Evaluation of inhibitor activity of bacterial sialidase from Clostridium perfringens against Newcastle disease virus in the cell culture model using chicken embryo fibroblast

Abstract

Objective: The Newcastle disease virus (NDV) is an infectious disease that causes very high eco¬nomic losses due to decreased livestock production and poultry deaths. The vaccine’s ineffec¬tiveness due to mutation of the genetic structure of the virus impacts obstacles in controlling the disease, especially in some endemic areas. This study aimed to provide an alternative treatment for NDV infection by observing the viral replication inhibitor activity of Clostridium perfringens sialidase in primary chicken embryo fibroblast (CEF) cells. Materials and Methods: The virus was adapted in CEF monolayer cells, then collected thrice using the freeze–thaw method and stored at −20°C for the next step in the challenge procedure. C. perfringens crude sialidase was obtained, but it was further purified via stepwise elution in ion exchange using Q Sepharose® Fast Flow and affinity chromatography with oxamic acid agarose. The purified sialidase was tested for its toxicity, ability to breakdown sialic acid, stopping viral replication, and how treated cells expressed their genes. Results: According to this study, purified C. perfringens sialidase at dosages of 187.5, 93.75, and 46.87 mU effectively hydrolyzes CEF cells’ sialic acid and significantly inhibits viral replication on the treated cells. However, sialidase dosages of 375 and 750 mU affected the viability of mono¬layer CEF cells. Interestingly, downregulation of toll-like receptor (TLR)3 and TLR7 (p < 0.05) in the sialidase-treated group indicates viral endocytosis failure. Conclusions: By stopping endocytosis and viral replication in host cells, sialidase from C. perfrin¬gens can be used as an alternative preventive treatment for NDV infection.

Clostridium perfringens Sialidase Interaction with Neu5Ac α-Gal Sialic Acid Receptors by In- Silico Observation and Its Impact on Monolayers Cellular Behaviour Structure

“…It demonstrates that removing of sialic acid from the cell surface due to sialidase able to cause slight changes in the pattern of glycosylation which are known to cause dramatic changes in cellular behavior. 14,33,34 In this study, the CEF cells appearance was characterized by an increase in the empty space area between cells which was most likely due to the loss of sialic acid on the cell surface. On the other hand, compared to the control group, administration of sialidase at a low dose of 30 mU did not cause a signi cant increase in perimeter and area or changes in cell structure.…”

“…Puri ed sialidase enzyme activity was observed using the Neuraminidase assay kit MAK121 (Sigma-Aldrich) with appropriate protocol procedures to obtain a quantitative value in U/ml. 13,14 Chicken Embryo Fibroblast Cell Culture Treatment By Sialidase CEF monolayer cultured cells were obtained from SPF embryonated chicken eggs (ValoBioMedia GmbH, German) age of 8-10 days. Embryo was taken and rinsed brie y with Phosphate Buffered Saline (PBS).…”

“…Further observations were made by observing in addition to microscopic observations of cell structure behavior as an indicator of cell response. 14 \varvec℃…”

“…Bacterial sialidase can catalyze the hydrolysis of terminal sialic acid linked by α (2,3) -, α (2,6) -, or α (2,8) bonds to a wide variety of substrates. [11][12][13][14] In addition, some of these enzymes can catalyze the transfer of sialic acid from sialoglycans to asialoglycoconjugates by means of the transglycosylation reaction mechanism. 15 Research regarding to the inhibition of viral infection by sialidase has been carried out in vitro using cell culture and in vivo by animal models.…”

In Vitro Antiviral Activity of NanB Bacterial Sialidase Against Avian Influenza H9N2 Virus in MDCK cells

“…The spectrophotometer was used to measure absorbance readings at a wavelength of 405 nm. (Kurnia et al 2022a;Nugroho et al 2022) The effect of sialidase on viral replication was evaluated by exposing MDCK cells, previously treated with the enzyme and subsequently challenged with the H9N2 virus to evaluate the effectiveness of sialidase in hindering viral replication. Monolayer MDCK cells on 96-well plates.…”