Breaking the Barrier: SARS-CoV-2 Infections in Wild and Companion Animals and Their Implications for Public Health.

The emergence of the novel coronavirus SARS-CoV-2 has led to significant interest in its potential transmission between animals and humans, especially pets. This review article summarises the literature on coronavirus infections in domestic animals, emphasising epidemiology, transmission dynamics, clinical manifestations, and public health implications. This article highlights current understandings of the relationship between infections in companion animals and humans, identifies research gaps, and suggests directions for future research. Cases of disease in cats, dogs, and other domestic animals, often occurring through close contact with infected owners, are reviewed, raising concerns about possible zoonotic and reverse zoonotic transmission. Precautions and recommendations for pet owners and healthcare workers are also discussed. The scientific evidence presented in the article highlights the need for a One Health approach that considers the health of people, animals, and the environment to combat future pandemics.

Pre-Clinical Safety and Immunogenicity Study of a Coronavirus Protein-Based Subunit Vaccine for COVID-19.

Creating an effective and safe vaccine is critical to fighting the coronavirus infection successfully. Several types of COVID-19 vaccines exist, including inactivated, live attenuated, recombinant, synthetic peptide, virus-like particle-based, DNA and mRNA-based, and sub-unit vaccines containing purified immunogenic viral proteins. However, the scale and speed at which COVID-19 is spreading demonstrate a global public demand for an effective prophylaxis that must be supplied more. The developed products promise a bright future for SARS-CoV-2 prevention; however, evidence of safety and immunogenicity is mandatory before any vaccine can be produced. In this paper, we report on the results of our work examining the safety, toxicity, immunizing dose choice, and immunogenicity of QazCoVac-P, a Kazakhstan-made sub-unit vaccine for COVID-19. First, we looked into the product's safety profile by assessing its pyrogenicity in vaccinated rabbit models and using the LAL (limulus amebocyte lysate) test. We examined the vaccine's acute and sub-chronic toxicity on BALB/c mice and rats. The vaccine did not cause clinically significant toxicity-related changes or symptoms in our toxicity experiments. Finally, we performed a double immunization of mice, ferrets, Syrian hamsters, and rhesus macaques (Macaca mulatta). We used ELISA to measure antibody titers with the maximum mean geometric titer of antibodies in the animals' blood sera totaling approximately 8 log2. The results of this and other studies warrant recommending the QazCoVac-P vaccine for clinical trials.

Phylogenetic analysis of prospective M. bovis antigens with the aim of developing candidate vaccines for bovine tuberculosis.

BACKGROUND: Bovine Tuberculosis is a respiratory disease caused by the pathogen Mycobacterium bovis (M. bovis) that infects cattle. Though rare, this disease can also affect humans, as well as domestic and wild animals, making it a serious concern. Therefore, searching for alternative and new vaccines with high efficiency and safety is the main goal in bovine tuberculosis prophylaxis. New vaccines, known as vector vaccines, have the potential to become safe and effective alternatives to the traditional BCG vaccine. In this study, two major immunodominant proteins of M. bovis Esat-6 and TB10.4 were utilized to create a vector vaccine for bovine tuberculosis. METHODS: The Esat-6 and TB10.4 genes were amplified by PCR. The amplified and purified PCR products were sequenced by the Sanger method. Assembly and multiple alignments of amplicon nucleotides were carried out in the MEGA 11 software. RESULT: Two genes of the local strain 0078-M. bovis-8/RIBSP were sequenced. The nucleotide sequences were deposited in the GenBank database. Comparative analysis of the nucleotide sequences of the ESAT-6 and TB10.4 genes established 100% identity of the compared strains of Mycobacterium. CONCLUSION: Through the use of phylogenetic analysis, it has been confirmed that the amplified genes are related to the mycobacteria genus. This discovery allows the development of a vector vaccine against bovine tuberculosis utilising these genes.

Evaluation of a novel real-time polymerase chain reaction assay for identifying H3 equine influenza virus in Kazakhstan.

Background and Aim: Equine influenza (EI) is a highly contagious disease that causes fever and upper respiratory tract inflammation. It is caused by influenza virus A, belonging to the Orthomyxoviridae family, with subtypes H3N8 and H7N7. This study presents data on the development of a real-time polymerase chain reaction (RT-PCR) assay using TaqMan probes to detect the H3 subtype of EI virus (EIV). Materials and Methods: The evaluation of the developed RT-PCR assay involved five strains of EIV as positive controls and ten nasopharyngeal swab samples collected from horses. RNA was isolated using the GeneJet Viral DNA and RNA Purification Kit, and primers and probes were designed using the Integrated DNA Technology PrimerQuest Tool. The assay was optimized by investigating the annealing temperature, primer and probes concentrations, sensitivity, and specificity. Sequencing was performed using the Thermo Fisher 3130 Genetic Analyzer, and the evolutionary history was inferred using the Neighbor-Joining method. Results: The designed primers and probes, targeting the H3 gene, were found to be specific to the EIV. The RT-PCR assay was capable of detecting as low as 50 femtogram (f) or 3 × 103 copies of genomic RNA. No cross-reactions were observed with other respiratory viral and bacterial pathogens, indicating the high specificity of the assay. To evaluate its effectiveness, ten nasopharyngeal swab samples collected from farms in North Kazakhstan regions during disease monitoring were analyzed. The accuracy of the analysis was confirmed by comparing the results with those obtained from a commercial RT-PCR assay for EI identification. The developed RT-PCR assay exhibited high sensitivity and specificity for detecting the EIV. Conclusion: The results demonstrate that the developed RT-PCR assay is suitable for diagnosing EI. This simple, highly sensitive, and specific assay for detecting H3 EIV can be a reliable tool for diagnosing and surveilling EI. Implementing this RT-PCR assay in veterinary practice will enhance and expedite the timely response to potential outbreaks of EI, thus positively impacting the overall epizootic well-being of EI in Kazakhstan.

Safety and Protective Efficacy of a Candidate Vector-Based Vaccine for Bovine Tuberculosis.

This study presents the results of a survey of the safety and protective efficacy of a candidate vector-based vaccine for bovine tuberculosis, using an influenza vector with the NS1 mutation and expressing M. bovis protective antigens ESAT-6 and TB10.4. We vaccinated Balb/c outbred mice two times at 21 days apart. Our experimental design includes mice immunised with the candidate vaccine with or without adjuvant 15% Montanide Gel. The candidate vaccine's safety was determined by biometric analysis, and protective efficacy was assessed by bacteriological and histological experiments following a virulent M. bovis-8 strain challenge. Our data indicated that the adjuvant-free version of the vaccine ensured complete protection from the M. bovis-8 infection in mice.

Preclinical Evaluation of TB/FLU-04L-An Intranasal Influenza Vector-Based Boost Vaccine against Tuberculosis.

Tuberculosis is a major global threat to human health. Since the widely used BCG vaccine is poorly effective in adults, there is a demand for the development of a new type of boost tuberculosis vaccine. We designed a novel intranasal tuberculosis vaccine candidate, TB/FLU-04L, which is based on an attenuated influenza A virus vector encoding two mycobacterium antigens, Ag85A and ESAT-6. As tuberculosis is an airborne disease, the ability to induce mucosal immunity is one of the potential advantages of influenza vectors. Sequences of ESAT-6 and Ag85A antigens were inserted into the NS1 open reading frame of the influenza A virus to replace the deleted carboxyl part of the NS1 protein. The vector expressing chimeric NS1 protein appeared to be genetically stable and replication-deficient in mice and non-human primates. Intranasal immunization of C57BL/6 mice or cynomolgus macaques with the TB/FLU-04L vaccine candidate induced Mtb-specific Th1 immune response. Single TB/FLU-04L immunization in mice showed commensurate levels of protection in comparison to BCG and significantly increased the protective effect of BCG when applied in a "prime-boost" scheme. Our findings show that intranasal immunization with the TB/FLU-04L vaccine, which carries two mycobacterium antigens, is safe, and induces a protective immune response against virulent M. tuberculosis.

Development and evaluation of a multiplex polymerase chain reaction in real-time for differential diagnosis of Moraxella-induced keratoconjunctivitis in livestock.

Background and Aim: Infectious bovine keratoconjunctivitis (IBK) is a prevalent ocular disease that affects livestock, leading to substantial economic losses due to reduced production and culling of infected animals. Moraxella spp. is common bacterial pathogens that can cause keratoconjunctivitis in livestock. Therefore, rapid and accurate diagnosis is crucial for effective treatment and disease control. This study aimed to develop a multiplex real-time polymerase chain reaction (mRT-PCR) assay for the detection and differentiation of Moraxella bovoculi, Moraxella ovis, and Moraxella bovis. Materials and Methods: Three reference strains of Moraxella as positive controls and 36 lacrimal swab samples collected from cattle were used to evaluate the developed mRT-PCR assay DNA extraction that was performed using the RIBO-sorb DNA/RNA extraction kit. Primers and probes were designed using the SpeciesPrimer pipeline. The annealing temperature, primer and probe concentrations, and sensitivity and specificity of the assay were optimized. Results: An mRT-PCR assay was developed to detect pathogens associated with IBK in cattle on the basis of optimized parameters. The specificity and sensitivity of this assay were confirmed using samples containing individual pathogens (O - M. ovis, B - M. bovis, and BO - M. bovoculi), combinations of two pathogens (O-B, B-BO, and O-BO), and when the DNA of all three pathogens was present in a single reaction (O-B-BO). The analytical sensitivity of mRT-PCR for detecting M. ovis and M. bovoculi DNA was 21 copies or 50 fg per reaction, whereas that for M. bovis was 210 copies or 500 fg per reaction. In addition, this assay has been tested on samples isolated from the affected eyes of cattle in the Akmola region of the Republic of Kazakhstan. Conclusion: For the first time in the Republic of Kazakhstan, the proposed mRT-PCR assay for the simultaneous detection of three Moraxella spp. pathogens has been developed. This assay exhibits the required specificity and high sensitivity for m RT-PCR, facilitating the timely implementation of effective measures for disease control and the prevention of economic losses. These losses are linked to a reduction in livestock breeding value, a reduction in meat and milk production, a reduction in the reproductive performance of heifers, resulting in fewer offspring, as well as costs related to the treatment of affected animals.

Coding Complete Genome Sequence of the SARS-CoV-2 Virus Strain, Variant B.1.1, Sampled from Kazakhstan.

This article describes the results of sequencing and analysis of the entire genome of the SARS-CoV-2 virus sampled in Kazakhstan in 2021. The whole-genome sequence of the strain was 29,751 bp. According to the results of phylogenetic analysis (according to the Pangolin COVID-19 database), the SARS-CoV-2/human/KAZ/B1.1/2021 strain studied here was assigned to variant B.1.1.

Complete Coding Genome Sequence of an Influenza A/H3N8 Equine Virus Isolated in Kazakhstan in 2007.

Here, we reported the complete coding sequence of the influenza A/equine/Otar/3/2007 (H3N8) equine virus, first isolated in Kazakhstan in 2007. The hemagglutinin (HA) sequences of the Kazakhstan isolates appeared to be closely related to viruses isolated in early 2000 in Asia. Phylogenetic analysis characterized the Kazakhstan isolates as a member of the Florida sublineage clade 2 by the HA protein sequence.

Safety and immunogenicity of the first Kazakh inactivated vaccine for COVID-19.

This article describes the results of a preclinical safety and immunogenicity study of QazCovid-in®, the first COVID-19 vaccine developed in Kazakhstan, on BALB/c mice, rats, ferrets, Syrian hamsters and rhesus macaques (Macaca mulatta). The study's safety data suggests that this immunobiological preparation can be technically considered a Class 5 nontoxic vaccine. The series of injections that were made did not produce any adverse effect or any change in the general condition of the model animals' health, while macroscopy and histology studies identified no changes in the internal organs of the BALB/c mice and rats. This study has demonstrated that a double immunization enhances the growth of antibody titers as assessed by the microneutralization assay (MNA) and the enzyme-linked immunosorbent assay (ELISA) in a pre-clinical immunogenicity test on animal models. The best GMT results were assessed in MNA and ELISA 7 days after re-vaccination; however, we noted that GMT antibody results in ELISA were lower than in MNA. A comparative GMT assessment after the first immunization and the re-immunization identified significant differences between model animal groups and a growth of GMT antibodies in all of them; also, differences between the gender groups were statistically significant. Moreover, the most marked MNA immune response to the QazCovid-in® vaccine was seen in the Syrian hamsters, while their SARS-CoV-2-specific antibody activity as assessed with ELISA was the lowest.

Efficacy and safety of an inactivated whole-virion vaccine against COVID-19, QazCovid-in®, in healthy adults: A multicentre, randomised, single-blind, placebo-controlled phase 3 clinical trial with a 6-month follow-up.

Background: Vaccination remains the primary measure to prevent the spread of the SARS-CoV-2 virus, further necessitating the use of effective licensed vaccines. Methods: From Dec 25, 2020, to July 11, 2021, we conducted a multicenter, randomised, single-blind, placebo-controlled phase 3 efficacy trial of the QazCovid-in® vaccine with a 180-day follow-up period in three clinical centres in Kazakhstan. A total of 3000 eligible participants aged 18 years or older were randomly assigned (4:1) to receive two doses of the vaccine (5 µg each, 21 days apart) or placebo administered intramuscularly. QazCovid-in® is a whole-virion formaldehyde-inactivated anti-COVID-19 vaccine, adjuvanted with aluminium hydroxide. The primary endpoint was the incidence of symptomatic cases of the SARS-CoV-2 infection confirmed by RT-PCR starting from day 14 after the first immunisation. The trial was registered with ClinicalTrials.gov NCT04691908. Findings: The QazCovid-in® vaccine was safe over the 6-month monitoring period after two intramuscular immunisations inducing only local short-lived adverse events. The concomitant diseases of participants did not affect the vaccine safety. Out of 2400 vaccinated participants, 31 were diagnosed with COVID-19; 43 COVID-19 cases were recorded in 600 placebo participants with onset of 14 days after the first dose within the 180-day observation period. Only one severe COVID-19 case was identified in a vaccine recipient with a comorbid chronic heart failure. The protective efficacy of the QazCovid-in® vaccine reached 82·0% (95% CI 71.1-88.5) within the 180-day observation period. Interpretation: Two immunisations with the inactivated QazCovid-in® vaccine achieved 82·0% (95% CI 71.1-88.5) protective efficacy against COVID-19 within a 180-day follow-up period. Funding: The work was funded by the Science Committee of the Ministry of Education and Science of Kazakhstan within the framework of the Scientific and Technical Program "Development of a vaccine against coronavirus infection COVID-19". State registration number 0.0927.

Serological exposure in Bactrian and dromedary camels in Kazakhstan to a MERS or MERS-like coronavirus.

Middle East Respiratory Syndrome Coronavirus (MERS-CoV) is a camel-borne zoonotic virus endemic across Eastern Africa and the Middle East, with evidence of circulation in Bangladesh and Mongolia. To determine if MERS-CoV was present in Kazakhstan, in 2017-2018, we collected swabs and sera from Bactrian camels (n = 3124) and dromedary (n = 5083). The total seropositivity was 0.54% in Bactrian camels and 0.24% in dromedaries; however, we did not detect MERS-CoV RNA in swab samples. There was no difference in the probability of infection between species or sex, but younger camels had a higher probability of being seropositive, suggesting a recent introduction of the virus to Kazakhstan. The infection of both camel species indicates that they both may play a role as natural reservoirs. These results reinforce the need for continual surveillance, especially at the camel-human interface to understand the risk of zoonotic exposure.

Development of the Inactivated QazCovid-in Vaccine: Protective Efficacy of the Vaccine in Syrian Hamsters.

In March 2020, the first cases of the human coronavirus disease COVID-19 were registered in Kazakhstan. We isolated the SARS-CoV-2 virus from clinical materials from some of these patients. Subsequently, a whole virion inactivated candidate vaccine, QazCovid-in, was developed based on this virus. To develop the vaccine, a virus grown in Vero cell culture was used, which was inactivated with formaldehyde, purified, concentrated, sterilized by filtration, and then adsorbed on aluminum hydroxide gel particles. The formula virus and adjuvant in buffer saline solution were used as the vaccine. The safety and protective effectiveness of the developed vaccine were studied in Syrian hamsters. The results of the studies showed the absolute safety of the candidate vaccine in the Syrian hamsters. When studying the protective effectiveness, the developed vaccine with an immunizing dose of 5 µg/dose specific antigen protected animals from a wild homologous virus at a dose of 104.5 TCID50 /mL. The candidate vaccine induced the formation of virus-neutralizing antibodies in vaccinated hamsters at titers of 3.3 ± 1.45 log2 to 7.25 ± 0.78 log2, and these antibodies were retained for 6 months (observation period) for the indicated titers. No viral replication was detected in vaccinated hamsters, protected against the development of acute pneumonia, and ensured 100% survival of the animals. Further, no replicative virus was isolated from the lungs of vaccinated animals. However, a virulent virus was isolated from the lungs of unvaccinated animals at relatively high titers, reaching 4.5 ± 0.7 log TCID50/mL. After challenge infection, 100% of unvaccinated hamsters showed clinical symptoms (stress state, passivity, tousled coat, decreased body temperature, and body weight, and the development of acute pneumonia), with 25 ± 5% dying. These findings pave the way for testing the candidate vaccine in clinical human trials.

Development and Evaluation of a Live Attenuated Egg-Based Camelpox Vaccine.

Camelpox is an infectious viral disease of camels reported in all the camel-breeding areas of Africa, north of the equator, the Middle East and Asia. It causes huge economic loss to the camel industry. We developed a live camelpox virus vaccine candidate using an attenuated strain and evaluated its safety, immunogenicity and protective efficacy in camels. The attenuated virus strain was generated from the camelpox wild-type strain M-96 by 40 consecutive passages on the chorioallantoic membrane of 11-day-old embryonated chicken eggs, henceforth called KM-40 strain. Reversion to virulence of the KM-40 strain was evaluated in camels by three serial passages, confirmed its inability to revert to virulence and its overdose administration was also found safe. Studies of immunogenicity and protective efficacy of the candidate vaccine KM-40 strain in camels was carried out using the dose of 5 x 104.0 EID50. Our data showed complete protection against the challenge infection using the virulent wild-type camelpox virus strain M-96 (dose of 105.0 EID50) which was evaluated at 1, 3, 6 and 12 months post vaccination. In summary, our candidate live attenuated egg-based camelpox vaccine strain KM-40 was found safe, protective, and thus has the potential to use safely in field conditions.

Safety and immunogenicity of a QazCovid-in® inactivated whole-virion vaccine against COVID-19 in healthy adults: A single-centre, randomised, single-blind, placebo-controlled phase 1 and an open-label phase 2 clinical trials with a 6 months follow-up in Kazakhstan.

BACKGROUND: A new inactivated whole-virion QazCovid-in® vaccine against COVID-19 was developed from SARS-CoV-2 isolated in Kazakhstan, inactivated by formaldehyde, and adjuvanted with aluminium hydroxide. Phase 1 and 2 clinical trials aimed at assessing the vaccine's safety, immunogenicity, and the duration of immunity induced by the QazCovid-in® vaccine after one or two immunisations. METHODS: From 23.09.2020 to 19.03.2021 we performed a randomised, single-blind, placebo-controlled phase 1 clinical trial and from 18.10.2020 to 17.04.2021 an open-label phase 2 clinical trials of the QazCovid-in® vaccine with a 6 months follow-up at a single centre in Almaty, the Republic of Kazakhstan. Eligible healthy adults aged 18 years and older with no history of laboratory-confirmed SARS-CoV-2 infection were randomly assigned to the treatment groups using a computerised randomisation scheme generator. In the phase 1 clinical trial, two doses of the vaccine (5 µg each) or placebo (0·9% NaCl) were administered intramuscularly to 44 subjects aged 18-50 years, 21 days apart. In the phase 2 trial, 200 healthy participants were randomised into four equal-sized groups according to the age (18-49 or ≥50 years) and either single (day 1) or double (day 1 and 21) vaccination protocol. The primary outcomes were safety and tolerability. The secondary outcome was immunogenicity. The cellular response was measured by a whole-blood cytokine release assay (phase 1 only). The trials were registered with ClinicalTrials.gov NCT04530357. FINDINGS: The QazCovid-in® vaccine was safe and well-tolerated and induced predominantly mild adverse events; no serious or severe adverse events were recorded in both trials. In the phase 1 trial, the percentage of subjects with a fourfold increase of antibody titres (sero conversion) in MNA was 59% after one vaccine dose and amounted to 100% after two doses. Neutralizing antibody titres reached the geometric mean titre (GMT) of 100 after administration of two doses. A statistically significant increase in the levels of pro-inflammatory cytokines after vaccination indicated the Th1-biased response. On day 180, 40% of placebo-treated subjects demonstrated a statistically significant increase in the levels of antibodies measured by both ELISA and MNA, which suggests the infection with SARS-CoV-2. In the phase 2 trial, 100% of subjects aged 18-49 years seroconverted for SARS-CoV-2 on day 21 after the first dose, as indicated by MNA yielding the GMTs of 32 or 30 in the one- and two-dose groups, respectively. Amongst ≥50-year-old subjects, the number of sero conversions in the two- and one-dose groups on day 21 was 94% and 92% with the respective GMTs of 25 and 24. After the second dose, the sero conversion rate reached 100%; however, the GMT was significantly lower when compared with the corresponding value measured in subjects aged 18-49 years (83 vs 143). In both trials, specific antibodies were detected in MNA and ELISA on study day 180, but the titres dropped in comparison to day 42. The results of this study serve as the rationale for the phase 3 study. INTERPRETATION: The QazCovid-in® vaccine is safe and well-tolerated and promotes pronounced humoral immunity which lasts for at least 6 months after double intramuscular immunisation. FUNDING: The work was funded by the Science Committee of the Ministry of Education and Science of the Republic of Kazakhstan within the framework of the Scientific and Technical Program "Development of a vaccine against coronavirus infection COVID-1900 . State registration number ?.0927.

Near-Complete Genome Sequence of an H5N1 Avian Influenza Virus Strain Isolated from a Swan in Southwest Kazakhstan in 2006.

We report the near-complete genome sequence of an influenza H5N1 virus strain isolated from a dead swan on the southeastern Caspian seashore in 2006. The results of the surface protein HA phylogenetic analysis showed that the A/swan/Mangystau/3/2006 virus belongs to the EA-nonGsGD clade.

Complete Genome Sequence of the Highly Pathogenic Strain A/Domestic Goose/Pavlodar/1/05 (H5N1) of the Avian Influenza Virus, Isolated in Kazakhstan in 2005.

Here, we present the complete genome sequence of a highly pathogenic strain of avian influenza A virus/domestic goose/Pavlodar/1/05 (H5N1) (GS/1/05), which belongs to clade 2.2. This strain of the influenza virus was isolated in northern Kazakhstan in 2005.

Safety and immunogenicity of trivalent inactivated influenza vaccine in adults 60 years of age and older: a phase II, a randomized, comparative trial in Kazakhstan.

BACKGROUND: The study was aimed at comparative evaluation of seasonal influenza vaccine RIBSP versus commercial vaccine VAXIGRIP® for immunogenicity and safety in the course of clinical trial phase II on healthy subjects up to 60 years. METHODS: The trial involved 150 subjects in randomized 2:1 groups that received either RIBSP vaccine or comparator vaccine VAXIGRIP®. One dose (0.5 ml) of either vaccine contained 15 µg of hemagglutinin of each influenza virus strain recommended by WHO for the Northern hemisphere in 2016-2017 flu season. The observation period lasted 21 days. The trial was registered at ClinicalTrials.gov identifier NCT03016143. RESULTS: Assessment of immunogenic activity of the vaccine under study showed that in 21 days the portion of participants with 4-fold seroconversions was 80.0% to А/H1N1; 65.0% to А/H3N2 and 64.0% to B virus. Antibody titer increase factor in the group of subjects that received RIBSP vaccine was 13.4 for А/H1N1; 5.2 for А/H3N2 and 5.2 for B virus. The subjects that received RIBSP vaccine demonstrated 88% seroprotection rate against А/H1N1; 75% against А/H3N2 and 61% against B virus. In the course of evaluating the vaccine safety, no serious adverse events were recorded. All changes of laboratory data were slight and single in most cases. All recorded local reactions have been light in character and these have been predicted reactions observed at vaccination against influenza. CONCLUSION: Comparison vaccines RIBSP and VAXIGRIP®, showed similar immunogenic activity. The RIBSP vaccine is safe and immunogenic for the elderly and conforms to international criteria in CPMP/BWP/214/96.

Analysis of the efficacy of an adjuvant-based inactivated pandemic H5N1 influenza virus vaccine.

This paper describes a preclinical study analyzing the immunogenicity and protective efficacy of Kazfluvac®, an adjuvant-based inactivated pandemic influenza A/H5N1 virus vaccine. In this study, laboratory animals (ferrets and mice) were vaccinated by the intramuscular or intraperitoneal route at an interval of 14 days with two doses of the vaccine containing different concentrations of influenza virus hemagglutinin (HA) protein. HA protein without adjuvant (aluminum hydroxide and Merthiolate) was used as a control. As a negative control, we utilized PBS. We assessed the protective efficacy of the candidate vaccine by analyzing the response to challenge with the influenza virus strain A/chicken/Astana/6/05 (H5N1). Our experimental results revealed substantially reduced clinical disease and an increased antibody response, as determined by hemagglutination-inhibition (HAI) test and microneutralization assay (MNA). This study showed that the candidate vaccine is safe and elicits an antigen-dose-dependent serum antibody response. In summary, we determined the optimum antigen dose in a Kazfluvac® adjuvant formulation required for induction of heightened immunogenicity and protective efficacy to mitigate H5N1 disease in experimental animals, suggesting its readiness for clinical studies in humans.

Safety and immunogenicity of the novel seasonal preservative- and adjuvant-free influenza vaccine: Blind, randomized, and placebo-controlled trial.

The producers of influenza vaccines are not capable today to meet the global demand for an influenza vaccine in case of pandemic, so the World Health Organization recommends to develop the own influenza vaccine production in each country. A domestic preservative- and adjuvant-free trivalent split vaccine against seasonal influenza was developed at the Research Institute for Biological Safety Problems. The paper presents the results of assessing safety and immunogenicity of the influenza split vaccine after single immunization of healthy volunteers aged 18-50 years in the course of Phase I Clinical Trials. This study was randomized, blind, and placebo-controlled. The volunteers were intramuscularly vaccinated with a dose of split vaccine or placebo. The study has shown that all local and systemic reactions had low degree of manifestation and short-term character, so there was no need in medication. Serious side effects were not observed. On day 21 post vaccination the portion of vaccinated persons with fourfold seroconversions to influenza А/H1N1pdm09 virus was 100.0%, to influenza А/H3N2 virus-95.5%, to influenza B virus-81.8%, and in placebo group this index was 0%. Seroprotection rates against influenza А/H1N1pdm09, А/H3N2 and B viruses were 95.5, 86.3, and 72.7%, respectively. Geometric mean titers (GMT) of antibodies by day 21 post vaccination reached 175.7 for influenza А/H1N1pdm09 virus, 64.2 for influenza А/H3N2 virus, and 37.6 for influenza B virus; in placebo group GMT growth was not observed. So, the seasonal influenza split vaccine is well tolerated and fits all immunogenicity criteria for human influenza vaccines.