Nanobodies: As a Versatile Tool for Diagnosis and Effective Management of Foot-and-Mouth Disease
Anu Malik1, Swati Dahiya1, Akhil Kumar Gupta1
1Department of Veterinary Microbiology, LUVAS, Hisar-125001, Haryana
Foot-and-mouth disease (FMD) is a highly contagious, economically devastating viral disease of almost all cloven hoofed animals including both domestic and wild ruminant species. The disease poses a huge threat to ruminant health and causes socio-economic losses to livestock and dairy sector worldwide. During 2013-14, the total loss due to FMD in cattle and buffaloes was estimated to be INR 208.97 billion in India. The disease is caused by FMD virus (FMDV) classified within the genus of Aphthovirus, family Picornaviridae. The genome consists of a single-stranded RNA, ∼8 kb in length, which encodes four structural proteins (VP1, VP2, VP3 and VP4) and a total of ten non-structural proteins (NSPs: Lpro, 2A, 2B, 2C, 3A, 3B1-3, 3Cpro, 3Dpol). The FMDV consists of seven serologically and genetically distinguishable serotypes named O, A, C, Asia-1 and South African Territories (SAT1, SAT2, and SAT3) with multiple subtypes within each serotype. In India, only three serotypes mainly O, A and Asia-1 are found and circulating in livestock population of which serotype O is the most prevalent.
Global FMD control strategy includes reliable and effective surveillance and competent laboratory diagnostic assays. Such diagnosis is typically carried out by the combination of virus isolation, nucleic acid recognition methods and serological tests. Till date numerous diagnostic tests are available for foot and mouth disease diagnosis which include several types of ELISAs as a platform for serological based diagnosis. One such type of ELISA is DIVA that helps to differentiate between infected and vaccinated animals thus contributing to effective control and management of foot-and-mouth disease. However, these diagnostic tests lack in terms of sensitivity, specificity and cost-effectiveness. Also, conventional type of ELISAs used for serotype detection employs use of polyclonal antibody sera raised in animals which may raise the problem of batch-to-batch variability and dependency on animal sacrifice. Nanobodies can be procured easily by cloning from heavy chain antibodies of immunized camel in a simple, inexpensive process that requires 2-6 boosters within 3-6 months. Nbs libraries generated from immunized camels retain full functional diversity unlike conventional antibody production that may yield libraries with reduced diversities due to reshuffling of VL and VH domains during library construction. These advantages favour use of Nanobodies over conventional antibodies.
Camelid derived single domain antibodies or Nanobodies (Nbs) have greater efficacy for serological detection of antibodies against FMD and are emerging novel tools in field of diagnostics. Unlike conventional antibodies, the antigen-binding fragment of these heavy chain antibodies consists of one single variable domain referred to as VHH or Nb. The Nbs are proven to be a valuable tool for diagnostics due to their unique signature physical, biochemical and structural properties. The Nbs are one of the smallest known antigen binding antibody fragments with reduced size, improved solubility, greater thermostability and higher affinity which makes them a powerful tool for diagnostic applications. These Nbs can be produced by cocktail of molecular techniques of cloning and panning by phage display technology. Nbs could be produced at larger scale by their recombinant expression in bacterial host or other eukaryotic expression system for their use in diagnostics and therapeutics.
The development of Nb based ELISA could clearly benefit routine disease diagnosis, the establishment of disease-free zones and the improvement of FMD control and management in endemic areas. The detection of antibodies to viral NSPs can be considered as important indicators of infection irrespective of vaccination status. The use of Nb based competitive ELISA has been reported for the detection of anti-FMDV NSP antibodies in cattle serum. The assay demonstrated high sensitivity and specificity to identify NSP antibodies of several FMD serotype infections with effective, robust performance and potentially low-cost production. The presence of antibodies to FMDV NSP is widely accepted as a reliable indicator of FMD infection in animal herds regardless of their vaccination status and provides critical input for FMD assessment.
Interestingly, Nbs derived from heavy chain-only camelid antibodies have emerged as a new hope in the development of rapid, accurate and cost-effective diagnostic tools in veterinary and biomedical fields that may prove beneficial in near future for the country. Hence, Nbs constitute a promising alternative tool for FMD diagnosis with proper assay validation in upcoming scenario.
https://pubmed.ncbi.nlm.nih.gov/27428417/
