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Review

Revolutionizing Veterinary Vaccines: Overcoming Cold-Chain Barriers Through Thermostable and Novel Delivery Technologies

by
Rabin Raut
1,*,
Roshik Shrestha
2,3,
Ayush Adhikari
4,
Arjmand Fatima
1 and
Muhammad Naeem
1,*
1
Department of Poultry Science, Auburn University, Auburn, AL 36849-5416, USA
2
Livestock Service Branch, Madhyabindu Municipality, Rampur, Bharatpur 13712, Nepal
3
Faculty of Animal Science, Veterinary Science and Fisheries, Agriculture and Forestry University, Rampur, Bharatpur 13712, Nepal
4
Nepal Polytechnic Institute, Bharatpur 13712, Nepal
*
Authors to whom correspondence should be addressed.
Appl. Microbiol. 2025, 5(3), 83; https://doi.org/10.3390/applmicrobiol5030083
Submission received: 23 June 2025 / Revised: 19 July 2025 / Accepted: 8 August 2025 / Published: 19 August 2025
Review Reports Versions Notes

Abstract

Veterinary vaccines are essential tools for controlling infectious and zoonotic diseases, safeguarding animal welfare, and ensuring global food security. However, conventional vaccines are hindered by cold-chain dependence, thermal instability, and logistical challenges, particularly in low- and middle-income countries (LMICs). This review explores next-generation veterinary vaccines, emphasizing innovations in thermostability and delivery platforms to overcome these barriers. Recent advances in vaccine drying technologies, such as lyophilization and spray drying, have improved antigen stability and storage resilience, facilitating effective immunization in remote settings. Additionally, novel delivery systems, including nanoparticle-based formulations, microneedles, and mucosal routes (intranasal, aerosol, and oral), enhance vaccine efficacy, targeting immune responses at mucosal surfaces while minimizing invasiveness and cost. These approaches reduce reliance on cold-chain logistics, improve vaccine uptake, and enable large-scale deployment in field conditions. The integration of thermostable formulations with innovative delivery technologies offers scalable solutions to immunize livestock and aquatic species against major pathogens. Moreover, these strategies contribute significantly to One Health objectives by mitigating zoonotic spillovers, reducing antibiotic reliance, and supporting sustainable development through improved animal productivity. The emerging role of artificial intelligence (AI) in vaccine design—facilitating epitope prediction, formulation optimization, and rapid diagnostics—further accelerates vaccine innovation, particularly in resource-constrained environments. Collectively, the convergence of thermostability, advanced delivery systems, and AI-driven tools represents a transformative shift in veterinary vaccinology, with profound implications for public health, food systems, and global pandemic preparedness.

1. Introduction

Vaccines are fundamental tools for controlling infectious diseases in both humans and animals. In veterinary medicine, immunization has been pivotal in managing diseases caused by pathogens such as infectious bursal disease virus (IBDV), classical swine fever virus (CSFV), and foot-and-mouth disease virus (FMDV) [1,2,3]. The global eradication of the rinderpest virus further exemplifies the transformative impact of veterinary vaccines [2]. Beyond their critical health benefits, vaccines also deliver significant economic advantages by reducing disease-related costs and minimizing hospitalizations. It is estimated that vaccination efforts worldwide have saved over USD 500 billion in human healthcare and related expenditures [3]. In addition to preventing outbreaks of new and exotic diseases, veterinary vaccination programs contribute to zoonotic disease control, safer food production, reduced reliance on antibiotics, and improved health of livestock and companion animals [4]. However, the ongoing emergence of zoonotic diseases in food and companion animals poses serious challenges to global public health. With new outbreaks anticipated in the coming decades, vaccination remains a cornerstone strategy for preventing disease transmission and ensuring the safety of both animal and human populations.
The origins of vaccination can be traced to the pioneering work of English physician and scientist Edward Jenner (1749–1823), who is widely recognized as the father of immunology. Following his development of the smallpox vaccine, the terms “vaccine” and “vaccinology” were introduced [5]. Jenner referred to his smallpox immunization method as variola vaccinae, emphasizing its derivation from cowpox [6]. Building upon Jenner’s foundational contribution, French scientist Louis Pasteur developed the first classical vaccines in the late 1800s, laying the groundwork for modern vaccinology [7]. In 1881, Pasteur extended the use of the term “vaccine” to describe immunogens for diseases beyond smallpox [8]. While the principles of vaccination have remained consistent since their inception, the practical implementation, particularly in the veterinary sector, faces persistent challenges, most notably, the reliance on cold-chain logistics and the thermal sensitivity of conventional vaccines. Most vaccines require a cold-chain system for transportation and storage, typically between 2 and 8 °C, from manufacturing to administration [9]. This requirement poses significant logistical and financial barriers, especially in low- and middle-income countries (LMICs) where access to reliable electricity and refrigeration is limited [10]. In endemic regions such as Nepal, vaccination campaigns are essential to control and prevent outbreaks of several OIE-listed diseases, including rabies, foot-and-mouth disease (FMD), hemorrhagic septicemia (HS), black quarter (BQ), Peste des Petits Ruminants (PPR), and classical swine fever (CSF) [10]. However, the lack of infrastructure in these regions makes the thermal stability of vaccines a critical concern for global veterinary immunization efforts.
To address these limitations, next-generation vaccine technologies such as nanovaccine platforms have emerged as promising alternatives. These platforms introduce a novel approach to vaccine delivery by leveraging nanoparticles (NPs)—including lipids, polymers, and proteins—to enhance antigen targeting and stability [11]. Nanoparticles facilitate precise delivery to immune cells, improving protection against a wide range of infectious and zoonotic diseases relevant to animal health [11]. Additionally, nanocarrier-based systems contribute to improved vaccine stability, protect against premature degradation, and possess strong adjuvant properties that amplify immune responses [12].
As the demand for effective and resilient veterinary vaccines grows, especially in resource-limited settings, innovations in thermostability and delivery systems will play a central role in expanding the reach and impact of immunization programs worldwide.

2. Challenges in Conventional Vaccination

Despite their proven benefits, livestock vaccination programs, especially in LMICs, continue to face major obstacles, including cold-chain dependence, injectable administration, and complex logistical barriers. As mentioned earlier, most veterinary vaccines require refrigeration between 2 °C and 8 °C to maintain potency [9]. However, in resource-limited regions, inadequate infrastructure and frequent power outages often compromise cold-chain integrity, leading to vaccine spoilage. Globally, cold-chain failures in logistics result in the loss of nearly 50% of human vaccines, a rate likely mirrored in veterinary settings, though direct data are limited [9]. Such losses represent substantial financial burdens. A study by Dumpa et al. estimates vaccine wastage costs to be USD 200–300 million annually, accounting for up to 80% of immunization program expenses [13]. Moreover, the administration of thermally degraded vaccines increases the risk of immunization failure and disease outbreaks [14].
Historical eradication efforts, such as rinderpest in cattle and smallpox in humans, demonstrate the critical importance of stable and effective vaccines [15]. Yet many current veterinary vaccines remain thermolabile, limiting their reach in remote and underserved areas. Injectable vaccine delivery further compounds the problem. It requires trained personnel and sterile equipment and carries occupational risks. In a Nigerian survey, 79.5% of veterinarians reported accidental needlestick injuries, often accompanied by adverse symptoms [16]. These injuries not only compromise worker safety but also pose risks of zoonotic transmission. In resource-constrained settings, the reuse of needles due to equipment shortages can exacerbate these dangers. A study demonstrated that shared needles can mechanically transmit viruses such as bluetongue virus, even in the absence of visible blood contamination [17]. Needle technology in animal vaccination includes both conventional needle–syringe (NS) methods and needle-free injection devices (NFIDs), each presenting unique benefits and limitations. While needle-based approaches are commonly utilized, they carry risks such as accidental needle-stick injuries, cross-contamination between animals, and the need for careful disposal. In contrast, NFIDs can help minimize these concerns, promote better animal welfare, and potentially boost vaccine effectiveness by delivering antigens directly to targeted immune cells [18]. Logistical and infrastructural challenges also hinder vaccine deployment. In pastoralist regions across East Africa and South Asia, vaccinators often must travel long distances on foot, motorcycles, or unreliable vehicles to access scattered herds. Seasonal rains may render dirt roads impassable, disrupting immunization campaigns during critical vaccination periods. Socio-cultural factors further reduce vaccine uptake, as some farmers hesitate due to concerns about injection-related stress or animal injury, ultimately limiting timely immunization and herd immunity [19].
Collectively, reliance on cold-chain-dependent injectables, coupled with logistical, infrastructural, and socio-cultural barriers, substantially constrains conventional livestock vaccination programs, particularly in LMICs. These challenges underscore the urgent need for thermostable, user-friendly alternatives that can ensure broader and more reliable immunization coverage.

3. Thermostability in Vaccine Development

Conventional live attenuated vaccines (both virus and bacteria) typically remain stable when frozen but degrade rapidly at temperatures above 8 °C due to the denaturation of surface proteins, breakdown of nucleic acids, and disruption of lipid membranes [20,21]. However, live attenuated viral vaccines are typically more heat-sensitive and tend to degrade faster at room temperature compared to inactivated bacterial vaccines. This increased vulnerability is due to the viral proteins being more prone to heat-induced denaturation, which can compromise the vaccine’s effectiveness [22].
To overcome these thermal limitations, vaccine developers increasingly rely on drying techniques that reduce degradation pathways, extend shelf life, and diminish cold-chain dependency. These advancements are especially critical for pandemic preparedness, bioterrorism stockpiling, and immunization in resource-limited regions [23,24].
Freeze-drying (lyophilization) remains the most widely adopted method for vaccine stabilization. This technique involves three key stages—freezing, primary drying (sublimation), and secondary drying (desorption)—to remove water under low-temperature, low-pressure conditions [22,25]. It preserves antigenic integrity without subjecting vaccines to the heat stress of terminal sterilization [22]. Stabilizing excipients such as trehalose and sucrose are commonly used to protect labile proteins and viral particles during drying. For example, a lyophilized live-attenuated Equine Herpesvirus Type 4 vaccine formulated with sucrose and gelatin maintained its infectivity for over six months at 4 °C, indicating strong potential for deployment in low-temperature cold-chain environments [25].
Spray drying is an emerging alternative that offers scalable, continuous production by rapidly converting liquid vaccines into dry powders using hot gas [26,27]. Unlike the compact cakes formed in lyophilization, spray drying yields loose powders ideal for mucosal delivery, potentially eliciting stronger localized immune responses [26]. However, the technique introduces challenges: antigens are exposed to shear forces, heat, and air–liquid interfaces that may compromise stability, sometimes necessitating secondary drying to reduce residual moisture [27,28].
Regardless of the drying method, stabilizers like trehalose and sucrose are critical for preserving vaccine efficacy. Trehalose is preferred for its high glass transition temperature and amorphous properties, which enhance its protective effects, particularly in vaccines such as HSV-2 candidates [27]. Sucrose, though effective through hydrogen bonding, can crystallize under certain conditions, potentially diminishing vaccine stability [28]. Combinations of sugars and polymers, such as dextran, can further improve thermostability by elevating glass transition temperatures and reducing moisture absorption [29]. Notably, trehalose consistently outperforms sucrose in spray-dried formulations, though both must be carefully handled to prevent moisture-induced degradation [30,31]. Together, these formulation and drying innovations lay the groundwork for thermostable vaccines that can be safely stored, transported, and administered without reliance on the cold chain. This advancement is crucial for improving vaccine access and strengthening resilience in global health systems.
Furthermore, maintaining antigen stability is a vital component in the development of veterinary vaccines, as it helps preserve their immunological effectiveness during storage, handling, and administration. This is particularly important in veterinary contexts where access to reliable cold-chain systems may be limited. To safeguard antigen structure and function, various stabilizers such as sugars (like trehalose and sucrose), proteins (such as gelatin and albumin), and polymers (including polyethylene glycol) are frequently employed to prevent heat-induced degradation and aggregation [32]. In large animal species, foot-and-mouth disease (FMD) continues to severely affect livestock, often leading to significant economic losses. Traditional vaccines based on inactivated viruses require costly, high-level biosecurity facilities and continuous cold-chain storage, limiting their practicality. To overcome these issues, researchers have developed recombinant empty capsids as an alternative. By modifying expression systems to reduce viral protease activity, essential for capsid protein processing, they successfully produced A-serotype capsids in eukaryotic cells using vaccinia and baculovirus vectors, achieving yields suitable for commercial use [30]. Additionally, a stability-enhancing mutation was introduced, and structural analysis confirmed that the modified capsids closely resemble the natural virus, as picornaviruses have been shown to become more thermostable through the introduction of specific mutations, such as FMDV. Immunization of cattle with these recombinant capsids led to strong, long-lasting immune responses and protection lasting at least 34 weeks [30]. This method offers several advantages over current vaccines, including lower production costs, elimination of infection risk, and improved heat stability. Similar techniques could be adapted for safe vaccine development against other picornaviruses affecting both animals and humans [33]. Achieving stable antigen formulations ultimately enhances vaccine reliability and lowers the chances of immunization failure across various animal species.

4. Innovations in Vaccine Delivery Systems

4.1. Nanoparticles in Vaccinations

In recent decades, nanoparticle (NP)-based technologies have emerged as powerful tools in next-generation vaccine development, offering multiple advantages over traditional platforms that rely on inactivated viruses or soluble subunit antigens [34].
Nanoparticle-based vaccines, or nanovaccines, are developed by encapsulating antigens within nanoparticles or conjugating them to their surfaces. These systems enhance antigen bioavailability, protect against proteolytic degradation, and enable controlled, sustained antigen release. These features contribute to stronger and longer-lasting immune responses compared to those from conventional soluble antigen vaccines [35]. Beyond serving as delivery vehicles, many nanoparticles also possess intrinsic immunostimulatory properties, enabling them to function as both antigen carriers and adjuvants [35,36].
Among the various nanoparticle types studied, inorganic NPs such as carbon, gold, and silica have shown notable promise in vaccine delivery [26,37,38]. These particles can be synthesized in a wide range of shapes and sizes, and their surfaces can be chemically modified for targeted delivery [39]. Inorganic NPs also improve antigen stability by shielding vaccine components from enzymatic degradation. For instance, gold nanoparticles have been successfully used to deliver bacterial and viral antigens, eliciting robust immune responses in mice against pathogens such as Mycobacterium tuberculosis, influenza virus, HIV, and FMDV. Notably, encapsulation of plasmid DNA encoding the hsp65 antigen of M. tuberculosis in gold NPs significantly reduced bacterial burden in infected mice [40,41,42].
The use of organic particles in vaccine development provides numerous advantages, particularly in boosting immune responses and overcoming limitations of conventional vaccination approaches. Organic nanoparticles like liposomes and polymer-based carriers can enclose delicate antigens and genetic materials, shielding them from degradation. For instance, lipid nanoparticles (LNPs) protect mRNA from enzymatic breakdown and hydrolysis, which is essential for the success of mRNA-based vaccines [42]. Due to their small size and surface characteristics, organic nanoparticles can effectively pass through biological barriers and be absorbed by cells, including crucial antigen-presenting cells such as dendritic cells. Surface modification with targeting ligands can further improve precision and uptake. Many organic nanoparticles naturally stimulate the immune system, helping to amplify the response to the antigen. Cationic lipid particles, for example, can activate immune cells and promote the production of inflammatory cytokines, contributing to stronger and more durable immune protection. Virus-like particles (VLPs) are a notable type of organic nanoparticle that resemble real viruses in structure but lack infectivity [43]. This allows them to provoke strong immune responses, and they have already proven effective in vaccines against diseases like Hepatitis B and HPV. Organic nanoparticles can be engineered with specific sizes, shapes, and surface features to optimize vaccine performance. They are also suitable for scalable and economical production, making them an appealing solution for increasing vaccine availability, especially in low-resource settings.
Silica nanoparticles, which contain abundant surface silanol groups, offer versatile opportunities for surface functionalization to improve targeted antigen delivery into host cells [43,44]. These tailored properties enhance cellular uptake and immune activation, positioning silica NPs as adaptable platforms for advanced vaccine development. Overall, nanoparticle-based vaccine platforms offer promising alternatives to conventional formulations by enhancing delivery efficiency, stability, and immunogenicity. Their adaptability and dual functionality make them especially valuable for veterinary applications in both high-resource and resource-limited settings.
NPs significantly contribute to the activation of antigen-presenting cells (APCs), particularly dendritic cells (DCs), which are crucial for the effectiveness of vaccines [45]. While NPs can exhibit some cytotoxicity, [46,47] the potential harm is minimal relative to the advantages they offer in vaccine delivery [46]. Inorganic nanoparticles can be toxic to cells, potentially causing inflammation, oxidative damage, and even harm to cellular DNA. Long-term exposure to specific inorganic nanoparticles, like titanium dioxide, has been associated with persistent inflammation and tissue fibrosis. Certain inorganic nanoparticles may provoke harmful immune reactions, such as a Th2-skewed response—associated with allergic reactions and anaphylaxis—as seen with aluminum-based adjuvants, according to Petrovsky (2015) [48]. Studies have reported that the prolonged presence and buildup of inorganic nanoparticles in the body may impair the immune system’s capacity to defend against infections. For example, contact with specific inorganic nanoparticles can stimulate the release of inflammatory cytokines and interfere with immune cell communication pathways [49].
Although inorganic nanoparticles hold great promise for enhancing vaccine effectiveness, it is important to consider concerns about their toxicity, buildup in the body, and possible long-term impacts on the immune system. Ongoing studies and thorough safety assessments are vital to grasp and reduce these risks, promoting the safe and ethical integration of inorganic nanoparticles in vaccine creation.

4.2. Microneedles for Vaccine Delivery

Microneedle vaccines use tiny needle arrays, typically between 50 and 900 μm long, to gently breach the skin’s outer barrier, the stratum corneum, without causing pain. Vaccine components—whether in liquid form, particle-based form, or as nucleic acids—are delivered straight into the epidermis and dermis, areas densely populated with immune cells such as dendritic and Langerhans cells. According to the NIH, these microneedles—usually composed of dissolvable materials like sugars or polymers—either break down in the skin to release the vaccine or are taken off after their surface coating dissolves [50]. Delivering antigens directly to the skin’s rich network of immune cells triggers a strong immune reaction [51]. Antigen-presenting cells like Langerhans and dendritic cells take up the antigens and activate both antibody-based and T-cell immune responses [52]. Research indicates that microneedle vaccines can elicit stronger antibody production and more effective virus elimination than traditional intramuscular shots, potentially offering enhanced protection against infections.
Microneedle vaccine systems provide a practical and accessible alternative for areas lacking adequate healthcare facilities, advanced medical tools, and consistent cold-chain infrastructure. In contrast to conventional vaccines that typically demand refrigeration and skilled healthcare workers for administration, microneedle patches remain stable at ambient temperatures, are simple to use, and can be applied by individuals with minimal training. Their painless, needle-free design enhances acceptance while avoiding the dangers of needle-stick injuries and the complications of hazardous waste disposal. Lightweight and portable, they are particularly effective for use in hard-to-reach or resource-limited regions where vaccine storage and delivery pose logistical difficulties. These features make microneedle systems an excellent option for large-scale vaccination efforts in underserved communities, ultimately promoting wider vaccine coverage and better health outcomes.
Microneedle-based delivery systems offer a promising alternative to conventional intramuscular (IM) or subcutaneous injections, with several studies reporting enhanced immune responses and improved vaccine efficacy. In comparative models, microneedle immunization has demonstrated superior memory responses. For example, mice vaccinated using microneedles exhibited undetectable lung viral titers following challenge, indicating robust protection, whereas intramuscularly vaccinated counterparts showed significantly higher viral loads [47]. Similarly, recombinant subunit vaccines composed of trimeric influenza hemagglutinin protein, when delivered via coated microneedles, induced stronger immune responses in mice than subcutaneous administration [53]. In another study, the Bacillus Calmette–Guérin (BCG) vaccine delivered through coated microneedle devices elicited enhanced immunity in guinea pigs compared to traditional intradermal injection [54]. These findings highlight microneedles as a minimally invasive, immunologically potent, and potentially thermostable delivery system with significant promise for vaccine applications, particularly in settings with limited healthcare infrastructure.

4.3. Mucosal Routes in Vaccination (Intranasal and Aerosol)

Mucosal vaccines delivered through intranasal or aerosol routes are designed to stimulate immune responses directly at mucosal surfaces, which serve as primary entry points for many infectious agents. These vaccines specifically target mucosa-associated lymphoid tissues (MALTs), especially the nasal-associated lymphoid tissue (NALT) and bronchus-associated lymphoid tissue (BALT), which are abundant in antigen-presenting cells like dendritic cells and macrophages [55,56]. These cells capture and process the antigens introduced via the mucosa, presenting them to T and B lymphocytes. This process leads to the generation of secretory IgA (sIgA) at the mucosal surface to neutralize pathogens locally, as well as systemic IgG and T-cell responses for broader protection. Intranasal and aerosol-based vaccines thus offer the advantage of inducing both mucosal and systemic immunity, making them particularly effective against respiratory infections [57]. To enhance their effectiveness and bypass mucosal tolerance, adjuvants are frequently incorporated. In humans, notable examples include the cholera toxin B subunit (CTB), which aids in antigen uptake and immune stimulation; CpG oligodeoxynucleotides, which activate Toll-like receptor 9 (TLR9) to promote Th1 responses; and chitosan, a mucoadhesive compound that improves antigen delivery [58]. These adjuvants play a crucial role in strengthening the immune response, positioning mucosal vaccines as a promising approach for controlling airborne diseases like influenza, COVID-19, and respiratory syncytial virus (RSV).
Delivering vaccines through mucosal routes offers a promising alternative to traditional parenteral administration, particularly when paired with NP formulations composed of biodegradable and biocompatible polymers that enhance antigen delivery to mucosal surfaces [59]. Studies have shown that liposome-based DNA and subunit influenza nanovaccines administered to mice can stimulate robust humoral, cellular, and mucosal immune responses [47]. In livestock, intranasal (IN) vaccination has emerged as an effective alternative for influenza control, particularly in pigs, where it targets the respiratory mucosal immune system more directly than IM approaches [48,49]. This route not only elicits strong local immune protection in the respiratory tract but also enhances immunity at distal mucosal and systemic sites. In contrast, IM administration of whole inactivated influenza A virus (IAV) vaccines, currently the most widely used, often fails to induce sufficient mucosal antibody and cellular immune responses, interferes with maternal antibodies in young piglets, and, in some cases, may exacerbate respiratory disease [50,51]. Overall, mucosal vaccination, particularly via intranasal routes, offers a safe and effective strategy for enhancing both respiratory and systemic immunity. Its immunological advantages make it a compelling alternative to traditional IM delivery in research and field settings.

4.4. Oral Route in Vaccination

Oral vaccination strategies, first developed in the 1960s, have since been widely adapted for veterinary use, particularly to target gastrointestinal and respiratory pathogens [60,61,62]. In livestock and poultry, oral vaccines are now routinely administered via feed or drinking water to prevent diseases such as erysipelas, rotavirus, avian encephalomyelitis, and Newcastle disease (ND) [59].
These vaccines predominantly stimulate mucosal immune responses in the gut-associated lymphoid tissue (GALT), particularly in Peyer’s patches [63]. Efficient viral uptake has been demonstrated in several species. For instance, an attenuated oral rabies vaccine was shown to replicate in the tonsils of dogs, indicating effective mucosal delivery [64]. However, interspecies differences may limit efficacy, as illustrated by reduced viral uptake in skunks [65]. In aquaculture, oral vaccines are gaining momentum due to their logistical and welfare advantages over injections. These include reduced stress, lower labour costs, and improved handling efficiency [66]. Commercially available formulations now protect several finfish species from significant pathogens affecting the aquaculture industry [59,67,68]. As a practical, stress-free, and scalable approach, oral vaccination continues to gain traction across terrestrial and aquatic species, offering a promising route for broad immunization coverage and enhanced mucosal immunity in diverse veterinary settings.

5. Synergy of Thermostability and Delivery Platforms

Integrating thermostable vaccine formulations with innovative delivery platforms, such as microneedles, oral feeds, and intranasal sprays, enhances vaccine accessibility and practical use, particularly in remote and resource-limited agricultural settings. Heat-stable solid forms like patches or pellets reduce reliance on cold-chain logistics, facilitating large-scale immunization far from centralized facilities [10]. For example, the Rift Valley fever (RVF) vaccine candidate CL13T strain maintains full viability after prolonged exposure to 37–45 °C, demonstrating promise for use in tropical climates with limited refrigeration [9].
Microneedles incorporate thermostable vaccine formulations into dissolvable needle arrays that can be stored at ambient temperature and administered intradermally with minimal discomfort. This makes them suitable for field use or self-administration [69]. Arya et al. [70] reported that a rabies DNA vaccine delivered via dissolving MNPs remained stable for over three weeks at 4 °C and elicited immune responses comparable to those to IM injections in canines.
Oral vaccine delivery similarly benefits from thermostable formulations, which can be incorporated into livestock feed, pellets, or oral drenches, enabling mass immunization without injections or specialized personnel. A study found that an orally administered attenuated PPR vaccine retained high viral titers (>103.1 TCID50) for over eight hours at ambient temperature and induced significant seroconversion in Nigerian sheep and goats [71]. Likewise, a thermostable Newcastle disease virus (NDV) vaccine (I-2 strain) delivered via treated drinking water or feed protected over 1600 backyard chickens in Nigeria within two weeks, demonstrating oral vaccine feasibility in low-resource settings [72].
Thermostability also enhances needle-free intranasal or aerosol vaccine platforms that target mucosal immunity at pathogen entry sites. Dry powder intranasal sprays can be engineered for thermal resilience. For instance, Cutlip is developing a thermostable intranasal spray vaccine against H5N1 avian influenza for dairy cattle, aiming to block viral replication at initial infection sites and reduce zoonotic spill-over risk [73]. Such non-invasive delivery methods allow easy administration by farmers or community health workers, broadening the potential for widespread use. Together, thermostable formulations and advanced delivery systems offer practical, scalable solutions for vaccine deployment in diverse and resource-limited agricultural settings, improving immunization reach and effectiveness. A comparative overview of the major next-generation veterinary vaccine platforms, delivery routes, thermostability features, and their practical advantages is summarized in Table 1.

6. Implications for “One Health” and “Global Health”

Thermostable and easily deliverable vaccines for animals offer benefits that extend beyond livestock health, playing a pivotal role in advancing One Health (OH) goals by integrating human health, food security, and sustainable development [10]. Animals often act as reservoirs or amplifiers of zoonotic pathogens such as influenza viruses, Brucella, and Rift Valley fever virus, which can spill over to humans [76]. Vaccinating animals against these pathogens directly reduces the risk of human infection. For example, mass vaccination of Mongolian sheep and cattle against brucellosis using Rev-1 and S19 vaccines was estimated to prevent approximately 49,000 disability-adjusted life years (DALYs) over a decade, with a societal benefit-to-cost ratio of 3.2 [77]. Similarly, immunizing poultry and livestock against highly pathogenic avian influenza and Rift Valley fever helps prevent human outbreaks. Recent work at the University of Maryland showed that a nasal vaccine blocking avian H5N1 virus entry in cattle significantly reduced the risk of zoonotic transmission [73].
Beyond public health, livestock vaccination supports food security and economic development, especially in resource-limited settings where animals are crucial sources of protein, income, and financial resilience. Healthy herds produce more milk, meat, and eggs, contributing to Sustainable Development Goals (SDGs) such as Zero Hunger and Good Health [76]. Outbreaks of diseases like ND and PPR can devastate smallholder farms, posing threats to livelihoods and women’s empowerment. Expanding access to thermostable, user-friendly vaccines, therefore, strengthens food system resilience and supports rural economies. Thermostable vaccines also offer environmental benefits by minimizing reliance on cold-chain logistics, which impose significant energy and greenhouse gas burdens. Maintaining ultra-cold storage consumes large amounts of energy and relies on hydrofluorocarbon refrigerants with high global warming potential [69,78].
From both veterinary and public health standpoints, animal vaccination reduces disease prevalence and serves as a barrier against zoonoses transmitted through direct contact or vectors [79]. Vaccinating at the animal–human interface is a key strategy for pandemic prevention and embodies OH principles by interrupting disease transmission across species [80]. Therefore, thermostable livestock vaccines are indispensable for advancing animal welfare, safeguarding public health, and achieving global health and sustainability targets.

7. Future Outlook: Role of AI in Vaccine Innovation

The 21st century has witnessed remarkable technological advances aimed at enhancing healthcare, public health, and global resilience [9,69]. Among these, AI, which mimics cognitive functions such as learning and problem-solving, has revolutionized several domains, including healthcare and veterinary science [81]. AI and machine learning (ML) are transforming diagnostics and treatment strategies, with the global AI healthcare market projected to reach $45.2 billion by 2025 [72]. In veterinary medicine, these technologies are increasingly employed to enhance disease detection, guide vaccine development, and support decision-making in field conditions, especially where resources are limited [75].
AI is becoming increasingly important in vaccinology through the use of tools like deep learning algorithms, natural language processing, reverse-vaccinology systems, and immunoinformatics platforms [82]. These technologies support rapid and accurate identification of vaccine candidates, including B- and T-cell epitopes, antigen structures, and protein modeling. Notable examples include DeepVacPred for predicting epitopes, AlphaFold for determining protein structures, and Vaxign for analyzing pathogen genomes to guide vaccine development [83]. In developing countries, where healthcare systems often face constraints such as inadequate infrastructure, poor disease monitoring, and logistical challenges, AI offers valuable solutions. It can forecast outbreaks, streamline vaccine delivery, customize vaccines for region-specific pathogens, and facilitate real-time data use through mobile health applications—ultimately improving vaccine access, reducing delays in development, and strengthening immunization efforts in low-resource environments.
In vaccine research, AI holds immense potential to accelerate the development of rapid and broadly protective vaccines [84]. By analyzing genetic sequences, protein structures, and immune interactions, AI can predict immunogenic epitopes and identify optimal antigenic targets, thereby improving vaccine efficacy [85]. This approach is particularly valuable in veterinary and zoonotic disease management, offering tools to combat emerging infectious threats and antimicrobial resistance [62].
The pivotal role of AI was demonstrated during the COVID-19 pandemic, where it accelerated mRNA vaccine development (e.g., Pfizer-BioNTech and Moderna) through AI-driven antigen design and trial optimization [86]. AI also enabled rapid and accurate disease detection. For instance, deep learning algorithms identified COVID-19 on CT chest scans in under 5 s, compared to over 10 minutes by radiologists, offering nearly 135-fold speed improvement without fatigue-related errors [10,84,85].
If veterinarians are responsible for needle-stick injuries during vaccine administration, integrating AI into veterinary practice could offer significant safety and efficiency benefits. AI can support the adoption and optimization of alternative delivery methods, such as needle-free injection systems or microneedle patches, by analyzing data on injury rates, vaccine efficacy, and animal responses to guide the best practices. It can also assist in developing intelligent robotic or automated vaccine delivery systems that reduce direct human–animal interaction during injection, lowering the risk of accidents. Additionally, AI-powered monitoring tools can help track and predict high-risk scenarios, allowing for better training protocols and workflow adjustments. Ultimately, the use of AI in this context is not only about enhancing vaccine delivery but also about protecting veterinary professionals, improving workplace safety, and ensuring more consistent and humane vaccination practices.
Beyond diagnostics, AI assists veterinarians in disease surveillance, positive sample selection, and resource management, proving invaluable for monitoring animal diseases and controlling foodborne pathogens [87]. Although challenges remain in developing thermostable vaccines and novel delivery systems, machine learning tools offer a promising path forward. By accelerating antigen discovery and streamlining formulation processes, AI is poised to drive the next generation of veterinary vaccines.

8. Conclusions

The development of next-generation veterinary vaccines represents a transformative leap in preventing disease outbreaks at the animal–human–environment interface. Advances in thermostable formulations and innovative delivery platforms, such as microneedles, oral feed-based systems, and intranasal sprays, are addressing long-standing challenges related to cold-chain dependency, field applicability, and accessibility in low-resource settings. These technologies not only improve vaccine stability and immunogenicity but also expand the reach of immunization programs, particularly in rural and underserved areas. By reducing zoonotic transmission risks, supporting food security, and minimizing environmental impact, these innovations directly contribute to OH objectives and sustainable development goals. Furthermore, the integration of AI in vaccine research is accelerating antigen discovery, formulation design, and field diagnostics, paving the way for more targeted and responsive immunization strategies. Moving forward, cross-sector collaboration among scientists, veterinarians, policymakers, and global health stakeholders will be critical to translating these promising innovations into equitable and impactful solutions worldwide.

Author Contributions

Conceptualization, R.R., R.S. and A.A.; writing—original draft preparation, R.R., R.S., A.A. and A.F.; review and editing, R.R., A.F. and M.N. All authors have read and agreed to the published version of the manuscript.

Funding

The authors received no external funding for this study.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Moennig, V. Introduction to Classical Swine Fever: Virus, Disease and Control Policy. Vet. Microbiol. 2000, 73, 93–102. [Google Scholar] [CrossRef] [PubMed]
  2. Horzinek, M.C. Rinderpest: The Second Viral Disease Eradicated. Vet. Microbiol. 2011, 149, 295–297. [Google Scholar] [CrossRef]
  3. Chung, Y.H.; Church, D.; Koellhoffer, E.C.; Osota, E.; Shukla, S.; Rybicki, E.P.; Pokorski, J.K.; Steinmetz, N.F. Integrating Plant Molecular Farming and Materials Research for Next-Generation Vaccines. Nat. Rev. Mater. 2022, 7, 372–388. [Google Scholar] [CrossRef]
  4. Roth, J.A. Veterinary Vaccines and Their Importance to Animal Health and Public Health. Procedia Vaccinol. 2011, 5, 127–136. [Google Scholar] [CrossRef]
  5. Nooraei, S.; Sarkar Lotfabadi, A.; Akbarzadehmoallemkolaei, M.; Rezaei, N. Immunogenicity of Different Types of Adjuvants and Nano-Adjuvants in Veterinary Vaccines: A Comprehensive Review. Vaccines 2023, 11, 453. [Google Scholar] [CrossRef]
  6. Bailey, I. Edward Jenner (1749–1823): Naturalist, Scientist, Country Doctor, Benefactor to Mankind. J. Med. Biogr. 1996, 4, 63–70. [Google Scholar] [CrossRef]
  7. Cappellano, G.; Abreu, H.; Casale, C.; Dianzani, U.; Chiocchetti, A. Nano-Microparticle Platforms in Developing next-Generation Vaccines. Vaccines 2021, 9, 606. [Google Scholar] [CrossRef]
  8. McVey, S.; Shi, J. Vaccines in Veterinary Medicine: A Brief Review of History and Technology. Vet. Clin. North Am. Small Anim. Pract. 2010, 40, 381. [Google Scholar] [CrossRef] [PubMed]
  9. Brandau, D.T.; Jones, L.S.; Wiethoff, C.M.; Rexroad, J.; Middaugh, C.R. Thermal Stability of Vaccines. J. Pharm. Sci. 2003, 92, 218–231. [Google Scholar] [CrossRef]
  10. Fanelli, A.; Mantegazza, L.; Hendrickx, S.; Capua, I. Thermostable Vaccines in Veterinary Medicine: State of the Art and Opportunities to Be Seized. Vaccines 2022, 10, 245. [Google Scholar] [CrossRef] [PubMed]
  11. Murugan, B.; Sagadevan, S. Nano-Vaccines: Opportunities and Challenges in Biomaterial-Based Vaccine Delivery. Biomater.-Inspired Nanomed. Target. Ther. 2024, 101–116. [Google Scholar]
  12. Means, T.K.; Hayashi, F.; Smith, K.D.; Aderem, A.; Luster, A.D. The Toll-like Receptor 5 Stimulus Bacterial Flagellin Induces Maturation and Chemokine Production in Human Dendritic Cells. J. Immunol. 2003, 170, 5165–5175. [Google Scholar] [CrossRef] [PubMed]
  13. Dumpa, N.; Goel, K.; Guo, Y.; McFall, H.; Pillai, A.R.; Shukla, A.; Repka, M.A.; Murthy, S.N. Stability of Vaccines. AAPS PharmSciTech 2019, 20, 42. [Google Scholar] [CrossRef]
  14. Kristensen, D.; Chen, D.; Cummings, R. Vaccine Stabilization: Research, Commercialization, and Potential Impact. Vaccine 2011, 29, 7122–7124. [Google Scholar] [CrossRef] [PubMed]
  15. Rinderpest—WOAH—World Organisation for Animal Health. Available online: https://www.woah.org/en/disease/Rinderpest/ (accessed on 11 June 2025).
  16. Mshelbwala, P.P.; Weese, J.S.; Idris, J.M. Prevalence of Needlestick Injury and Its Potential Risk Among Veterinarians in Nigeria. 2016. Available online: https://onlinelibrary.wiley.com/doi/10.1155/2016/7639598 (accessed on 11 June 2025).
  17. Darpel, K.E.; Barber, J.; Hope, A.; Wilson, A.J.; Gubbins, S.; Henstock, M.; Frost, L.; Batten, C.; Veronesi, E.; Moffat, K.; et al. Using Shared Needles for Subcutaneous Inoculation Can Transmit Bluetongue Virus Mechanically between Ruminant Hosts. Sci. Rep. 2016, 6, 20627. [Google Scholar] [CrossRef]
  18. Rey, M.; Rodriguez-Lecompte, J.; Undi, M.; Joseph, T.; Morrison, J.; Yitbarek, A.; Wittenberg, K.; Tremblay, R.; Crow, G.; Ominski, K. Efficacy of Needle-Free Injection on Antibody Production against Clostridium Chauvoei in Beef Calves under Field Conditions. Can. Vet. J. 2015, 56, 405–407. [Google Scholar] [PubMed]
  19. Sacarrão-Birrento, L.; Harrison, L.J.S.; Pienaar, R.; Toka, F.N.; Torres-Acosta, J.F.J.; Vilela, V.L.R.; Hernández-Castellano, L.E.; Arriaga-Jordán, C.M.; Soltan, Y.A.; Ungerfeld, R.; et al. Challenges for Animal Health and Production in the Tropics and Mediterranean for the next 55 Years. Trop. Anim. Health Prod. 2024, 56, 381. [Google Scholar] [CrossRef]
  20. Josefsberg, J.O.; Buckland, B. Vaccine Process Technology. Biotech. Bioeng. 2012, 109, 1443–1460. [Google Scholar] [CrossRef]
  21. Kumar, R.; Srivastava, V.; Baindara, P.; Ahmad, A. Thermostable Vaccines: An Innovative Concept in Vaccine Development. Expert. Rev. Vaccines 2022, 21, 811–824. [Google Scholar] [CrossRef] [PubMed]
  22. Ghaemmaghamian, Z.; Zarghami, R.; Walker, G.; O’Reilly, E.; Ziaee, A. Stabilizing Vaccines via Drying: Quality by Design Considerations. Adv. Drug Deliv. Rev. 2022, 187, 114313. [Google Scholar] [CrossRef] [PubMed]
  23. Hansen, L.J.J.; Daoussi, R.; Vervaet, C.; Remon, J.-P.; De Beer, T.R.M. Freeze-Drying of Live Virus Vaccines: A Review. Vaccine 2015, 33, 5507–5519. [Google Scholar] [CrossRef]
  24. Thorat, B.N.; Sett, A.; Mujumdar, A.S. Drying of Vaccines and Biomolecules. Dry. Technol. 2022, 40, 461–483. [Google Scholar] [CrossRef]
  25. Kutumbetov, L.; Myrzakhmetova, B.; Tussipova, A.; Zhapparova, G.; Tlenchiyeva, T.; Bissenbayeva, K.; Nurabayev, S.; Kerimbayev, A. Development and Preclinical Evaluation of a Lyophilized Vaccine Against Equine Herpesvirus Type 4 (EHV-4). Vaccines 2025, 13, 604. [Google Scholar] [CrossRef] [PubMed]
  26. Kanojia, G.; Have, R.T.; Soema, P.C.; Frijlink, H.; Amorij, J.-P.; Kersten, G. Developments in the Formulation and Delivery of Spray Dried Vaccines. Hum. Vaccines Immunother. 2017, 13, 2364–2378. [Google Scholar] [CrossRef] [PubMed]
  27. LeClair, D.A.; Li, L.; Rahman, N.; Cranston, E.D.; Xing, Z.; Thompson, M.R. Stabilization of HSV-2 Viral Vaccine Candidate by Spray Drying. Int. J. Pharm. 2019, 569, 118615. [Google Scholar] [CrossRef]
  28. Flood, A.; Estrada, M.; McAdams, D.; Ji, Y.; Chen, D. Development of a Freeze-Dried, Heat-Stable Influenza Subunit Vaccine Formulation. PLoS ONE 2016, 11, e0164692. [Google Scholar] [CrossRef]
  29. Bacon, A.; Makin, J.; Sizer, P.J.; Jabbal-Gill, I.; Hinchcliffe, M.; Illum, L.; Chatfield, S.; Roberts, M. Carbohydrate Biopolymers Enhance Antibody Responses to Mucosally Delivered Vaccine Antigens. Infect. Immun. 2000, 68, 5764–5770. [Google Scholar] [CrossRef]
  30. Porta, C.; Kotecha, A.; Burman, A.; Jackson, T.; Ren, J.; Loureiro, S.; Jones, I.M.; Fry, E.E.; Stuart, D.I.; Charleston, B. Rational Engineering of Recombinant Picornavirus Capsids to Produce Safe, Protective Vaccine Antigen. PLoS Pathog. 2013, 9, e1003255. [Google Scholar] [CrossRef]
  31. Emami, F.; Vatanara, A.; Park, E.J.; Na, D.H. Drying Technologies for the Stability and Bioavailability of Biopharmaceuticals. Pharmaceutics 2018, 10, 131. [Google Scholar] [CrossRef]
  32. Bezbaruah, R.; Chavda, V.P.; Nongrang, L.; Alom, S.; Deka, K.; Kalita, T.; Ali, F.; Bhattacharjee, B.; Vora, L. Nanoparticle-Based Delivery Systems for Vaccines. Vaccines 2022, 10, 1946. [Google Scholar] [CrossRef]
  33. Fox, H.; Knowlson, S.; Minor, P.D.; Macadam, A.J. Genetically Thermo-Stabilised, Immunogenic Poliovirus Empty Capsids; a Strategy for Non-Replicating Vaccines. PLoS Pathog. 2017, 13, e1006117. [Google Scholar] [CrossRef] [PubMed]
  34. Dhakal, S.; Renukaradhya, G.J. Nanoparticle-Based Vaccine Development and Evaluation against Viral Infections in Pigs. Vet. Res. 2019, 50, 90. [Google Scholar] [CrossRef] [PubMed]
  35. Demento, S.L.; Cui, W.; Criscione, J.M.; Stern, E.; Tulipan, J.; Kaech, S.M.; Fahmy, T.M. Role of Sustained Antigen Release from Nanoparticle Vaccines in Shaping the T Cell Memory Phenotype. Biomaterials 2012, 33, 4957–4964. [Google Scholar] [CrossRef]
  36. Tandrup Schmidt, S.; Foged, C.; Smith Korsholm, K.; Rades, T.; Christensen, D. Liposome-Based Adjuvants for Subunit Vaccines: Formulation Strategies for Subunit Antigens and Immunostimulators. Pharmaceutics 2016, 8, 7. [Google Scholar] [CrossRef] [PubMed]
  37. Chen, Y.-S.; Hung, Y.-C.; Lin, W.-H.; Huang, G.S. Assessment of Gold Nanoparticles as a Size-Dependent Vaccine Carrier for Enhancing the Antibody Response against Synthetic Foot-and-Mouth Disease Virus Peptide. Nanotechnology 2010, 21, 195101. [Google Scholar] [CrossRef]
  38. Wang, T.; Zou, M.; Jiang, H.; Ji, Z.; Gao, P.; Cheng, G. Synthesis of a Novel Kind of Carbon Nanoparticle with Large Mesopores and Macropores and Its Application as an Oral Vaccine Adjuvant. Eur. J. Pharm. Sci. 2011, 44, 653–659. [Google Scholar] [CrossRef]
  39. Pati, R.; Shevtsov, M.; Sonawane, A. Nanoparticle Vaccines against Infectious Diseases. Front. Immunol. 2018, 9, 2224. [Google Scholar] [CrossRef] [PubMed]
  40. Silva, C.L.; Bonato, V.L.D.; Coelho-Castelo, A.A.M.; De Souza, A.O.; Santos, S.A.; Lima, K.M.; Faccioli, L.H.; Rodrigues, J.M. Immunotherapy with Plasmid DNA Encoding Mycobacterial Hsp65 in Association with Chemotherapy Is a More Rapid and Efficient Form of Treatment for Tuberculosis in Mice. Gene Ther. 2005, 12, 281–287. [Google Scholar] [CrossRef] [PubMed]
  41. Tao, W.; Gill, H.S. M2e-Immobilized Gold Nanoparticles as Influenza A Vaccine: Role of Soluble M2e and Longevity of Protection. Vaccine 2015, 33, 2307–2315. [Google Scholar] [CrossRef]
  42. Poon, C.; Patel, A.A. Organic and Inorganic Nanoparticle Vaccines for Prevention of Infectious Diseases. Nano Ex. 2020, 1, 012001. [Google Scholar] [CrossRef]
  43. Donaldson, B.; Lateef, Z.; Walker, G.F.; Young, S.L.; Ward, V.K. Virus-like Particle Vaccines: Immunology and Formulation for Clinical Translation. Expert. Rev. Vaccines 2018, 17, 833–849. [Google Scholar] [CrossRef] [PubMed]
  44. Fang, Z.; Li, X.; Xu, Z.; Du, F.; Wang, W.; Shi, R.; Gao, D. Hyaluronic Acid-Modified Mesoporous Silica-Coated Superparamagnetic Fe3O4 Nanoparticles for Targeted Drug Delivery. IJN 2019, 14, 5785–5797. [Google Scholar] [CrossRef]
  45. Nanoparticle-Based Vaccines. In Nanopharmaceuticals; Elsevier: Amsterdam, The Netherlands, 2020; pp. 135–150. ISBN 978-0-12-817778-5.
  46. Eidi, H.; Joubert, O.; Némos, C.; Grandemange, S.; Mograbi, B.; Foliguet, B.; Tournebize, J.; Maincent, P.; Le Faou, A.; Aboukhamis, I.; et al. Drug Delivery by Polymeric Nanoparticles Induces Autophagy in Macrophages. Int. J. Pharm. 2012, 422, 495–503. [Google Scholar] [CrossRef]
  47. Eidi, H.; Joubert, O.; Attik, G.; Duval, R.E.; Bottin, M.C.; Hamouia, A.; Maincent, P.; Rihn, B.H. Cytotoxicity Assessment of Heparin Nanoparticles in NR8383 Macrophages. Int. J. Pharm. 2010, 396, 156–165. [Google Scholar] [CrossRef]
  48. Petrovsky, N. Comparative Safety of Vaccine Adjuvants: A Summary of Current Evidence and Future Needs. Drug Saf. 2015, 38, 1059–1074. [Google Scholar] [CrossRef]
  49. Bhatti, R.; Shakeel, H.; Malik, K.; Qasim, M.; Khan, M.A.; Ahmed, N.; Jabeen, S. Inorganic Nanoparticles: Toxic Effects, Mechanisms of Cytotoxicity and Phytochemical Interactions. Adv. Pharm. Bull. 2021, 12, 757–762. [Google Scholar] [CrossRef] [PubMed]
  50. Pan, P.; Liu, Q.; Wang, L.; Wang, C.; Hu, L.; Jiang, Y.; Deng, Y.; Li, G.; Chen, J. Recent Advances in Multifunctional Microneedle Patches for Wound Healing and Health Monitoring. Adv. NanoBiomed Res. 2023, 3, 2200126. [Google Scholar] [CrossRef]
  51. Menon, I.; Bagwe, P.; Gomes, K.B.; Bajaj, L.; Gala, R.; Uddin, M.N.; D’Souza, M.J.; Zughaier, S.M. Microneedles: A New Generation Vaccine Delivery System. Micromachines 2021, 12, 435. [Google Scholar] [CrossRef]
  52. Sullivan, S.P.; Koutsonanos, D.G.; Del Pilar Martin, M.; Lee, J.W.; Zarnitsyn, V.; Choi, S.-O.; Murthy, N.; Compans, R.W.; Skountzou, I.; Prausnitz, M.R. Dissolving Polymer Microneedle Patches for Influenza Vaccination. Nat. Med. 2010, 16, 915–920. [Google Scholar] [CrossRef]
  53. Weldon, W.C.; Martin, M.P.; Zarnitsyn, V.; Wang, B.; Koutsonanos, D.; Skountzou, I.; Prausnitz, M.R.; Compans, R.W. Microneedle Vaccination with Stabilized Recombinant Influenza Virus Hemagglutinin Induces Improved Protective Immunity. Clin. Vaccine Immunol. 2011, 18, 647–654. [Google Scholar] [CrossRef]
  54. Hiraishi, Y.; Nandakumar, S.; Choi, S.-O.; Lee, J.W.; Kim, Y.-C.; Posey, J.E.; Sable, S.B.; Prausnitz, M.R. Bacillus Calmette-Guerin Vaccination Using a Microneedle Patch. Vaccine 2011, 29, 2626–2636. [Google Scholar] [CrossRef]
  55. Chen, K.; Cerutti, A. Vaccination Strategies to Promote Mucosal Antibody Responses. Immunity 2010, 33, 479–491. [Google Scholar] [CrossRef]
  56. Seefeld, M.L.; Templeton, E.L.; Lehtinen, J.M.; Sinclair, N.; Yadav, D.; Hartwell, B.L. Harnessing the Potential of the NALT and BALT as Targets for Immunomodulation Using Engineering Strategies to Enhance Mucosal Uptake. Front. Immunol. 2024, 15, 1419527. [Google Scholar] [CrossRef] [PubMed]
  57. Zhou, M.; Xiao, H.; Yang, X.; Cheng, T.; Yuan, L.; Xia, N. Novel Vaccine Strategies to Induce Respiratory Mucosal Immunity: Advances and Implications. MedComm 2025, 6, e70056. [Google Scholar] [CrossRef]
  58. Tizard, I.R. The Administration of Vaccines. Vaccines Vet. 2020, 87–104. [Google Scholar]
  59. Stratmann, T. Cholera Toxin Subunit B as Adjuvant––An Accelerator in Protective Immunity and a Break in Autoimmunity. Vaccines 2015, 3, 579–596. [Google Scholar] [CrossRef]
  60. Mishra, N.; Goyal, A.K.; Tiwari, S.; Paliwal, R.; Paliwal, S.R.; Vaidya, B.; Mangal, S.; Gupta, M.; Dube, D.; Mehta, A. Recent Advances in Mucosal Delivery of Vaccines: Role of Mucoadhesive/Biodegradable Polymeric Carriers. Expert. Opin. Ther. Pat. 2010, 20, 661–679. [Google Scholar] [CrossRef] [PubMed]
  61. Howard, B.D. A Prototype Live Oral Cholera Vaccine. Nature 1971, 230, 97–99. [Google Scholar] [CrossRef] [PubMed]
  62. Kwong, K.W.-Y.; Xin, Y.; Lai, N.C.-Y.; Sung, J.C.-C.; Wu, K.-C.; Hamied, Y.K.; Sze, E.T.-P.; Lam, D.M.-K. Oral Vaccines: A Better Future of Immunization. Vaccines 2023, 11, 1232. [Google Scholar] [CrossRef]
  63. Zhong, K.; Chen, X.; Zhang, J.; Jiang, X.; Zhang, J.; Huang, M.; Bi, S.; Ju, C.; Luo, Y. Recent Advances in Oral Vaccines for Animals. Vet. Sci. 2024, 11, 353. [Google Scholar] [CrossRef]
  64. Zhou, M.; Wang, L.; Zhou, S.; Wang, Z.; Ruan, J.; Tang, L.; Jia, Z.; Cui, M.; Zhao, L.; Fu, Z.F. Recombinant Rabies Virus Expressing Dog GM-CSF Is an Efficacious Oral Rabies Vaccine for Dogs. Oncotarget 2015, 6, 38504. [Google Scholar] [CrossRef]
  65. Vos, A.; Freuling, C.M.; Hundt, B.; Kaiser, C.; Nemitz, S.; Neubert, A.; Nolden, T.; Teifke, J.P.; Te Kamp, V.; Ulrich, R. Oral Vaccination of Wildlife against Rabies: Differences among Host Species in Vaccine Uptake Efficiency. Vaccine 2017, 35, 3938–3944. [Google Scholar] [CrossRef]
  66. Du, Y.; Hu, X.; Miao, L.; Chen, J. Current Status and Development Prospects of Aquatic Vaccines. Front. Immunol. 2022, 13, 1040336. [Google Scholar] [CrossRef]
  67. Babai, I.; Samira, S.; Barenholz, Y.; Zakay-Rones, Z.; Kedar, E. A Novel Influenza Subunit Vaccine Composed of Liposome-Encapsulated Haemagglutinin/Neuraminidase and IL-2 or GM-CSF. I. Vaccine Characterization and Efficacy Studies in Mice. Vaccine 1999, 17, 1223–1238. [Google Scholar] [CrossRef]
  68. Wang, D.; Christopher, M.E.; Nagata, L.P.; Zabielski, M.A.; Li, H.; Wong, J.P.; Samuel, J. Intranasal Immunization with Liposome-Encapsulated Plasmid DNA Encoding Influenza Virus Hemagglutinin Elicits Mucosal, Cellular and Humoral Immune Responses. J. Clin. Virol. 2004, 31, 99–106. [Google Scholar] [CrossRef] [PubMed]
  69. Kolluru, C.; Gomaa, Y.; Prausnitz, M.R. Development of a Thermostable Microneedle Patch for Polio Vaccination. Drug Deliv. Transl. Res. 2019, 9, 192–203. [Google Scholar] [CrossRef]
  70. Arya, J.M.; Dewitt, K.; Scott-Garrard, M.; Chiang, Y.-W.; Prausnitz, M.R. Rabies Vaccination in Dogs Using a Dissolving Microneedle Patch. J. Control. Release 2016, 239, 19–26. [Google Scholar] [CrossRef] [PubMed]
  71. El-Yuguda, A.-D.; Baba, S.S.; Ambali, A.G.; Egwu, G.O. Field Trial of a Thermostable Peste Des Petits Ruminants (PPR) Vaccine in a Semi-Arid Zone of Nigeria. World J. Vaccines 2014, 2014. [Google Scholar] [CrossRef]
  72. Bell, J.G.; Fotzo, T.M.; Amara, A.; Agbede, G. A Field Trial of the Heat Resistant V4 Vaccine against Newcastle Disease by Eye-Drop Inoculation in Village Poultry in Cameroon. Prev. Vet. Med. 1995, 25, 19–25. [Google Scholar] [CrossRef]
  73. A Spray in a Cow’s Nose Could Soon Protect It, and People, From Bird…. Available online: https://today.umd.edu/a-spray-in-a-cows-nose-could-soon-protect-it-and-people-from-bird-flu (accessed on 11 June 2025).
  74. Neutra, M.R.; Kozlowski, P.A. Mucosal Vaccines: The Promise and the Challenge. Nat. Rev. Immunol. 2006, 6, 148–158. [Google Scholar] [CrossRef] [PubMed]
  75. Agrebi, S.; Larbi, A. Use of Artificial Intelligence in Infectious Diseases. In Artificial Intelligence in Precision Health; Elsevier: Amsterdam, The Netherlands, 2020; pp. 415–438. [Google Scholar]
  76. Lane, J.K.; Kelly, T.; Bird, B.; Chenais, E.; Roug, A.; Vidal, G.; Gallardo, R.; Zhou, H.; VanHoy, G.; Smith, W. A One Health Approach to Reducing Livestock Disease Prevalence in Developing Countries: Advances, Challenges, and Prospects. Annu. Rev. Anim. Biosci. 2025, 13, 277–302. [Google Scholar] [CrossRef]
  77. Roth, F.; Zinsstag, J.; Orkhon, D.; Chimed-Ochir, G.; Hutton, G.; Cosivi, O.; Carrin, G.; Otte, J. Human Health Benefits from Livestock Vaccination for Brucellosis: Case Study. Bull. World Health Organ. 2003, 81, 867–876. [Google Scholar]
  78. Bozorgi, A.; Pazour, J.; Nazzal, D. A New Inventory Model for Cold Items That Considers Costs and Emissions. Int. J. Prod. Econ. 2014, 155, 114–125. [Google Scholar] [CrossRef]
  79. Udainiya, S.; Tiwari, A.; Mishra, A.; Dubey, A. Chapter 37—Zoonotic Diseases of Dogs and Cats. In Introduction to Diseases, Diagnosis, and Management of Dogs and Cats; Rana, T., Ed.; Developments in Microbiology; Academic Press: Cambridge, MA, USA, 2024; pp. 559–572. ISBN 978-0-443-18548-9. [Google Scholar]
  80. Entrican, G.; Francis, M.J. Applications of Platform Technologies in Veterinary Vaccinology and the Benefits for One Health. Vaccine 2022, 40, 2833–2840. [Google Scholar] [CrossRef]
  81. Esonu, D.; Armson, B.; Babashani, M.; Alafiatayo, R.; Ekiri, A.B.; Cook, A.J.C. Epidemiology of Peste Des Petits Ruminants in Nigeria: A Review. Front. Vet. Sci. 2022, 9, 898485. [Google Scholar] [CrossRef]
  82. Hederman, A.P.; Ackerman, M.E. Leveraging Deep Learning to Improve Vaccine Design. Trends Immunol. 2023, 44, 333–344. [Google Scholar] [CrossRef] [PubMed]
  83. Bhattacharya, M.; Lo, Y.-H.; Chatterjee, S.; Das, A.; Wen, Z.-H.; Chakraborty, C. Deep Learning in Next-Generation Vaccine Development for Infectious Diseases. Mol. Ther. Nucleic Acids 2025, 36, 102586. [Google Scholar] [CrossRef] [PubMed]
  84. Olawade, D.B.; Teke, J.; Fapohunda, O.; Weerasinghe, K.; Usman, S.O.; Ige, A.O.; David-Olawade, A.C. Leveraging Artificial Intelligence in Vaccine Development: A Narrative Review. J. Microbiol. Methods 2024, 106998. [Google Scholar] [CrossRef] [PubMed]
  85. Garcia-del Rio, L.; Diaz-Rodriguez, P.; Pedersen, G.K.; Christensen, D.; Landin, M. Sublingual Boosting with a Novel Mucoadhesive Thermogelling Hydrogel Following Parenteral CAF01 Priming as a Strategy against Chlamydia Trachomatis. Adv. Healthc. Mater. 2022, 11, 2102508. [Google Scholar] [CrossRef]
  86. Abubaker Bagabir, S.; Ibrahim, N.K.; Abubaker Bagabir, H.; Hashem Ateeq, R. Covid-19 and Artificial Intelligence: Genome Sequencing, Drug Development and Vaccine Discovery. J. Infect. Public. Health 2022, 15, 289–296. [Google Scholar] [CrossRef]
  87. Wang, H.; Cui, W.; Guo, Y.; Du, Y.; Zhou, Y. Machine Learning Prediction of Foodborne Disease Pathogens: Algorithm Development and Validation Study. JMIR Med. Inform. 2021, 9, e24924. [Google Scholar] [CrossRef] [PubMed]
Table 1. Overview of next-generation veterinary vaccines, including delivery platforms, target species, thermostability features, immune response sites, key advantages, limitations, and representative examples.
Table 1. Overview of next-generation veterinary vaccines, including delivery platforms, target species, thermostability features, immune response sites, key advantages, limitations, and representative examples.
Vaccine Type/PlatformDelivery RouteTarget Species/GroupThermostability FeaturesImmune Response/Target SiteAdvantagesChallenges/LimitationsKey Examples & References
NanovaccinesOral, intranasal, injectableLivestock, poultry, and companion animalsThermostability varies; often enhanced via NP formulationSystemic and mucosal (IgG, IgA, T-cells)Enhanced antigen stability, delivery, and intrinsic adjuvant activityComplex formulation; regulatory hurdlesGold NP for M. tuberculosis, influenza NP vaccine[40]
MicroneedlesSkin (intradermal)Canines, livestockHighly thermostable (ambient-stable for weeks)Skin immune cells, systemic immunityMinimal discomfort, user-friendly, and ambient-stableRequires formulation optimization, scalability issuesRabies DNA microneedle vaccine, BCG microneedle vaccine [54]
Liquid-based Intranasal VaccinesIntranasalPigs, poultry, cattleThermostable dry powder (in some formulations)Respiratory mucosa (IgA, T-cells)Strong mucosal immunity, needle-free, easy field usePrecise dosing, cold chain needed for some live formsH5N1 vaccine in cattle, influenza in pigs [74]
Dry Powder Intranasal Vaccines IntranasalPoultry, cattleDry, thermostable powder formMucosal IgA, systemic cross-protectionNon-invasive, ambient-stable, easy storageDifficult dose uniformity, delivery technique-dependentH5N1 spray [67]
Oral Vaccines (Drinking Water, Feed-Based)Oral (drinking water, pellets, feed mix)Poultry, small ruminants, and aquaculture speciesThermostable pellets/powders; liquids are more sensitiveGALT, Peyer’s patches (IgA)Easy mass administration, scalable, feed-based, thermostable optionsVariable uptake, degradation (especially in liquid form)NDV vaccine via feed or water, PPR vaccine [72]
AI-assisted vaccine designN/A (research phase)All animal speciesEnables the design of thermostable antigensOptimized antigen targets, surveillance data integrationRapid design, field diagnostics, surveillance supportRequires data, advanced infrastructuremRNA-based vaccine[75]
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Raut, R.; Shrestha, R.; Adhikari, A.; Fatima, A.; Naeem, M. Revolutionizing Veterinary Vaccines: Overcoming Cold-Chain Barriers Through Thermostable and Novel Delivery Technologies. Appl. Microbiol. 2025, 5, 83. https://doi.org/10.3390/applmicrobiol5030083

AMA Style

Raut R, Shrestha R, Adhikari A, Fatima A, Naeem M. Revolutionizing Veterinary Vaccines: Overcoming Cold-Chain Barriers Through Thermostable and Novel Delivery Technologies. Applied Microbiology. 2025; 5(3):83. https://doi.org/10.3390/applmicrobiol5030083

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Raut, Rabin, Roshik Shrestha, Ayush Adhikari, Arjmand Fatima, and Muhammad Naeem. 2025. "Revolutionizing Veterinary Vaccines: Overcoming Cold-Chain Barriers Through Thermostable and Novel Delivery Technologies" Applied Microbiology 5, no. 3: 83. https://doi.org/10.3390/applmicrobiol5030083

APA Style

Raut, R., Shrestha, R., Adhikari, A., Fatima, A., & Naeem, M. (2025). Revolutionizing Veterinary Vaccines: Overcoming Cold-Chain Barriers Through Thermostable and Novel Delivery Technologies. Applied Microbiology, 5(3), 83. https://doi.org/10.3390/applmicrobiol5030083

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