In Vitro Fertilization in Kazakh Whiteheaded Cattle: A Comparative Study.

In vitro fertilization (IVF) technologies have great potential in the preservation of endangered species. In the current study, an IVF experiment was carried out to evaluate whether reproductive technologies are suitable for Kazakh Whiteheaded cattle, aimed at preserving this breed whose population has reduced drastically over the last thirty years. The reproduction characteristics of Kazakh Whiteheaded cows were compared to Aberdeen Angus cows. Transvaginal ultrasound-guided ovum pick up sessions were carried out followed by in vitro embryo production and embryo transfer and pregnancy diagnosis. The total and viable oocytes per OPU procedure were 12.8 ± 1.18 and 8.7 ± 0.85 for the Aberdeen Angus breed, and 8.8 ± 1.04 and 6.2 ± 0.83 for the Kazakh Whiteheaded breed. Similarly, the mean number of cleaved oocytes and morula/blastocyst stage embryos produced by OPU/IVF were 4.8 ± 0.49 and 1.4 ± 0.15 for the Aberdeen Angus breed, and 2.4 ± 0.46 and 0.18 ± 0.05 for the Kazakh Whiteheaded breed (p ≤ 0.02). From fifty Kazakh Whiteheaded donor animals, 2585 oocytes were aspirated following six ovum pick up sessions. One thousand eight hundred and seventy-six (72.5%) oocytes were chosen for maturation and were further fertilized. The number of embryos cleaved was 720 (38.3% out of oocytes fertilized) on day four post-fertilization. Of these cleaved embryos, 56 (7.5%) developed into the late morula/blastocyst stage on day seven post-fertilization, averaging 1.12 embryos per donor animal. Pregnancy was detected in 12 recipients; 4 healthy calves have been born to date. The outcomes of our study have demonstrated that reproductive technologies can be applicable in preserving the endangered Kazakh Whiteheaded cattle. The findings in this report will enhance knowledge of the reproductive characteristics of endangered domestic animals and help develop sophisticated reproductive protocols for animals with unique reproductive mechanisms.

Development of an Inactivated Camelpox Vaccine from Attenuated Camelpox Virus Strain: Safety and Protection in Camels.

This article describes the preparation of an inactivated vaccine from an attenuated strain of camelpox. The attenuated camelpox virus (CMLV) was grown in lamb kidney cells and in Vero cells. CMLV was accumulated to a significantly higher (p ≤ 0.05) titer in lamb kidney cells (7.75 ± 0.08 log TCID50/mL) than in Vero cells (4.00 ± 0.14 log TCID50/mL). During virus inactivation, a concentration of 0.05% beta-propiolactone (BPL) completely inactivated the virus in 6 h at a temperature of 22 ± 1 °C, while a concentration of 0.2% formaldehyde inactivated the virus in 8 h. However, a viral antigen inactivated by BPL was used for vaccine preparation. The inactivated viral antigen was adsorbed with aluminum hydroxide gel, and as a result, an inactivated candidate vaccine was prepared. While the safety of the candidate vaccine was tested in camels and white mice, the protective efficacy of the vaccine was tested only in camels. In the safety evaluation of the inactivated vaccine, the vaccine was not observed to cause any adverse effects in mice and camels. During the immunogenicity study in camels, antibody formation started (0.2 ± 0.16 log2) at Day 21 post-vaccination (PV), and the antibody titer peaked (1.33 ± 0.21 log2) at Day 60 PV and decreased at Day 90 PV (0.50 ± 0.22 log2). Furthermore, no antibodies were detected in vaccinated camels from Days 180 to 365 PV. Camels that received vaccination and were subsequently exposed to wild-type virus evinced a healthy state despite lacking antibodies. In contrast, unvaccinated camels exhibited susceptibility to camelpox upon challenge.

Near-Complete Genome Sequence of a SARS-CoV-2 Variant B.1.1.7 Virus Strain Isolated in Kazakhstan.

This research describes the genome sequence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) obtained from a patient with symptoms of coronavirus disease 2019 (COVID-19) who was infected in the Republic of Kazakhstan. Strain SARS-CoV-2/human/KAZ/Britain/2021 consists of 29,815 nucleotides and belongs to lineage B.1.1.7, according to the Pangolin COVID-19 database.

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.

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.

Duration of Protective Immunity in Sheep Vaccinated with a Combined Vaccine against Peste des Petits Ruminants and Sheep Pox.

In this study, the ability of the combined vaccine against peste des petits ruminants (PPR) (Nigeria strain 75/1) and sheep pox (SPP) (NISKhI strain) to form a protective immune response for 12 months in Kazakh breed fine-fleeced sheep aged 6-12 months was demonstrated. The duration of the protective immunity of immunized sheep from PPR and from SPP was evaluated using a serum neutralization test (SNT), followed by testing of the resistance of vaccinated sheep to infection with the field strain Kentau-7 of the PPRV and the virulent strain A of the SPPV. The PPR antibody response was additionally measured by c-ELISA. A single immunization of sheep with a combined vaccine in a volume of 2.0 mL, containing the PPR and SPP vaccine viruses in the titers of 103.0 TCID50/mL, provided reliable protection of animals from two infections simultaneously for 12 months (observation period). At the same time, in sheep immunized with the combined vaccine, antibodies of PPRV persisted for up to 12 months, with slight fluctuations. The combined vaccine induced 100% clinical protection against the field strain of PPRV and the virulent strain of SPPV in immunized sheep for up to 12 months, while unvaccinated animals became ill with the manifestation of clinical signs specific to PPRV and SPPV.

The combined effects of temperature and relative humidity parameters on the reproduction of Stomoxys species in a laboratory setting.

In this study, Stomoxys species (S. calcitrans, S. sitiens and S. indica) were examined to improve on the current technique for mass rearing using a method of combined incubation parameters. Moreover, the reproductive potential of immature forms at various stages of development was defined. Immature forms of stable flies were incubated according to species. There was no significant difference in the number of immature forms obtained among species incubated under the same conditions. Six incubation parameters were used in combination, at temperatures (T) of 32°C, 27°C and 22°C and relative humidity (RH) of 90% and 70% RH. The combined method resulted in a higher number of eggs hatching at 32°C and 90% humidity as well as an increase in the number of larva pupated and emergence of imago at 27°C and 70% humidity.

Beta-propiolactone inactivated bivalent bluetongue virus vaccine containing Montanide ISA-71VG adjuvant induces long-term immune response in sheep against serotypes 4 and 16 even after 3 years of controlled vaccine storage.

In this study, we developed and evaluated the beta-propiolactone inactivated bivalent bluetongue virus (BTV) serotypes 4 and 16 vaccine delivered with Montanide™ ISA-71VG adjuvant. The safety, stability and immunological profile of the fresh and after three years of long-term storage of the vaccine formulation was analyzed. We observed after long-term storage that the vaccine emulsion was stable as indicated by unchanged pH and viscosity. The stored vaccine formulation induced virus neutralizing antibodies (VNA) in sheep against both the bluetongue virus serotypes at 7-10 day post-vaccination (dpv). VNA titers reached the peak by 60 dpv and detectable during the entire study period. Antibodies against bluetongue virus structural protein VP7 were detected by ELISA in all BTV vaccinated experimental animal groups. Partial clinical protection was observed in vaccinates against challenge virulent BTV-4 and BTV-16 serotypes by 10 dpv, while complete protection was observed at 14 dpv. The levels of viremia was decreased in challenged sheep by 10 dpv while the viremia was undetectable by 14 dpv. In summary, our newly formulated bivalent BTV (BTV-4 and BTV-16) vaccine delivered with Montanide™ ISA-71VG adjuvant was found safe and stable for over three years and induced protective response in sheep.

Duration of protective immunity after a single vaccination with a live attenuated bivalent bluetongue vaccine.

The prevention of bluetongue is typically achieved with mono- or polyvalent modified- live-attenuated virus (MLV) vaccines. MLV vaccines typically elicit a strong antibody response that correlates directly with their ability to replicate in the vaccinated animal. They are inexpensive, stimulate protective immunity after a single inoculation, and have been proven effective in preventing clinical bluetongue disease. In this study, we evaluated the safety, immunogenicity, and efficacy of a bluetongue vaccine against Bluetongue virus serotypes 4 and 16 in sheep. All the animals remained clinically healthy during the observation period. The vaccinated animals showed no clinical signs except fever (>40.8 °C) for 2-4 days. Rapid seroconversion was observed in the sheep, with the accumulation of high antibody titers in the vaccinated animals. No animal became ill after the challenge, indicating that effective protection was achieved. Therefore, this vaccine, prepared from attenuated bluetongue virus strains, is safe, immunogenic, and efficacious.