Wild migratory birds are the principal natural reservoir of influenza A viruses (IAVs) and play a central role in their evolution and global dissemination. Ukraine lies along the Black Sea–Mediterranean Flyway, making surveillance of avian influenza viruses in wild birds essential within a One Health framework, although genomic data from the region remain limited. Between October 2025 and January 2026 In some places you say December 2025 and in other places you say January 2026. I do not understand the differences, 604 cloacal and oropharyngeal swabs and environmental fecal samples were collected from migratory birds and mixed-species flocks in the Tuzlivski lymany National Nature Park and surrounding areas, together with samples from nearby domestic poultry. Influenza A virus was detected by real-time RT-PCR targeting the matrix gene, followed by H5/H7 screening, Oxford Nanopore sequencing of complete hemagglutinin genes, and phylogenetic analysis. Influenza A virus RNA was identified in 30/604 samples (4.97%), whereas all samples were negative for H5 and H7 subtypes. Complete hemagglutinin sequences revealed three low-pathogenic subtypes (H1, H4, and H6) that clustered within established Eurasian lineages closely related to contemporary avian viruses circulating across Europe, Asia, and North America. No influenza A virus or Newcastle disease virus was detected in domestic poultry. These findings provide an important genomic baseline for avian influenza surveillance in Ukraine and demonstrate the value of portable nanopore sequencing and integrated One Health surveillance for monitoring viral diversity and supporting early detection of emerging influenza threats.

Background and Aim: The One Health approach integrates human, animal, plant, and environmental health through multisectoral collaboration and is increasingly recognized as essential for addressing zoonotic diseases, antimicrobial resistance (AMR), food security, and ecosystem degradation. Ukraine has formally adopted One Health principles through national strategies and international partnerships; however, the ongoing full-scale military conflict has profoundly disrupted health, veterinary, and environmental systems, challenging effective implementation. This study aimed to evaluate the current status, achievements, and constraints of the One Health approach in Ukraine, with particular emphasis on the effects of armed conflict on governance, surveillance capacity, and intersectoral coordination, and to outline strategic priorities for strengthening One Health resilience. Materials and Methods: A mixed-methods approach was used, combining bibliometric analysis of Scopus-indexed literature on zoonoses, AMR, food security, and environmental safety with targeted case studies and a review of policy documents. National legal frameworks, international guidelines, and reports from global organizations were systematically analyzed to assess institutional capacity and operational readiness. Results: Ukraine has established a solid policy foundation for One Health, notably through the national Strategy for Biosafety and Biosecurity, which is grounded in the One Health principle and aligned with quadripartite frameworks. Active initiatives address priority zoonoses (rabies, leptospirosis, tuberculosis), AMR surveillance, and food safety. Nevertheless, implementation remains fragmented. Armed conflict has caused extensive damage to laboratories, displaced the workforce, created surveillance blind spots, and disrupted multisectoral communication. AMR trends have intensified due to healthcare strain, while environmental and plant health components remain under-integrated despite their relevance to food security and long-term resilience. The Ukrainian experience demonstrates that policy commitment alone is insufficient in the context of conflict. Effective One Health operationalization requires institutionalized governance mechanisms, interoperable surveillance systems, and sustained investment in human resources and laboratory infrastructure. Environmental and plant health integration remains a critical gap. Conclusion: Reinforcing the One Health framework is essential for Ukraine’s recovery and long-term health security. Sustained international technical and financial support, coupled with national institutionalization of One Health principles, is crucial to rebuilding integrated surveillance, mitigating biological risks, and enhancing resilience in conflict-affected settings.

Using rapid nanopore whole-genome sequencing, we assembled the genome of Bacillus anthracis strain ter21 (5,229,480 bp), a Tsiankovskii-I group isolate cultured from a fatal case in 2021 of a pony from a zoo in Ternopil, Ukraine, identifying virulence plasmid pXO1 that encodes anthrax toxin, and pXO2.

The development of new domestic probiotics targeted at immunomodulatory effects is both timely and holds scientific and practical value. To carry out preclinical studies of the 'Combio' probiotic, which is a mixture of probiotic bacteria Bacillus subtilis, Bacillus licheniformis, Bacillus amyloliquefaciens, Enterococcus faecium and other substances developed by the team of authors of SSRILDVSE, under the conditions of several toxicological experiments on laboratory (white rats, rabbits) and target animals toxico-biochemical parameters were identified. The findings of the trial of the acute toxicity of the 'Combio' probiotic showed that the LD50 value could not be calculated, since the death of laboratory animals was not registered within 14 days after administration; the maximum administered dose of the preparation (by absolute weight) was 30,000.0 mg/kg of body weight, which allows it to be assigned to toxicity class VI – relatively harmless substances (LD50>15,000.0 mg/kg of body weight), and according to the degree of danger to IV class – low-risk substances (LD50>5000.0 mg/kg body weight). When applied to the skin (acute dermal toxicity) and the mucous membrane of the eye of rabbits in doses from 750.0 to 3000.0 mg/kg of body weight, the 'Combio' probiotic did not have an irritating effect, and according to the degree of danger, it can be classified as IV class – low- hazardous substances (LD50>2500.0 mg/kg of body weight). According to the results of extended oral feeding of the probiotic in doses of 1500.0; 7500.0 and 15000.0 mg/kg of feed it showed no evidence of hematological, hepatic, or nephrotoxic effects on the laboratory animals under subacute toxicological conditions. On the contrary, it showed the ability to induce metabolic responses in the bodies of white rats (according to dynamics of hematopoiesis and liver protein-synthesizing functions, especially at therapeutic doses). The dynamics of changes in metabolic indicators during extended 60-day oral feeding of probiotics to target animals and poultry in the dose range showed no evidence of hematological, immunological, or hepatoxic effects and correlated with the hematological indicators, which indicated the restoration of nonspecific resistance and the protein profile in the bodies of experimental animals and a sign of increased metabolic respond and immune responsiveness in pigs organisms. The findings regarding the effect of the new 'Combio' probiotic based on the bacteria Bacillus spp. in several experiments on laboratory and target (pigs) animals give grounds to assert that it is ecologically safe and promising for the result – improving the survival rate of target animals and poultry, increasing their body weight gain, optimizing feed conversion, and enhancing the overall quality of production.

We report the complete genome sequence of an avian orthoavulavirus 13 strain, isolated from a white-fronted goose in the Odesa region of Ukraine in 2013. The detection of avian orthoavulavirus 13 in Ukraine confirms that the geographic distribution of this virus extends beyond Asia.

Amid the devastating war in Ukraine, a parallel battle rages on – that against infectious diseases. The COVID-19 pandemic starkly underlined the vital importance of global health surveillance for emerging and re-emerging human and animal pathogens. This challenge has become even more acute in war-torn regions like Ukraine, where healthcare infrastructure is under siege. For viruses with pandemic potential, such as the influenza virus, worldwide surveillance initiatives are essential in tracking the virus’s movement and spread among both animals and humans. Such monitoring serves dual purposes. Firstly, it guides the formulation of the yearly vaccine, ensuring it remains effective against prevailing strains. Secondly, it alerts livestock farmers in advance, enabling the timely rollout of mitigation measures. Moreover, discerning whether a disease outbreak is of natural origin, or an intentional release becomes critical. To achieve this, the global community must share transparent and timely information through a robust network of laboratories with cutting-edge biotechnological capabilities. The war’s ramifications have severely impacted these efforts. Yet, the stakes are too high to let these facilities fade away. This situation leads to the ultimate question: Who will step up to rebuild Ukraine’s public health and disease research laboratories?

Newcastle disease virus (NDV) infects a wide range of bird species worldwide and is of importance to the poultry industry. Although certain virus genotypes are clearly associated with wild bird species, the role of those species in the movement of viruses and the migratory routes they follow is still unclear. In this study, we performed a phylogenetic analysis of nineteen NDV sequences that were identified among 21,924 samples collected from wild and synanthropic birds from different regions of Ukraine from 2006 to 2015 and compared them with isolates from other continents. In synanthropic birds, NDV strains of genotype II, VI, VII, and XXI of class II were detected. The fusion gene sequences of these strains were similar to strains detected in birds from different geographical regions of Europe and Asia. However, it is noteworthy to mention the isolation of vaccine viruses from synanthropic birds, suggesting the possibility of their role in viral transmission from vaccinated poultry to wild birds, which may lead to the further spreading of vaccine viruses into other regions during wild bird migration. Moreover, here we present the first publicly available complete NDV F gene from a crow (genus Corvus). Additionally, our phylogenetic results indicated a possible connection of Ukrainian NDV isolates with genotype XXI strains circulating in Kazakhstan. Among strains from wild birds, NDVs of genotype 1 of class I and genotype I of class II were detected. The phylogenetic analysis highlighted the possible exchange of these NDV strains between wild waterfowl from the Azov-Black Sea region of Ukraine and waterfowl from different continents, including Europe, Asia, and Africa.

The article presents the results of a molecular genetic study of two isolates of the Pseudorabies virus that were isolated from pigs in Ukraine. Bioinformatic analysis of the gE gene fragment of Aujeszky's disease virus (Pseudorabies virus) isolates was carried out in order to determine the phylogenetic relationships and homology of nucleotide sequences. Fragments of the Aujeszky disease virus genome corresponding to the C-terminal region of the gE gene were selected for sequencing and further analysis. As a result of the conducted studies, it was demonstrated that the nucleotide sequences of the analyzed samples differ from each other by the presence of ACG insert in the tandem repeats region. Comparison of the studied sequences with the sequences of strains/isolates of the Aujeszky's disease virus found in Europe and Asia, presented in the GenBank database, indicates that such an insert is characteristic for the Min-A and HNJZ strains (position 1487 in the gE gene) isolated in Asia. Analysis of the homology of nucleotide sequences showed that the sequence of the gE gene fragment of sample No. 1 is 100% identical to the sequences of strains 89V87 and 00V72 isolated in Belgium. The homology of the nucleotide sequence of the gE gene fragment of sample No. 3 with strains 89V87 and 00V72 was 99.13%. In order to clarify the analyzed samples belonging to a particular genogroup (genetic cluster), a phylogenetic dendrogram was constructed. This demonstrates the phylogenetic relationships between strains/isolates of the Aujeszky's disease virus. It was found that the analyzed samples belong to the genetic cluster uniting European strains/isolates, and the studied isolates are most genetically close to strains 89V87 and 00V72.

New tests for the detection and typing of animal pathogens have been developed for veterinary medicine. Careful systematization is required to determine the place of molecular-based tools’ applications in the existing system of epizootological and epidemiological surveillance. Today, molecular genetic tests, including PCR, are used in veterinary medicine and agriculture for the following purposes:- surveillance and diagnosis of infectious and certain invasive diseases, - typing of animal pathogens, the study of their eco-geographic features, the drift of genetic variability and evolution, - research of molecular mechanisms of the immune response and the host-pathogen interactions, - quality and safety control of agricultural products, including food and feeds, - control of the quality and safety of genetic resources of animals, - control of the circulation of pathogens in the environment, - analysis of the origin and certification of breeds of productive and non-productive animals, etc. The application of molecular genetic methods of monitoring and early diagnosis is regulated by the Manual and Code of the World Organization for Animal Health (WOAH), the Program for the Global Control of Infectious Diseases of the World Health Organization, the guidelines on the monitoring of infectious diseases of animals and the control of the safety of agricultural products of the FAO. A large number of tests based on molecular diagnostic methods are recommended for use in infectious disease control programs, both emerging and economically significant, in the USA, Canada, and the countries of the European Union. This paper summarises the current PCR-based development scope and ways of its implementation in practical veterinary medicine.

African swine fever (ASF) is a viral disease, endemic to Africa, that causes high mortality when introduced into domestic pig populations. Since the emergence of p72-genotype II African swine fever virus (ASFV) in Georgia in 2007, an ASF epidemic has been spreading across Europe and many countries in Asia. The epidemic first reached Ukraine in 2012. To better understand the dynamics of spread of ASF in Ukraine, we analyzed spatial and temporal outbreak data reported in Ukraine between 2012 and mid-2023. The highest numbers of outbreaks were reported in 2017 (N = 163) and 2018 (N = 145), with overall peak numbers of ASF outbreaks reported in August (domestic pigs) and January (wild boars). While cases were reported from most of Ukraine, we found a directional spread from the eastern and northern borders towards the western and southern regions of Ukraine. Many of the early outbreaks (before 2016) were adjacent to the border, which is again true for more recent outbreaks in wild boar, but not for recent outbreaks in domestic pigs. Outbreaks prior to 2016 also occurred predominantly in areas with a below average domestic pig density. This new analysis suggests that wild boars may have played an important role in the introduction and early spread of ASF in Ukraine. However, in later years, the dynamic suggests human activity as the predominant driver of spread and a separation of ASF epizootics between domestic pigs and in wild boars. The decline in outbreaks since 2019 suggests that the implemented mitigation strategies are effective, even though long-term control or eradication remain challenging and will require continued intensive surveillance of ASF outbreak patterns.

Emerging RNA virus infections are a growing concern among domestic poultry industries due to the severe impact they can have on flock health and economic livelihoods. Avian paramyxoviruses (APMV; avulaviruses, AaV) are pathogenic, negative-sense RNA viruses that cause serious infections in the respiratory and central nervous systems. APMV was detected in multiple avian species during the 2017 wild bird migration season in Ukraine and studied using PCR, virus isolation, and sequencing. Of 4090 wild bird samples collected, mostly from southern Ukraine, eleven isolates were grown in ovo and identified for APMV serotype by hemagglutinin inhibition test as: APMV-1, APMV-4, APMV-6, and APMV-7. To build One Health’s capacity to characterize APMV virulence and analyze the potential risks of spillover to immunologically naïve populations, we sequenced virus genomes in veterinary research labs in Ukraine using a nanopore (MinION) platform. RNA was extracted and amplified using a multiplex tiling primer approach to specifically capture full-length APMV-1 (n = 5) and APMV-6 (n = 2) genomes at high read depth. All APMV-1 and APMV-6 fusion (F) proteins possessed a monobasic cleavage site, suggesting these APMVs were likely low virulence, annually circulating strains. Utilization of this low-cost method will identify gaps in viral evolution and circulation in this understudied but important critical region for Eurasia.

Porcine circovirus type 2 (PCV2) is responsible for a number of porcine circovirus-associated diseases (PCVAD) that can severely impact domestic pig herds. For a non-enveloped virus with a small genome (1.7 kb ssDNA), PCV2 is remarkably diverse, with eight genotypes (a–h). New genotypes of PCV2 can spread through the migration of wild boar, which are thought to infect domestic pigs and spread further through the domestic pig trade. Despite a large swine population, the diversity of PCV2 genotypes in Ukraine has been under-sampled, with few PCV2 genome sequences reported in the past decade. To gain a deeper understanding of PCV2 genotype diversity in Ukraine, samples of blood serum were collected from wild boars (n = 107) that were hunted in Ukraine during the November–December 2012 hunting season. We found 34/107 (31.8%) prevalence of PCV2 by diagnostic PCR. For domestic pigs, liver samples (n = 16) were collected from a commercial market near Kharkiv in 2019, of which 6 out of 16 (37%) samples were positive for PCV2. We sequenced the genotyping locus ORF2, a gene encoding the PCV2 viral capsid (Cap), for 11 wild boar and six domestic pig samples in Ukraine using an Oxford Nanopore MinION device. Of 17 samples with resolved genotypes, the PCV2 genotype b was the most common in wild boar samples (10 out of 11, 91%), while the domestic pigs were infected with genotypes b and d. We also detected genotype b/d and b/a co-infections in wild boars and domestic pigs, respectively, and for the first time in Ukraine we detected genotype f in a wild boar from Poltava. Building a maximum-likelihood phylogeny, we identified a sublineage of PCV2 genotype b infections in both wild and domestic swine, suggesting a possible epizootic cluster and an ecological interaction between wild boar and domestic pig populations in northeastern Ukraine.

[This corrects the article DOI: 10.3389/fvets.2021.688078.].

The purpose of this work was to adapt the method for detection of genetic material of PCV-II developed by us for use in the field conditions in the absence of laboratory equipment and proper working conditions. To develop the technique, a liver sample from a pig was used, which was characterized as positive for the presence of PCV-II genetic material. Nucleic acid extraction was performed using an express method of our own modification. The isothermal amplification reaction was carried out using reagents manufactured by Thermo Fisher Scientific (Germany) and BioLabs (Great Britain) following the manufacturers’ recommendations when using the PCV-F3, PCV-B3, PCV-FIP, PCV-BIP primer systems. A WB-4MS water bath (Biosan, Latvia) and a 380 ml thermal mug (ZIZ, Ukraine) were used for isothermal amplification. Freeze dryer ALPHA 1-2 LD plus manufactured by Christ (Germany) was used for lyophilization of the reaction mixture. The reaction temperature was monitored using a ТТЖ-М thermometer (PJSC “Skloprylad”, Ukraine), the temperature was recorded using a SterilDisk logger (Tecnosoft, Italy). Gel Doc XR+ transilluminator (Bio-Rad, USA), SYBR Green I intercalating dye produced by Invitrogen (USA) and portable ultraviolet light source (YATO, Poland) were used to visualize and control the results of isothermal amplification. The previously developed isothermal amplification method was adapted for use in field conditions on the example of the identification of PCV-II. The developed method of isothermal amplification is technically simplified and does not require the use of special laboratory equipment

The etiological agent of infectious ovine epididymitis is Brucella ovis and for its direct indication in clinical samples several PCR protocols are proposed. This study describes a design and selection of the oligonucleotides for real-time PCR targeting conservative BOV_A0504 gene. The specificity of a real-time PCR was validated using 25 B. ovis field isolates and 14 microorganisms of closely related species. The detection limit of B. ovis in bacterial culture was determined as 3.5×101 CFU/mL with Ct value of 37.8. There are no detectable fluorescence signals in the clinical samples from intact animals, whereas bacteriologically confirmed material such as urine and testicle tissue samples were positive. It confirms that the assay is highly specific for detection of B. ovis DNA. Thus, the proposed real-time PCR assay enables fast detection and quantification of B. ovis in clinical material, which can be used as additional test for estimation of the health status of a sheep herd

Abstract A Knowledge, Attitudes, and Practices (KAP) questionnaire was designed to collect information on farmers’ knowledge of ASF and their practices surrounding that could impact the spread of the disease. The questionnaire was distributed, and data collected, from 233 backyard farmers from five selected Oblasts (Rivne, Kharkiv, Odessa, Zakarpattia and Kiev). Kruskal‐Wallis tests were conducted to identify factors that could influence knowledge, and Dunn tests were performed to determine differences between groups when the Kruskal‐Wallis tests were significant. Spearman tests were carried out to explore the association between knowledge and risky practices. Results show that comprehensive knowledge on ASF is not common in backyard farmers and that risky practices that influence the spread of ASF are regularly performed. Of the respondents, 47% felt well‐informed about how ASF can be transmitted and 31.8% felt confident about recognizing clinical signs of ASF. The independent variable “Oblast” was identified as a significant factor (p = 0.0015) associated with differences in knowledge on clinical signs. We demonstrated statistically significant differences of knowledge between backyard farmers from different Oblasts. Knowledge of preventive measures was positively correlated with risky handling practices related to edible pork products (p = 0.0053) and non‐edible pork products (p = 0.0417). In conclusion, our results show that backyard farmers have knowledge gaps on ASF and practice various risky behaviours that might favour the spread of the disease in Ukraine. There are regional differences in ASF knowledge and risky practices that should be taken into consideration in future evidence‐based ASF prevention and control programs, including public awareness activities.

Isolation of the virus from biological material from a two-month-old calf with pathology of the respiratory system from a herd with a morbidity rate of 48% was performed. After detection the presence of IRT antigens in the lungs of the dead animal, the pathogen was isolated on a continuous culture of calf kidney cells, where a characteristic cytopathic effect was observed. The genetic material of the bovine herpesvirus type 1 (Bovine herpesvirus-1, BHV-1) was identified by polymerase chain reaction in the test sample. The virus isolate was adapted to continuous cell cultures of calf kidneys, sheep kidney, cow embryo lung and calf trachea, and the most suitable biological system was determined, where adsorption and reproductive properties of the virus were more pronounced. It was found that the highest titer of infectious activity of BHV-1 isolate (6.1 lg TCD50/cm3) was obtained on continuous culture of lung embryonic cells of a cow embryo after its reproduction during three consecutive passages (observation period)

To date, there is no information regarding the occurrence of porcine circovirus type 3 (PCV-3) in pigs in Ukraine. Aim. The aim of this work was to study the probable occurrence of the little-studied PCV-3 in pigs with different health status in Dnipropetrovsk, Donetsk, Kyiv, and Kharkiv regions of Ukraine. Methods. Blood, semen, liver, spleen, lung samples and nasal swabs of sows and boars of different ages and with different health status, belonging to farms from Dnipro, Donetsk, Kyiv, and Kharkiv regions of Ukraine, were used for the study. PCV-3 genomic material was detected by the standard polymerase chain reaction using specific primers, flanking a fragment of the rep gene of the virus with the length of 418 bp. To visualize the amplicons, horizontal gel electrophoresis was used and ethidium bromide staining after electrophoresis, followed by photographing the gels using Image Lab 5.2.1 software. Results. DNA of PCV-3 was found in two liver samples and four nasal swabs in two different farms, obtained from clinically healthy pigs, which suggests the possibility of the circulation of this infectious agent at the subclinical level of infection at the farm under investigation. No PCV-3 coinfection with the causative agents of porcine reproductive and respiratory syndrome (PRRS), Aujeszky’s disease, PCV-2, and mycoplasmas was found at this farm. Conclusions. Porcine circovirus type 3 (PCV-3) – (a little-studied causative agent of swine disease) was detected in 6 out of 61 samples, originating from two farms in the Kyiv and Kharkiv regions, obtained from clinically healthy animals) for the first time in Ukraine. This indicates possible circulation of the pathogen among pig farms in Ukraine and demonstrates the need to create and implement a target risk analysis, an extensive survey, as well as to develop control measures of the disease spreading (both organizational and technical preventive). Molecular genetic surveying and subsequent monitoring of PCV-3 among domestic and wild animals, which can cross the borders, will give a possibility to determine the risks of its spreading and related economic and epidemiological consequences. The whole-genome DNA sequencing of the detected virus isolates is planned to determine the relation of Ukrainian strains of the virus to other strains circulating in Europe and other parts of the world. Better understanding the risks, epidemiology and pathology, associated with this new virus for the Ukrainian pig breeding industry, will help to prevent and control its further spread and harmful effects.

This work aimed to obtain positive control using recombinant DNA technology for detection by PCR of a new poorly studied pathogen — porcine circovirus type 3. Recombinant positive control was designed using Clone Manager Basic. As a vector in the creation of recombinant control we used plasmid pTZ57R/T, as an insert — a fragment of the gene rep PCV-3 with the length of 418 nucleotide pairs, obtained by classical PCR. Transformation of competent cells of E. coli strain DH5a was carried out by chemical poration, followed by plating on LB-medium with the addition of ampicillin at a final concentration of 100 μg/ml. The selection of E. coli cell colonies was performed by the marker of antibiotic resistance to ampicillin. The presence of a specific insert was checked by PCR with electrophoretic visualization of the results. The developed recombinant positive control can be used for the monitoring of biological samples from pigs for the presence of genetic material PCV-3 using molecular technologies