon-optimised E2 (oE2) protein, a major immunogenic determinant of BVDV. The in vivo functionality of the system was validated in mice immunisation trials comparing oE2 plus Quil A (50 mu g of oE2 plus 10 mu g of Quil A, a conventional adjuvant) to the oE2/SV-140 (50 mu g of oE2 adsorbed to 250 mu g of SV-140) or oE2/SV-140 together with 10 mu g of Quil A. Compared to the oE2 plus Quil A, which generated BVDV specific antibody responses at a titre of 10(4), the oE2/SV-140 group induced a 10 times higher antibody response. In addition, the cell-mediated response, which is essential to recognise and eliminate the invading pathogens, was also found to be higher [1954-2628 spot forming units (SFU)/million cells] in mice immunised with oE2/SV-140 in comparison to oE2 plus Quil A (512-1369 SFU/million cells). Our study has demonstrated that SV can be used as the next-generation nanocarriers and adjuvants for enhanced veterinary vaccine delivery. (C) 2014 Elsevier Ltd. All rights reserved.

Synthesis of Silica Vesicles with Controlled Entrance Size for High Loading, Sustained Release, and Cellular Delivery of Therapeutical Proteins

A rationally designed two-step synthesis of silica vesicles is developed with the formation of vesicular structure in the first step and fine control over the entrance size by tuning the temperature in the second step. The silica vesicles have a uniform size of ≈50 nm with excellent cellular uptake performance. When the entrance size is equal to the wall thickness, silica vesicles after hydrophobic modification show the highest loading amount (563 mg/g) towards Ribonuclease A with a sustained release behavior. Consequently, the silica vesicles are excellent nano-carriers for cellular delivery applications of therapeutical biomolecules.