Search Results (174)

The development of serum-free media (SFM) is critical to advance cell culture techniques used in viral vaccine production and address the ethical concerns and contamination risks associated with fetal bovine serum (FBS). This study evaluated the effects of marine microalgal extracts and growth factor cocktails on the activity of Madin-Darby canine kidney (MDCK) and Vero cells. Five marine microalgal species were used: Spirulina platensis (SP), Dunaliella salina (DS), Haematococcus pluvialis (HP), Nannochloropsis salina (NS), and Tetraselmis sp. (TS). DS and SP extracts significantly increased the proliferation rate of both MDCK and Vero cells. DS had a proliferation rate of 149.56% and 195.50% in MDCK and Vero cells, respectively, compared with that in serum-free medium (SFM). Notably, DS and SP extracts significantly increased superoxide dismutase (SOD) activity, which was 118.61% in MDCK cells and 130.08% in Vero cells for DS, and 108.72% in MDCK cells and 125.63% in Vero cells for SP, indicating a reduction in intracellular oxidative stress. Marine microalgal extracts, especially DS and SP, are feasible alternatives to FBS in cell culture as they promote cell proliferation, ensure safety, and supply essential nutrients while reducing oxidative stress. Full article

Background: WHO and WOAH advocate for annual high-coverage canine rabies vaccination campaigns as the most sustainable approach to eliminate the risk of dog rabies transmission to humans. It is estimated that Cambodia has one of the highest human rabies deaths per capita of any country (5.8 human deaths per 100,000 people), highlighting the urgent need to implement an effective canine rabies vaccination program. To this end, a coalition of government and non-government organizations conducted a pioneering short-duration dog rabies vaccination campaign over 10 days across Phnom Penh and Kandal Provinces in May 2023. Methods: Over 10 working days, 120 vaccination teams, each consisting of two vaccinators and one tuk-tuk driver, traveled door-to-door to deliver parenteral rabies vaccines to all dogs that could be held by the teams or members of the community. Spatial team management and data collection were conducted through the WVS Data Collection Application. Results: A total of 74,983 dogs were vaccinated, giving a mean vaccination rate of 62.5 dogs per team per day. An additional 2145 cats were vaccinated. Of all dogs encountered by the teams, 84.0% could be vaccinated, with 99.1% of those being identified as owned. Post-vaccination surveys of 4356 households estimated a mean vaccination coverage of 70.7% in owned dogs across the districts of Phnom Penh Province. Conclusion: Short-duration, large-scale canine rabies vaccination campaigns can achieve high vaccination coverage using a door-to-door approach in urban centers of Cambodia. Data gathered through the campaign yielded insights to support the refinement and planning of a wider rabies control strategy and is anticipated to drive further support for subsequent campaigns in Cambodia. Full article

The study conducted in Morocco focused on addressing the challenges posed by canine parvovirus (CPV-2) through comprehensive research, vaccine development, and efficacy assessment. Through real-time PCR screening and genotyping, CPV-2 variants were identified circulating in the region. An inactivated vaccine, derived from a CPV-2 strain isolated from a symptomatic dog, was produced and evaluated for safety and efficacy. The vaccine, from the strain named “CaPV M/3-2022”, demonstrated safety in vaccinated puppies, with no adverse reactions observed during the trial period. Efficacy trials showed that vaccinated puppies remained healthy and exhibited lower viral excretion post-challenge compared to unvaccinated controls. These results indicate that the vaccine effectively protects against illness related to CPV-2 and reduces viral shedding. The study provides valuable insights into CPV-2 epidemiology in Morocco, offers a promising vaccine solution, and underscores the importance of vaccination in controlling CPV-2 outbreaks and protecting canine health. Full article

Madin–Darby canine kidney (MDCK) cells are commonly used to produce cell-based influenza vaccines. However, the role of the low-serum medium on the proliferation of MDCK cells and the propagation of the influenza virus has not been well studied. In the present study, we used 5 of 15 culture methods with different concentrations of a mixed medium and neonatal bovine serum (NBS) to determine the best culture medium. We found that a VP:M199 ratio of 1:2 (3% NBS) was suitable for culturing MDCK cells. Furthermore, the stable growth of MDCK cells and the production of the influenza virus were evaluated over long-term passaging. We found no significant difference in terms of cell growth and virus production between high and low passages of MDCK cells under low-serum culture conditions, regardless of influenza virus infection. Lastly, we performed a comparison of the transcriptomics and proteomics of MDCK cells cultured in VP:M199 = 1:2 (3% NBS) with those cultured in VP:M199 = 1:2 (5% NBS) before and after influenza virus infection. The transcriptome analysis showed that differentially expressed genes were predominantly enriched in the metabolic pathway and MAPK signaling pathway, indicating an activated state. This suggests that decreasing the concentration of serum in the medium from 5% to 3% may increase the metabolic activity of cells. Proteomics analysis showed that only a small number of differentially expressed proteins could not be enriched for analysis, indicating minimal difference in the protein levels of MDCK cells when the serum concentration in the medium was decreased from 5% to 3%. Altogether, our findings suggest that the screening and application of a low-serum medium provide a background for the development and optimization of cell-based influenza vaccines. Full article

Human monocytic ehrlichiosis, an emerging tick-borne disease, is caused by Ehrlichia chaffeensis. Infections with the pathogen are also common in the canine host. Our previous studies demonstrated that functional disruption within the E. chaffeensis phage head-to-tail connector protein gene results in bacterial attenuation, creating a modified live attenuated vaccine (MLAV). The MLAV confers protective immunity against intravenous and tick transmission challenges one month following vaccination. In this study, we evaluated the duration of MLAV protection. Dogs vaccinated with the MLAV were challenged with wild-type E. chaffeensis via intravenous infection at 4-, 8-, and 12-months post-vaccination. Immunized dogs rapidly cleared the wild-type pathogen infection and tested positive for bacteremia less frequently than unvaccinated controls. While immune responses varied among dogs, vaccinees consistently mounted IgG and CD4+ T-cell responses specific to E. chaffeensis throughout the assessment period. Our findings demonstrate that MLAV-mediated immune protection persists for at least one year against wild-type bacterial infection, marking a major advancement in combating this serious tick-borne disease. The data presented here serve as the foundation for further studies, elucidating the molecular mechanisms underlying virulence and vaccine development and aiding in preventing the diseases caused by E. chaffeensis and other tick-borne rickettsial pathogens. Full article

Coronaviruses are widespread in mammals and birds, causing mostly digestive and respiratory problems. In cats, feline coronaviruses undergo mutations while replicating, giving rise to the fatal coronavirus causing the feline infectious peritonitis (FIP) disease. Several mutations in viral genes, among them 3c and M, are involved in the development of FIP. In order to study these viral shifts, samples of 43 organs, feces, and ascites collected from cats showing clinical signs of feline infectious peritonitis were tested, and the sequences obtained for the 3c and M genes were analyzed. The 3c gene nucleotides showed truncations commonly observed in feline infectious peritonitis virus. Additionally, the sequences corresponding to the 3c genes obtained from different organs of the same individual displayed high similarities, supporting the internal mutation theory. The analyses of the M gene and putative polypeptides showed similarities with canine coronaviruses, supporting the recombination theory between feline and canine coronaviruses. Infectious coronaviral strains are still challenging because of the difficulty in obtaining an effective vaccine for their prevention, and also because of the limited alternatives for therapy of FIP in cats. Full article

Oncolytic viruses and morbilliviruses in particular, represent an interesting therapeutic approach for tumors with a poor prognosis and frequent resistance to conventional therapies. Canine histiocytic sarcomas (HS) exemplify such a neoplasm in need for new curative approaches. Previous investigations demonstrated a limited success of an acute intratumoral application of canine distemper virus (CDV) on xenotransplanted canine histiocytic sarcoma cells (DH82 cells), while persistently CDV-infected DH82 cell transplants exhibited a complete spontaneous regression. Therefore, the present study focuses on an intratumoral application of persistently CDV vaccine strain Onderstepoort-infected DH82 (DH82 Ond p.i.) cells into non-infected subcutaneous DH82 cell transplants in a murine model. DH82 cell transplants that received 10 applications, two days apart, showed a transient growth retardation as well as larger areas of intratumoral necrosis, lower mitotic rates, and a decreased intratumoral vascularization compared to controls. Viral mRNA was detected in all neoplasms following application of DH82 Ond p.i. cells until 66 days after the last injection. Furthermore, infectious virus was present until 62 days after the last injection. Although complete regression was not achieved, the present application regimen provides promising results as a basis for further treatments, particularly with genetically modified viruses, to enhance the observed effects. Full article

Morbillivirus canis (canine distemper virus (CDV)) is recognized as a multihost pathogen responsible for a transmissible disease affecting both domestic and wild animals. A considerable portion of wildlife populations remain unvaccinated due to a lack of safety and immunogenicity data on existing vaccines for the prevention of CDV infection in these species. This review aimed to assess the current state of CDV vaccination research for both domestic and wild animals and to explore novel vaccine candidates through in vivo studies. It also sought to synthesize the scattered information from the extensive scientific literature on CDV vaccine research, identify key researchers in the field, and highlight areas where research on CDV vaccination is lacking. A scoping review was conducted across four databases following the PRISMA-ScR protocol, with information analyzed using absolute and relative frequencies and 95% confidence intervals (CIs) for study number proportions. Among the 2321 articles retrieved, 68 met the inclusion criteria and focused on CDV vaccines in various animal species, such as dogs, ferrets, minks, and mice. Most of the scientific community involved in this research was in the USA, Canada, France, and Denmark. Various vaccine types, including MLV CDV, recombinant virus, DNA plasmids, inactivated CDV, and MLV measles virus (MeV), were identified, along with diverse immunization routes and schedules employed in experimental and commercial vaccines. Safety and efficacy data were summarized. Notably, 37 studies reported postimmunization CDV challenge, primarily in dogs, revealing the survival rates of vaccinated animals. In summary, CDV vaccines generally demonstrate an acceptable safety profile in dogs and show promise as a means of controlling CDV. However, significant gaps in vaccine research persist, particularly concerning wildlife reservoirs, indicating the need for further investigation. Full article

Carnivore protoparvovirus-1, feline parvovirus (FPV), and canine parvovirus (CPV) continue to spread in companion animals all over the world. As a result, FPV and CPV underwent host-to-host transfer in carnivorous wild-animal hosts. Here, a total of 82 fecal samples of suspected cat FPV infections were collected from Henan Province from 2020 to 2022. The previously published full-length sequence primers of VP2 and NS1 genes were used to amplify the targeted genes of these samples, and the complete gene sequences of 11 VP2 and 21 NS1 samples were obtained and analyzed. Analysis showed that the amino acid homology of the VP2 and NS1 genes of these isolates was 96.1–100% and 97.6–100%, respectively. The phylogenetic results showed that the VP2 and NS1 genes of the local isolates were mainly concentrated in the G1 subgroup, while the vaccine strains were distributed in the G3 subgroup. Finally, F81 cells were inoculated with the local endemic isolate Luoyang-01 (FPV-LY strain for short) for virus amplification, purification, and titer determination, and the pathogenesis of FPV-LY was detected. After five generations of blind transmission in F81 cells, cells infected with FPV-LY displayed characteristic morphological changes, including a round, threadlike, and wrinkled appearance, indicative of viral infection. The virus titer associated with this cytopathic effect (CPE) was measured at 1.5 × 10⁶ TCID₅₀/mL. Subsequent animal regression tests confirmed that the virus titer of the PFV-LY isolate remained at 1.5 × 10⁶ TCID₅₀/mL, indicating its highly pathogenic nature. Cats exposed to the virus exhibited typical clinical symptoms and pathological changes, ultimately succumbing to the infection. These results suggest that the gene mutation rate of FPV is increasing, resulting in a complex pattern of gene evolution in terms of host preference, geographical selection, and novel genetic variants. The data also indicate that continuous molecular epidemiological surveillance is required to understand the genetic diversity of FPV isolates. Full article

There is a growing interest in the composition of amniotic fluid (AF) in both humans and animals. In addition to its nutritional and protective functions for the foetus, current knowledge demonstrates that AF also serves advanced diagnostic, prognostic, and therapeutic roles. Newborn dogs have an underdeveloped immune system, making them highly susceptible to dangerous pathogens such as canine parvovirus (CPV-2), canine infectious hepatitis virus (CAdV-1), and canine distemper virus (CDV), thus exposing them to a high risk of mortality in the first weeks of life. Immunoglobulins G (IgGs) represent the only antibody isotype capable of crossing the placenta in a small amount and have been detected also in canine AF. The primary aim of this study was to investigate the reliability of AF collected at birth as a marker of passive immunity in canine species. For this purpose, total and specific IgGs against CPV-2, CAdV-1, and CDV were investigated and quantified in both maternal plasma and AF collected at the time of caesarean section. The vaccination status of the bitches was also taken into consideration. Since the immune system can be influenced by gestational age, with preterm infants having immature innate and adaptive immunity, IgG concentrations were correlated with amniotic lecithin, sphingomyelin, cortisol, surfactant protein A, and pentraxin 3 levels. In a previous study from our group on foetal maturity these molecules were measured in the same samples. Finally, correlations between their amniotic content and neonatal outcomes were investigated. This study demonstrates that AF analysis at birth can provide valuable insights into neonatal immunity in puppies, offering a non-invasive method to detect potential early health risks, for improved puppy care and management. Full article

Influenza vaccines, which are recommended by the World Health Organization (WHO), are the most effective preventive measure against influenza virus infection. Madin–Darby canine kidney (MDCK) cell culture is an emerging technology used to produce influenza vaccines. One challenge when purifying influenza vaccines using this cell culture system is to efficiently remove impurities, especially host cell double-stranded DNA (dsDNA) and host cell proteins (HCPs), for safety assurance. In this study, we optimized ion-exchange chromatography methods to harvest influenza viruses from an MDCK cell culture broth, the first step in influenza vaccine purification. Bind/elute was chosen as the mode of operation for simplicity. The anion-exchange Q chromatography method was able to efficiently remove dsDNA and HCPs, but the recovery rate for influenza viruses was low. However, the cation-exchange SP process was able to simultaneously achieve high dsDNA and HCP removal and high influenza virus recovery. For the SP process to work, the clarified cell culture broth needed to be diluted to reduce its ionic strength, and the optimal dilution rate was determined to be 1:2 with purified water. The SP process yielded a virus recovery rate exceeding 90%, as measured using a hemagglutination units (HAUs) assay, with removal efficiencies over 97% for HCPs and over 99% for dsDNA. Furthermore, the general applicability of the SP chromatography method was demonstrated with seven strains of influenza viruses recommended for seasonal influenza vaccine production, including H1N1, H3N2, B (Victoria), and B (Yamagata) strains, indicating that the SP process could be utilized as a platform process. The SP process developed in this study showed four advantages: (1) simple operation, (2) a high recovery rate for influenza viruses, (3) a high removal rate for major impurities, and (4) general applicability. Full article

Diagnosing canine visceral leishmaniasis (CVL) in Brazil faces challenges due to the limitations regarding the sensitivity and specificity of the current diagnostic protocol. Therefore, it is urgent to map new antigens or enhance the existing ones for future diagnostic techniques. Immunoinformatic tools are promising in the identification of new potential epitopes or antigen candidates. In this study, we evaluated peptides selected by epitope prediction for CVL serodiagnosis in ELISA assays. Ten B-cell epitopes were immunogenic in silico, but two peptides (peptides No. 45 and No. 48) showed the best performance in vitro. The selected peptides, both individually and in combination, were highly diagnostically accurate, with sensitivities ranging from 86.4% to 100% and with a specificity of approximately 90%. We observed that the combination of peptides showed better performance when compared to peptide alone, by detecting all asymptomatic dogs, showing lower cross-reactivity in sera from dogs with other canine infections, and did not detect vaccinated animals. Moreover, our data indicate the potential use of immunoinformatic tools associated with ELISA assays for the selection and evaluation of potential new targets, such as peptides, applied to the diagnosis of CVL. Full article

Canine distemper virus (CDV) can cause fatal infections in giant pandas. Vaccination is crucial to prevent CDV infection in giant pandas. In this study, two bacterium-like particle vaccines F₃-GEM and H₄-GEM displaying the trimeric F protein or tetrameric H protein of CDV were constructed based on the Gram-positive enhanced-matrix protein anchor (GEM-PA) surface display system. Electron microscopy and Western blot results revealed that the F or H protein was successfully anchored on the surface of GEM particles. Furthermore, one more bacterium-like particle vaccine F₃ and H₄-GEM was also designed, a mixture consisting of F₃-GEM and H₄-GEM at a ratio of 1:1. To evaluate the effect of the three vaccines, mice were immunized with F₃-GEM, H₄-GEM or F₃ and H₄-GEM. It was found that the level of IgG-specific antibodies and neutralizing antibodies in the F₃ and H₄-GEM group was higher than the other two groups. Additionally, F₃ and H₄-GEM also increased the secretion of Th1-related and Th2-related cytokines. Moreover, F₃ and H₄-GEM induce IgG and neutralizing antibodies’ response in dogs. Conclusions: In summary, F₃ and H₄-GEM can provoke better immune responses to CDV in mice and dogs. The bacterium-like particle vaccine F₃ and H₄-GEM might be a potential vaccine candidate for giant pandas against CDV infection. Full article

Chagas disease (CD) is a vector-borne Neglected Zoonotic Disease (NZD) caused by a flagellate protozoan, Trypanosoma cruzi, that affects various mammalian species across America, including humans and domestic animals. However, due to an increase in population movements and new routes of transmission, T. cruzi infection is presently considered a worldwide health concern, no longer restricted to endemic countries. Dogs play a major role in the domestic cycle by acting very efficiently as reservoirs and allowing the perpetuation of parasite transmission in endemic areas. Despite the significant progress made in recent years, still there is no vaccine against human and animal disease, there are few drugs available for the treatment of human CD, and there is no standard protocol for the treatment of canine CD. In this review, we highlight human and canine Chagas Disease in its different dimensions and interconnections. Dogs, which are considered to be the most important peridomestic reservoir and sentinel for the transmission of T. cruzi infection in a community, develop CD that is clinically similar to human CD. Therefore, an integrative approach, based on the One Health concept, bringing together the advances in genomics, immunology, and epidemiology can lead to the effective development of vaccines, new treatments, and innovative control strategies to tackle CD. Full article

Compared with the traditional vaccine produced in embryonated chicken eggs, cell-based manufacturing represented by the Madin–Darby canine kidney (MDCK) cell line has a larger production scale and reduces the risk of egg shortage in a pandemic. Establishing a culture system that enables high production of the influenza virus is a key issue in influenza vaccine production. Here, a serum-free suspension culture of MDCK (sMDCK) cells was obtained from adherent MDCK (aMDCK) cells by direct adaptation. Viral infection experiments showed that viral yields of influenza A/B virus in sMDCK cells were higher than in aMDCK cells. Transcriptome analysis revealed that numerous interferon-stimulated genes (ISGs) exhibited reduced expression in sMDCK cells. To further clarify the mechanism of high viral production in sMDCK cells, we demonstrated the antiviral role of RIG-I and IFIT3 in MDCK cells by knockdown and overexpression experiments. Furthermore, suppression of the JAK/STAT pathway enhances the viral accumulation in aMDCK cells instead of sMDCK cells, suggesting the reduction in the JAK/STAT pathway and ISGs promotes viral replication in sMDCK cells. Taken together, we elucidate the relationship between the host innate immune response and the high viral productive property of sMDCK cells, which helps optimize cell production processes and supports the production of cell-based influenza vaccines. Full article