Development of gold nanoparticles-lateral flow test as a novel field diagnostic assay for detecting foot-and-mouth disease and lumpy skin disease viruses

Abstract

Background and objectives: Rapid diagnosis is a cornerstone for controlling and preventing viral disease outbreaks. The present study is aimed to develop a rapid field diagnostic test based on gold nanoparticles for the detection of lumpy skin diseases (LSD), and foot and mouth diseases (FMD) in animals with high sensitivity and specificity.

Materials and methods: FMD and LSD vaccines were used as a source of viruses' antigens for preparing monoclonal antibodies and conjugated with gold nanoparticles that characterized using various techniques such as UV-visible spectrometry, and transmission electron microscopy (TEM). Monoclonal antibodies (mAbs) for each serotype produced in experimental rats and used to capture antibodies for FMDV and/or LSDV. ELISA was used to screen 469 milk samples and 1165 serum samples from naturally infected cattle, buffaloes, sheep, and goats for validation of the lateral flow test (LFT). LSDV DNA was extracted from 117 blood and skin biopsy samples collected from naturally infected cattle during the 2019 outbreak.

Results: The specificity and sensitivity of GNP-LFT were evaluated and compared to Ag-ELISA, Western blot tests (WB), and PCR. A total of 95 FMDV positives out of 469 (20.25%) milk samples and 268 FMDV positives out of 1165 (23.3%) serum samples from natural infected cattle, buffaloes, sheep, and goats examined by ELISA to valid GNPS-LFT Viral LSDV DNA was detected in 60/117 (51.5%) and 31/60 (52.9%). While the GNPS-LFT assay results were 49/117 (41.9%) and 29/60 (48.3%) blood and skin biopsy samples, respectively. The diagnostic sensitivity and specificity of the GNP-LFT test were 72% and 82%, respectively. All vesicular fluid and epithelium samples collected from infected animals were identified as positive by the GNP-LFT and Ag-ELISA. Ag-ELISA, on the other hand, was 90% and 100%. While the developed GNP-LFT used LSDV polyclonal antibodies were similar to ELISA and IgG-WB with a sensitivity of 72.8% and a specificity of 88.8%, respectively.

Conclusion: The GNPS-LFT is a novel immunoassay based on mono or polyclonal antibodies conjugated with gold nanoparticles that provides an accurate, rapid, specific, and sensitive tool for field rapid diagnosis of FMDV and LSDV.

Keywords: Foot-mouth disease; Gold nanoparticles; Lateral flow test; Lumpy skin disease virus; Rapid diagnostic tests; Skin lesion biopsy.

Fig. 1.

(A, B and D): Electron microscopy micrograph of gold nanoparticles was showed (spherical particles with particle size diameter < 30 nm, characterized by TEM and UV–Vis spectroscopy showed 10 to 20 nm sizes with maximum absorption at 520 nm, and larger size with Mabs conjugation, the size range from 25–30 nm the color intensity (wine red).

Fig. 2.

3ABC-ELISA: WB: Western blot test, GNPs-LFT, IC: immune-chromogenic test 3ABC-enzyme-linked immunosorbent assay

Fig. 3.

Detection of LSDV by different molecular assays in whole blood and skin biopsy samples collected from cattle in different governorates in Egypt.