Nano silver is used in veterinary virus vaccines (IBR/IPB, RVFV, H5N1, H5N2, Newcastle Disease Virus (NDV), IBDV, FPV)
Traditional veterinary viral vaccines, such as inactivated vaccines and live attenuated vaccines, have achieved tremendous success in controlling many viral diseases in livestock and chickens worldwide. world. However, many recent viral outbreaks due to various emerging and re-emerging viruses continue to be reported annually worldwide. Therefore, it is necessary to develop new control regimens. Nanoparticle research has received significant attention over the past two decades as a promising platform with significant successes in veterinary medicine, replacing traditional viral vector vaccines. However, the field of nanoparticle applications is still in the early stages of development. Here, we discuss the different preparation methods, characteristics, physical properties, antiviral effects and pharmacokinetics of well-developed nanoparticles as well as the potential of nanoparticles or nano silver vaccine as a promising antiviral platform for veterinary medicine.
introduce
Nanotechnology has been a rapidly growing field since 1974 and has led to the development of many new nanoparticles with average diameters ranging from 1 to 100 nanometers (nm) [77, 79]. The prefix nano is derived from the Latin word “nanus” meaning “very small”, as 1 nm corresponds to 10 -9 meters (m) [ 77 ].
Currently, nanotechnology is being applied in many different fields, including agriculture, infection control [ 80 ], and biomedicine [ 10 , 69 ]. Nanoparticles have several physical and biological properties, such as large surface area, improved reactivity properties, enormous size-to-volume ratio, durability, bioactivity, bioavailability, regulated particle length, regulated drug release, site-specific targeting, and regulated drug delivery [ 49 ]. Furthermore, nanoparticles can penetrate cells, tissues, and organs, making them effective drug delivery tools [ 18 ]. Various medicinal products can also be attached to the surface of nanoparticles [ 57 , 69 ]. To overcome difficult problems, traditional treatments may not be sufficient and new approaches need to be considered, which may provide future findings and criteria for these problems. current issue [ 88 ].
The economies of many countries rely on animal-based industries, and with the emergence of many viral diseases, new disease prevention and control protocols are urgently needed [ 67 ). Nanotechnology has shown incredible potential in enhancing the delivery of drugs and vaccines in the veterinary sector [ 10 ]. The growing growth of the nanoparticle field will lead to the development of new therapeutics to cure viral or bacterial infections, as well as enhance the healing of deep wounds.
Additionally, these newly developed nanoparticles can successfully deliver drugs to different cells to treat diseases [ 15 , 50 ]. Another incredible development in nanotechnology is nanotherapeutics, a medical technique that integrates drugs and diagnostics that aims to enhance the effectiveness of currently used drugs. Furthermore, this integration offers great opportunities to improve and design these agents, allowing for treatment delivery as well as detection methods before and during treatment [ 47 , 66 ]. One of the most active and encouraging areas of nanotechnology is nano pharmaceutical products, which have many advantages in veterinary medicine [ 59 , 100 ].
Characterization of nanoparticles
Nanoparticle properties depend on measured parameters such as morphology, particle size, surface hydrophobicity and surface charge. Advanced techniques, such as transmission electron microscopy (TEM), atomic force microscopy (AFM) and scanning electron microscopy (SEM), Dynamic Light Scattering (DLS) can used to measure particle size, morphology and corresponding particle size distribution. Common nanoparticle characterization methods are summarized and listed in Table 2. The surface charge of nanoparticles has a significant impact on the physical stability and performance of the polymer. Therefore, the zeta potential technique is widely used as a tool to indirectly measure the surface charge of nanoparticles. It can also be used to evaluate the surface hydrophobicity and nature of materials encapsulated inside nanocapsules or coated on their surfaces [ 74 ]. On the other hand, several techniques have been used over the past decade to measure the surface hydrophobicity of nanoparticles, including hydrophobic interaction chromatography. Modern techniques such as X-ray photon correlation spectroscopy allow the identification of specific chemical groups on the nanoparticle surface, as well as the determination of nanoparticle hydrophobicity [ 101 ].
Additionally, several techniques have been used to determine drug quantity and drug release, such as high-performance liquid chromatography (HPLC) or ultraviolet (UV) spectroscopy. The HPLC method is used to measure the loading capacity of nanoparticle conjugated drugs, which can be expressed as moles of drug per mg of polymer, mg of drug per mg of polymer, or as a percentage relative to the polymer [ 25 , 48 , 77 ].
Nanoparticle preparation
The fabrication of nanoparticles is often based on the chemical and physical properties of drugs and polymers. Nanoparticles can be made from a variety of materials, including synthetic polymers, polysaccharides, and proteins. However, several factors need to be considered during the selection of polymers to use for drug delivery, such as toxicity, nanoparticle size, antigenicity of the final product, surface charge, hydrophobicity, biocompatibility and biodegradability [ 5 , 83 ]. As discussed below, nanoparticles are commonly prepared by emulsification, ionic gelation, and polymerization.
Emulsion method
The dispersion of synthetic polymers with the drug under investigation is the basis of this process [ 17 ]. The size of nanoparticles is influenced by polymer concentration, type and concentration of stabilizer, and stirring speed during preparation [ 93 ]. This method can be used to prepare lipophilic drugs with the flexibility of being able to be combined with different transformation methods for their preparation. This method can be modified to change the properties of nanoparticles or create suitable conditions for hydrophilic drugs [ 4 ]. These different conversion methods can include spontaneous emulsification to form oil-in-water-in-oil emulsions [ 35 ] or double emulsions combined with evaporation [ 58 ]. Another method, called saltation, involves dissolving the drug and polymer in a water-miscible solvent. This process can be performed at room temperature and is especially useful for preparing heat-sensitive materials.
Gel formation or ion coagulation method
This method is based on the fabrication of nanoparticles by mixing oppositely charged particles [ 99 ]. It is also suitable for the preparation of hydrophilic polymer-based nanoparticles. Furthermore, strong electrostatic interactions between the two aqueous phases contribute to the formation of precipitates by this method [ 72 ].
Coincidence method
In this method, nanoparticle molecules are chemically produced in the presence of an aqueous medium. The candidate drug is then added to the polymerization medium or adsorbed onto the nanoparticles after completion of the polymerization process. The polymerization process uses various stabilizers and surfactants, which are typically removed in an ultracentrifugation step, followed by resuspension of the particles in a surfactant-free medium. The desired nano capsule size can be achieved by optimizing the surfactant and stabilizer concentrations [33 , 96].
Nano vaccines and antiviral nanoparticles in veterinary medicine
Efficacy of nanoparticles against livestock viruses
Foot-and-mouth disease virus (FMDV) is an acute infectious disease caused by foot-and-mouth disease virus (FMDV), an RNA virus of the Picornaviridae family. FMDV causes disease in cows, sheep, goats, pigs, deer, and other animals with forked hooves [ 11 , 24 , 43 , 68 , 70 ]. Inactivated FMDV vaccines have been a proven part of best practices for prevention and control since the 1990s. However, the potential for virus escape from manufacturing facilities can cause causing unpredictable spread of the disease [ 86 ].
Many studies have shown that gold nanoparticles can be an excellent adjuvant in combination with current FMDV vaccines as they can also stimulate the nuclear factor kappa-light signaling pathway β-chain of activated B cells (NF-κB) and production of specific cytokines and cytotoxic T cells [ 98 ]. According to a recent study, the combination of synthetic gold star nanoparticles (AuSN) with FMDV-like particles (VLP) led to the formation of non-toxic VLP-AuSN complexes in different cell lines tested. Furthermore, a detailed mechanistic analysis showed that AuSN could effectively promote FMDV VLP entry into cells and improve macrophage activation when compared with FMD VLP alone [ 98 ]. . Furthermore, the protection rate in the AuSN-adjuvanted group was found to be significantly higher after viral challenge than in the conventional mineral oil-adjuvanted group (ISA206). This is very promising because in the future we may be able to use lower doses of nanovaccines to fight FMD, thereby reducing production costs and facilitating rapid and widespread distribution to countries. different.
Another group reported that injection of gold nanoparticles combined with a synthetic VP1 peptide corresponding to the Capsid protein of FMDV with complete Freund’s adjuvant resulted in maximal antibody production in guinea pigs, hyperplasia Gamma interferon (IFN-γ) production and increased peritoneal macrophage activity. Interestingly, in the same study, the use of gold nanoparticles as hapten carriers increased the immune response even without the full administration of Freund’s adjuvant [ 26 ].
Rift Valley fever virus (RVFV) is a mosquito-borne virus that causes devastating disease in ruminants and can be transmitted to humans. In humans, RVFV causes influenza-like illness, but it can also lead to a more complex situation with high morbidity and mortality [ 44 ]. Currently, there are no RVFV vaccines licensed for human use. Therefore, effective treatment is essential. Nano silver have long been reported to have potent antiviral activity against many viruses belonging to different families [ 82 ].
A recent report showed the potential application of Nano silver to control RVFV in which silver nanoparticles were formulated as Argovit™ [ 13 ]. The antiviral activity of Argovit was evaluated in two ways: in vitro on Vero cells and in vivo in type I-interferon receptor-deficient mice. First, different concentrations of Argovit were added to cells previously infected with RVFV or injected into RVFV-infected animals at a lethal dose. Second, RVFV was preincubated with different concentrations of Argovit before inoculation into mice and/or Vero cells. Argovit’s ability to control RVFV infection is limited. However, incubation of the virus with Argovit before infection resulted in a significant reduction in RVFV infectivity in both in vivo and in vitro experiments [ 13 ].
Bovine herpes virus Infectious bovine rhinotracheitis/infectious pustular balanitis (IBR/IPB) is a highly contagious viral disease caused by bovine herpesvirus type 1 (BoHV-1), a DNA virus caused by double strands of the Herpesviridae family. This virus infects cattle worldwide, leading to significant economic losses [ 56 ]. A recent study showed that silver nanoparticles (Ag-NPs) at a dose of 24 μg/mL could moderately inhibit viral infection in MDBK cells [ 28 ].
Peste des petits ruminant virus (PPRV) is a highly contagious transboundary viral disease that mainly affects sheep and goats. PPR is endemic in Egypt, causing major economic losses as well as high morbidity and mortality rates (up to 100%) in affected flocks [ 30 ]. The disease is caused by PPRV, a single-stranded negative-sense RNA virus belonging to the genus Morbillivirus , subfamily Orthoparamyxovirinae , family Paramyxoviridae [ 85 ]. The PPRV vaccine currently on the market is an attenuated live cell culture vaccine but has not shown success in controlling the epidemic worldwide due to insufficient coverage and unstable vaccines (especially is in subtropical countries), low protection during epidemics and poor cross-breeding. protection between PPRV strains circulating in the field and vaccine strains [ 64 ]. A study reported the in vitro activity of nano silver (SNPs) against PPRV infection in Vero cells, in which nano silver significantly inhibited virus entry at a minimum inhibitory concentration of 11.11 µg/ml by interacting with the virion surface and core, but they do not exert a direct virucidal effect on cell-free virions. Nano silver showed higher stability after storage at 37°C for seven days [ 61 ].
Efficacy of nanoparticles against avian viruses
Avian influenza virus (AIV) is a highly contagious virus that causes significant morbidity and mortality in poultry populations, and some strains can pose a pandemic threat to humans [ 34 , ninety four ]. Despite the widespread use of several inactivated AIV vaccines, they have proven ineffective, requiring the development of new technology to improve immunity and enhance their effectiveness. A recent study showed that chimeric vaccine antigen H5 (H5M) combined with polyanhydride nanoparticles (PAN) resulted in sustained release of encapsulated antigens [ 62 ]. Furthermore, this vaccine candidate is immunogenic when encapsulated in PAN and/or delivered using a modified vaccinia Ankara (MVA) vector. Interestingly, both platforms (MVA vector and PAN packaging) induced humoral and cellular immunity in specific pathogen-free (SPF) and commercial chicks. Additionally, protective levels of antibodies were elicited against highly pathogenic avian influenza (HPAI) caused by homologous H5N1 and heterologous H5N2 strains. However, little is known about the toxicological properties of silver nanoparticles in vivo in avian and/or livestock species. Biological effects may vary depending on the animal species studied, age, sex and other factors, including the physical properties of the silver nanoparticles used as well as the dose and route of administration. and delivery time [ 7 , 103 ].
Newcastle disease virus (NDV) is one of the most important viral diseases in poultry in terms of global distribution and causes heavy economic losses. ND is caused by NDV, which belongs to the genus Orthoavulavirus , subfamily Avulavirinae and family Paramyxoviridae [ 85 ]. An intensive NDV vaccination program in Egypt using traditional live attenuated and inactivated NDV vaccines has been unsuccessful and outbreaks continue to be reported due to new and highly virulent virus strains. induced floating [ 45 ].
A previous report showed that polyrhodanine nanoparticles had potent anti-NDV activity in eggs, suggesting that this non-toxic material could be used to control NDV in chickens, as it reduced the egg infectious dose by 50%. EID 50) of NDV strains. isolated from the outbreak in Tehran, Iran, in 2009 [ 75 ]. Interestingly, egg embryos injected with 0.1, 1, 10, and 100 parts per million (ppm) polyrhodanine showed no pathological tissue damage, abnormalities, or deformities, and there were also no changes in blood biochemical parameters. blood serum [ 75 ].
Another interesting study showed that microalgae-mediated nano silver (AgNPs) had significant in vitro antiviral activity against NDV infection in Huh7 cells [ 60 ]. Furthermore, microalgae extract has significant activity against NDV with an unclear mechanism of action, but it appears to be through inhibition of viral entry into infected cells, as Nano silver interacts directly with NDV envelope glycoprotein. In another study, nanoparticles and polymer-adjuvanted mucosal inactivated vaccines were developed for the prevention of ND and avian influenza (H9N2), administered to SPF chickens by spray or intranasal route. These vaccines significantly increased phagocytic index, interleukin-6 (IL-6) levels, and IFN-γ responses, and they protected chickens against challenge with both viruses. The authors recommended widespread adoption of these vaccines in vaccination strategies against avian influenza subtypes H9N2 and NDV [ 29 ].
Since mucosal immunity plays an important role in protecting against NDV [ 105 , 106 ], a DNA vaccine containing the NDV fusion (F) gene was encapsulated in Ag@SiO2 hollow nanoparticles (pFDNA -Ag@SiO2-NP) or chitosan-coated polymeric nanoparticles (PLGA) showed low toxicity, high stability, and did not destroy the biological activity of plasmid DNA in vitro. Furthermore, intranasal vaccination of chickens with pFDNA-Ag@SiO2-NP induced higher serum anti-NDV IgG and IgA antibody levels, enhanced lymphocyte proliferation, and promoted the expression IL-2, IL-4 and IFN-γ [ 108 ]. Further studies are needed to develop NDV mucosal vaccines incorporated in nanoparticles, as they are considered safe and effective carriers for NDV-DNA vaccines.
In other studies, the efficacy, stability, and safety of a live NDV vaccine (LaSota strain) encapsulated in chitosan nanoparticles were evaluated [ 19 , 104 ]. The packaged vaccine was found to be safe and highly stable, and after viral challenge, vaccinated chickens were vaccinated orally and/or intranasally with the vaccine. The nanoparticle vaccine provided complete protection, while only partial protection was observed in chickens vaccinated with live LaSota or inactivated NDV. vaccines alone [ 19 , 104 ]. Furthermore, the comparison between the live attenuated NDV and IBV combination vaccines on the market with the live attenuated NDV-IBV vaccine encapsulated in two types of chitosan nanoparticles showed that the chitosan adjuvanted vaccine is safe. , more stable and effective while creating stronger cells and mucosa. immune response protects chickens against challenge with virulent NDV and IBV [ 107 ]. This is promising because the majority of currently approved NDV and IBV vaccines provide partial protection due to an inadequate cellular immune response. Inadequate protection could facilitate the emergence of new virus variants that cause multiple outbreaks, which then lead to shortages in animal protein supplies. The use of these newly developed nanovaccines could help minimize the emergence of new virus variants and reduce the cost of animal protein production.
Infectious bursitis virus (IBDV) is a highly contagious immunosuppressive viral disease that affects chicks between 3 and 6 weeks of age with significant economic impact worldwide [ 89 ]. The disease is caused by IBDV, a non-enveloped, double-stranded RNA virus belonging to the genus Avibirnavirus of the family Birnaviridae [ 84 ]. Current commercial IBDV vaccines are inactivated or attenuated and cause some side effects. On the other hand, IBDV peptide and subunit vaccines are extremely safe but have poor immunogenicity [ 92 ]. Therefore, there is an urgent need to develop new, more potent vaccines to control IBDV infection. Interestingly, a research group noted a significant increase in both humoral and cellular immune responses in broiler chickens vaccinated with PLGA nanoparticles when compared with chickens vaccinated with traditional IBDV [ 3 ]. Another study showed that graphene oxide (GO) sheets and silver nanoparticle-anchored graphene oxide sheets (GO-Ag) have antiviral effects against non-enveloped IBDV and enveloped feline coronavirus (FCoV). [ 16 ]. Interestingly, they found that although GO had no antiviral activity against IBDV, it reduced FCoV infection by 16%, while GO-Ag inhibited IBDV and FCoV infection. 23% and 25%, respectively [ 16 ].
Another study also showed that nano silver has in vivo IBDV prevention and treatment effects using enzyme-linked immunosorbent assay (ELISA) [ 78 ]. The study tested the preventive effect of AgNPs against IBDV by mixing IBDV with nano silver two hours before inoculating the mixture into embryonated eggs, while to test the therapeutic effect, AgNPs were injected 48 hours later. when inoculating the virus into embryonated eggs. Interestingly, nano silver, especially at a concentration of 20 ppm, was effective against IBDV using both methods without significant differences [ 78 ].
Other veterinary viruses
Feline coronavirus (FCoV) is the causative agent of feline infectious peritonitis (FIP) and there is currently no effective vaccine. Diphyllin (a nanoparticle vacuolar ATPase inhibitor) has previously been tested as an antiviral agent against FCoV type II. Interestingly, diphyllin interfered with FCoV replication in fcwf-4 cells by inhibiting endogenous acidification. Diphyllin also showed in vivo efficacy against FCoV when administered intravenously (I/V) to mice and demonstrated high safety [ 53 ]. Another interesting study showed antiviral effects of both CulNP and Nano silver against feline calicivirus (FCV), a surrogate for human norovirus [ 12 , 95 ]. Additionally, polymeric nanoparticles such as PLGA stimulated significant IgA secretion in dairy calves when compared with the commercially available modified live bovine parainfluenza 3 virus (BPI3V) vaccine [ 14 , 71].
Several studies have shown that aluminum-magnesium silicate (AMS) nanoparticles have high in vitro antiviral activity against PPRV [36], canine parvovirus [41], AIV [39], NDV [40], ovulation syndrome virus 76 [ 38 ], IBDV [ 37 ] and fowl pox virus (FPV). In the last case, no hemagglutination activity was observed after virus treatment with AMS NPs [ 42 ]. Several other nano vaccines have been successfully developed against various veterinary viruses, including a polyanhydride-NP-encapsulated mucosal vaccine against respiratory syncytial virus in bovine (BRSV) [ 73 ] and swine influenza virus vaccines encapsulated in polyanhydride NPs for intranasal vaccination of pigs [ 21 ], PLGA NPs [ 22 ] or CS-polymer-based NPs [ 23 ] enhances both humoral and cellular immune responses and protects vaccinated pigs from swine flu virus challenge. Furthermore, pseudorabies virus (a porcine herpes virus) has also been shown to be inhibited by some nanoparticles [ 8 , 46 , 102 ]. Further studies are needed to evaluate the efficacy of previously described antiviral nanoparticles against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which was first reported described in December 2019 in Wuhan City, Hubei Province, China. As of May 16, 2021, more than 136 million cases and 3.38 million deaths have been reported in more than 220 countries and territories worldwide [ 31 , 32 , 51 , 97 ].
CONCLUSION
Previous studies on the development and use of nanoparticles and nano silver vaccines in veterinary medicine have shown significant success over the past decade when compared with traditional vaccines. However, further field studies are needed to study the effects of nano silver vaccines in immunosuppressed animals and determine the optimal application for different animal species.
Resource: Nanoparticles as a novel and promising antiviral platform in veterinary medicine
Mohamed Fawzy, Gasser M. Khairy, Ahmed Hesham, Ali A. Rabaan, Ahmed G. El-Shamy, and Abdou Nagy