Early Stage Development of a Newcastle Disease Vaccine Candidate in Corn

Abstract

Newcastle disease (ND) is a viral disease that causes labored breathing, periorbital oedema, and ataxia in the majority of avian species. The available vaccines against Newcastle disease virus (NDV) are limited, owing to their low reactivity and multiple dosage requirements. Plant-based machinery provides an attractive and safe system for vaccine production. In the current study, we attempted to express fusion (F) and hemagglutinin-neuraminidase (HN) proteins (the protective antigens against NDV) under constitutive 35S and seed-specific Zein promoters, respectively. Almost 2-7.1-fold higher expression of F gene mRNA in transgenic corn leaves and 8-28-fold higher expression of HN gene mRNA in transgenic corn seeds were observed, when the expression was analyzed by real-time PCR on a relative basis as compared to non-transgenic control plant material (Leaves and seeds). Similarly, 1.66 µg/ml of F protein in corn leaves, i.e., 0.5% of total soluble protein, and 2.4 µg/ml of HN protein in corn seed, i.e., 0.8% of total seed protein, were found when calculated through ELISA. Similar levels of immunological response were generated in chicks immunized through injection of E. coli-produced pET F and pET HN protein as in chickens orally fed leaves and seeds of maize with expressed immunogenic protein. Moreover, the detection of anti-NDV antibodies in the sera of chickens that were fed maize with immunogenic protein, and the absence of these antibodies in chickens fed a normal diet, confirmed the specificity of the antibodies generated through feeding, and demonstrated the potential of utilizing plants for producing more vaccine doses, vaccine generation at higher levels and against other infectious diseases.

Producing Vaccines against Enveloped Viruses in Plants: Making the Impossible, Difficult

“…Despite early work showing the viability of displaying NDV epitopes using CMV [ 160 , 161 , 162 ], most research into plant-derived NDV vaccines has focussed on expression of HN or F subunits. Various expression hosts have been explored for NDV vaccine production including potato [ 163 ], rice [ 164 ], maize [ 165 , 166 ], rapeseed [ 167 ], N. benthamiana [ 168 ], N. tabacum [ 169 ], and tobacco cell culture [ 170 ] with varying success. Modifications to constructs, such as the use of appropriate signal peptides and inclusion of ER retention signals, can enhance NDV antigen yield [ 168 ]; however, the highest reported accumulation of NDV antigen stands at 3% of total soluble protein in maize [ 165 ], with more typical yields reported across multiple plant species ranging from 0.18–0.8% of total soluble protein [ 166 , 167 , 170 ] and some yielding below 0.07% [ 169 ].…”

“…As the main target for edible NDV vaccines are farmed poultry, seed-based antigen production has great appeal given both the ease of introduction into the diet via existing feeding methods, and the long-term stability of recombinant antigens in seeds stored under ambient conditions [ 100 ]. For NDV, chicks fed transgenic rapeseed or maize seeds produce antibodies against both HN and F proteins when these are expressed as fused ectodomains [ 167 ] or by co-expression of subunits via a single plasmid [ 166 ]. However, antibody responses tended to be significantly slower and less robust than those stimulated by existing commercial vaccines.…”

Recent Trends in Immunology and Vaccination Against Newcastle Disease Virus

“…Results showed approximately 7 and 28 folds higher expressions of F and HN genes, respectively. Their analysis showed that chicken fed with these transgenic leaves produced specific antibodies against NDV antigen proteins (Shahid et al 2020).…”

Seed-based System for Cost-Effective Production of Vaccine Against Chronic Respiratory Disease in Chickens

“…Till now, variety of vaccines against veterinary diseases have been expressed using plant tissues as bioreactor [5]. The proof-of-concept for such studies has already been accomplished in previous studies and plants have been found suitable host for expression of edible vaccines for Newcastle disease [6], influenza, Ebola, infectious bronchitis virus (IBV), infectious bursal disease virus (IBDV) in chickens [7]. The plant made ZMapp antibodies against Ebola and influenza were not only encouraging but also paved a way for plants to be further utilized as bioreactor to combat infectious diseases in cost effective manner.…”

“…Worldwide, poultry is the largest livestock group, and according to estimates with market value of $309 billion in 2019 and expected to grow nearly 6% annually till 2023 (Poultry world, 2019). The most prevalent problem in poultry especially at large scale setup includes zoonotic diseases which not only is threat to poul-try producers but also is a great risk for poultry genetic resources [6]. Thus, suitable veterinary strategies against such diseases are in crucial need of poultry setups.…”