In planta expression of nanobody-based designer chicken antibodies targeting Campylobacter

Abstract

Campylobacteriosis is a widespread infectious disease, leading to a major health and economic burden. Chickens are considered as the most common infection source for humans. Campylobacter mainly multiplies in the mucus layer of their caeca. No effective control measures are currently available, but passive immunisation of chickens with pathogen-specific maternal IgY antibodies, present in egg yolk of immunised chickens, reduces Campylobacter colonisation. To explore this strategy further, anti-Campylobacter nanobodies, directed against the flagella and major outer membrane proteins, were fused to the constant domains of chicken IgA and IgY, combining the benefits of nanobodies and the effector functions of the Fc-domains. The designer chimeric antibodies were effectively produced in leaves of Nicotiana benthamiana and seeds of Arabidopsis thaliana. Stable expression of the chimeric antibodies in seeds resulted in production levels between 1% and 8% of the total soluble protein. These in planta produced antibodies do not only bind to their purified antigens but also to Campylobacter bacterial cells. In addition, the anti-flagellin chimeric antibodies are reducing the motility of Campylobacter bacteria. These antibody-containing Arabidopsis seeds can be tested for oral passive immunisation of chickens and, if effective, the chimeric antibodies can be produced in crop seeds.

New and alternative strategies for the prevention, control, and treatment of antibiotic-resistant Campylobacter

“…Recently, the same team made chimeric antibodies by fusing nanobodies recognizing Campylobacter MOMP and flagellin with the constant domains of IgY and IgA of chicken, and successfully expressed the chimeric antibodies in plant leaves and seeds. 136 The plant produced antibodies showed binding activities to native MOMP and intact Campylobacter cells, and the plant-derived flagellinspecific antibodies reduced the motility of Campylobacter. These results demonstrate potential use of genetically engineered nanobodies for control of Campylobacter infection.…”

A Rationally Designed Bovine IgA Fc Scaffold Enhances in planta Accumulation of a VHH-Fc Fusion Without Compromising Binding to Enterohemorrhagic E. coli

“…Despite the therapeutic value of IgA, transition to clinical testing has been hampered by technical difficulties in producing and purifying the full-size IgA in conventional microbial and mammalian platforms because of native IgA's large size (380 kDa; compared to 150 kDa for IgG) (Reinhart and Kunert, 2015). There are several reports of the production of IgA in plants and in yeast intended for oral passive enteromucosal immunization of animals (Virdi et al, 2013(Virdi et al, , 2019Vanmarsenille et al, 2018;Saberianfar et al, 2019). While accumulation of IgA in some of those reports has met the 1% TSP/100 mg/kg FW/100 mg/L culture medium rule of thumb benchmark for commercial viability (Rybicki, 2009;Schillberg et al, 2019), an improvement in accumulation levels by way of stabilization would reduce the cost of production.…”

Applications of nanobodies in plant science and biotechnology

“…Since the plant expression of SV H H-IgA still involves introduction of three genes (including V H H-IgA, J-chain, and secretory component) which is complex and time-consuming, the authors further investigated the simplified single-gene-encoded monomeric V H H-IgA (MV H H-IgA) and found it to be effective as well in preventing ETEC infection (Virdi et al 2019 ). Similar strategies have been successfully adopted to express nanobodies directed against pathogens in plant tissues to protect livestock and poultry from enteric diseases (Harmsen et al 2005 ; Saberianfar et al 2019 ; Vanmarsenille et al 2018 ). Besides, several nanobodies against toxins (Barrera et al 2015 ; Richard et al 2013 ), proteins involved in tumorigenesis (Ismaili et al 2007 ; Mirzaee et al 2018 ; Modarresi et al 2018 ; Park et al 2020 ; Rajabi-Memari et al 2006 ; Soleimanizadeh et al 2018 ), or key regulators of inflammatory diseases (Abe et al 2014 ; Conrad et al 2011 ; Giersberg et al 2010 ; Winichayakul et al 2009 ) have been produced successfully in plants, and are proved to be effective.…”