Search

The extreme genetic and immunobiological heterogeneity exhibited by the African swine fever virus (ASFV) has been a significant impediment in the development of an efficacious vaccine against this disease. Consequently, the lack of internationally accepted protocols for the laboratory evaluation of candidate vaccines has become a major concern within the scientific community. The formulation of such protocols necessitates the establishment of a consensus at the international level on methods for the determination of homologous and heterologous isolates/strains of ASFV. The present article provides a comprehensive description of biological techniques employed in the classification of ASFV by seroimmunotypes. These techniques involve a holistic evaluation of ASFV isolates/strains based on their antigenic properties as determined by the hemadsorption inhibiting test (HAdI) using type-specific sera and an immunological test (IT) conducted on pigs inoculated with attenuated strains. The article outlines the methods for setting up the HAdI test, an IT on pigs, and the processes involved in the acquisition of type-specific serums for the HAdI test. It is pertinent to note that the definitive classification of seroimmunotype can only be ascertained after conducting an IT on pigs. The findings from the HAdI test or the phylogenetic analysis of the EP402R gene should be considered preliminary in nature.

Understanding the immunological mechanisms of protection and the viral proteins involved in the induction of a protective immune response to the African swine fever virus (ASFV) is still limited. In the last years, the CD2v protein (gp110-140) of the ASFV has been proven to be a serotype-specific protein. Current work is devoted to the investigation of the possibility of creating protection against virulent ASFV strain Mozambique-78 (seroimmunotype III) in pigs previously vaccinated with vaccine strain FK-32/135 (seroimmunotype IV) and then immunized with the pUBB76A_CD2v plasmid, containing a chimeric nucleotide sequence from the CD2v protein gene (EP402R, nucleotides from 49 to 651) from the MK-200 strain (seroimmunotype III). Vaccination with the ASFV vaccine strain FK-32/135 protects pigs from the disease caused by the strain with homologous seroimmunotype—France-32 (seroimmunotype IV). Our attempt to create balanced protection against virulent strain Mozambique-78 (seroimmunotype III) by induction of both humoral factors of immunity (by vaccination with strain FK-32/135 of seroimmunotype IV) and serotype-specific cellular immunity (by immunization with the plasmid pUBB76A_CD2v of seroimmunotype III) was unsuccessful.

The spread of African swine fever (ASF) in Eurasia has forced a return to the development of live vaccines based on naturally or experimentally attenuated strains of the virus including those resulting from genetic manipulations. This process includes evaluation of the immunomodulating properties of the vaccines. In this report we provide our assessment of two tests for immunobiological evaluation of a candidate live vaccine against ASF from the attenuated ASF virus (ASFV) strain KK-202: (i) investigation of the effect of the attenuated ASFV strain KK-202 on the protectiveness of the vaccine ASFV strain FK-32/135 and a vaccine against classical swine fever (CSF) from the strain LK-VNIIVViM; (ii) determination of the phagocytic activity of blood neutrophils in pigs inoculated with ASFV strains differing in virulence. A simultaneous or sequential inoculation of attenuated strain KK-202 (seroimmunotype II) and vaccine strain FK-32/135 (seroimmunotype IV) into pigs resulted in the loss of protection against the virulent strain France-32 (seroimmunotype IV). Following the simultaneous or sequential inoculations of the ASFV strain KK-202 and the CSF virus (CSFV) vaccine produced from the strain LK-VNIIVViM, the neutralizing antibody titers against the CSFV observed in the experimental groups (after vaccination and after the challenge infection with the virulent CSFV strain Shimen) were not different from those found in animals of the control group. The phagocytic activity of blood neutrophils was shown to increase from 30% in the norm to 50%–94% depending on the virulence of the ASFV strains inoculated into pigs. The results of this work demonstrate the ability of the attenuated ASFV strains to modulate the development of the cellular link of protective immunity without negative impact on the humoral immune response. The informative value of the described immunobiological tests in vivo and in vitro seems to be a more preferable alternative in comparison to the commonly used in vitro tests, which do not always correlate with the development of protection against ASF.

African swine fever virus (ASFV) is an extremely genetically and phenotypically heterogeneous pathogen. Previously, we have demonstrated that experimental inoculation of pigs with an attenuated strain, Katanga-350 (genotype I, seroimmunotype I) (ASFV-Katanga-350), can induce protective immunity in 80% of European domestic pigs against the homologous virulent European strain Lisbon-57. At least 50% of the surviving pigs received protection from subsequent intramuscular infection with a heterologous virulent strain, Stavropol 01/08 (genotype II, seroimmunotype VIII) (ASFV-Stavropol 01/08). In this study, we assessed clinical signs, the levels of viremia, viral DNA, anti-ASFV antibodies and post-mortem changes caused by subsequent intramuscular injection with ASFV-Katanga-350 and heterologous ASFV-Stavropol 01/08. Inoculation of pigs with the ASFV-Katanga-350 did not protect animals from the disease in the case of the subsequent challenged ASFV-Stavropol 01/08. However, 40% of pigs were protected from death. Moreover, the surviving animals showed no pathomorphological changes or the presence of an infectious virus in the organs after euthanasia at 35 days post challenging. The ability/inability of attenuated strains to form a certain level of protection against heterologous isolates needs a theoretical background and experimental confirmation.

African swine fever (ASF) is an emerging viral contagious disease affecting domestic pigs (DP) and wild boar (WB). ASF causes significant economic damage to the pig industry worldwide due to nearly 100% mortality and the absence of medical treatments. Since 2019, an intensive spread of ASF has been observed in the Russian Far East region. This spread raises concerns for epidemiologists and ecologists given the potential threat to the WB population, which is an essential member of the region's wild ungulates and provides a notable share of food resources for predatory species. This study aims to determine the genotype of ASF virus circulating in the region, reveal the spatio-temporal patterns of the ASF outbreaks' emergence, and assess the potential reduction of the regional fauna because of expected depopulation of WB. The first historical case of ASF in the study region was caused by an African swine fever virus (ASFV) isolated from DPs and belonging to Genotype 2, CVR1; IGR-2 (TRS +). Sequencing results showed no significant differences among ASFV strains currently circulating in the Russian Federation, Europe, and China. The spatiotemporal analysis with the space-time permutations model demonstrated the presence of six statistically significant clusters of ASF outbreaks with three clusters in DPs and one cluster in WBs. DP outbreaks prevail in the north-west regions of the study area, while northern regions demonstrate a mixture of DP and WB outbreaks. Colocation analysis did not reveal a statistically significant pattern of grouping of one category of outbreaks around the others. The possible damage to the region's fauna was assessed by modeling the total body mass of wild ungulates before and after the wild boars' depopulation, considering a threshold density of WB population of 0.025 head/km2, according to the currently in force National Plan on the ASF Eradication in Russia. The results suggest the total mass of ungulates of the entire study region will likely decrease by 8.4% (95% CI: 4.1–13.0%), while it may decrease by 33.6% (19.3–46.1%) in the Primorsky Krai, thereby posing an undeniable threat to the predatory species of the region.

The African swine fever virus (ASFV) is the cause of a recent pandemic that is threatening the global pig industry. The virus infects domestic and wild pigs and manifests with a variety of clinical symptoms, depending on the strain. No commercial vaccine is currently available to protect animals from this virus, but some attenuated and recombinant live vaccine candidates might be effective against the disease. This article describes the immunobiological characteristics of one such candidate—the laboratory-attenuated ASFV strain, Katanga-350—which belongs to genotype I. In this study, we assessed clinical signs and post-mortem changes, the levels of viremia and the presence of viral DNA caused by injection of ASF virus strains Katanga-350, Lisbon-57, and Stavropol 08/01. Intramuscular injection of this strain protected 80% of pigs from a virulent strain of the same genotype and seroimmunotype (Lisbon-57). At least 50% of the surviving pigs received protection from subsequent intramuscular infection with a heterologous (genotype II, seroimmunotype VIII) virulent strain (Stavropol 08/01). Virus-specific antibodies were detectable in serum and saliva samples between 8–78 days after the first inoculation of the Katanga-350 strain (the observational period). The results suggested that this strain could serve as a basis for the development of a recombinant vaccine against ASF viruses belonging to seroimmunotype I.

Leptospirosis is a re-emerging zoonotic infectious disease caused by pathogenic bacteria of the genus Leptospira. Regional differences in the disease manifestation and the role of ecological factors, specifically in regions with a subarctic and arctic climate, remain poorly understood. We here explored environmental and socio-economic features associated with leptospirosis cases in livestock animals in the Russian Arctic during 2000–2019. Spatial analysis suggested that the locations of the majority of 808 cases were in “boreal” or “polar” climate regions, with “cropland,” “forest,” “shrubland,” or “settlements” land-cover type, with a predominance of “Polar Moist Cropland on Plain” ecosystem. The cases demonstrated seasonality, with peaks in March, June, and August, corresponding to the livestock pasturing practices. We applied the Forest-based Classification and Regression algorithm to explore the relationships between the cumulative leptospirosis incidence per unit area by municipal districts (G-rate) and a number of socio-economic, landscape, and climatic factors. The model demonstrated satisfactory performance in explaining the observed disease distribution (R2 = 0.82, p < 0.01), with human population density, livestock units density, the proportion of crop area, and budgetary investments into agriculture per unit area being the most influential socio-economic variables. Climatic factors demonstrated a significantly weaker influence, with nearly similar contributions of mean yearly precipitation and air temperature and number of days with above-zero temperatures. Using a projected climate by 2100 according to the RCP8.5 scenario, we predict a climate-related rise of expected disease incidence across most of the study area, with an up to 4.4-fold increase in the G-rate. These results demonstrated the predominant influence of the population and agricultural production factors on the observed increase in leptospirosis cases in livestock animals in the Russian Arctic. These findings may contribute to improvement in the regional system of anti-leptospirosis measures and may be used for further studies of livestock leptospirosis epidemiology at a finer scale.

Elizabethkingia anophelis is an emerging multidrug-resistant pathogen that causes severe nosocomial and community-acquired infections worldwide. We report the first case of E. anophelis isolation in Russia and the first isolation from raw cow’s milk. The ML-44 demonstrated resistance to 28 antimicrobials of 33 tested in the disk-diffusion test. Whole genome-based phylogeny showed ML-44 strain clustered together with the F3201 strain isolated from a human patient in Kuwait in 1982. Both strains were a part of the “endophytica” clade. Another clade was formed by subsp. anophelis strains. Each of the E. anophelis compared genomes carried 18 to 21 antibiotic resistance determinants. The ML-44 chromosome harbored nine efflux system genes and three beta-lactamase genes, along with six other antimicrobial resistance genes. In total, 72 virulence genes were revealed. The set of virulence factors was quite similar between different E. anophelis strains and included LPS and capsule encoded genes, type IV pili, oxidative stress response genes, and genes encoding TIVSS and TVISS effectors. The particular interest caused the mip and zmp1 gene homologs, which can be essential for intracellular survival. In sum, our findings suggest that raw milk might be a source of E. anophelis harboring a set of virulence factors and a broad resistance to generally used antimicrobials.

Totally, 45 L. monocytogenes strains isolated from meat, poultry, dairy, and fish products in the Central European part of Russia in 2001–2005 and 2019–2020 were typed using a combined MLST and internalin profile (IP) scheme. Strains belonged to 14 clonal complexes (CCs) of the phylogenetic lineages I and II. Almost half of the strains (20 of 45) belonged to six CCs previously recognized as epidemic clones (ECs). ECI and ECV strains were isolated during both studied periods, and ECII, ECIV, ECVI, and ECVII strains were isolated in 2001–2005, but not in 2019–2020. ECI, ECIV, ECV, and ECVII strains were isolated from products of animal origin. ECII and ECVI were isolated from fish. Testing of invasion efficiencies of 10 strains isolated in different years and from different sources and belonging to distinct CCs revealed a statistically significant difference between phylogenetic lineage I and II strains but not between ECs and non-EC CCs or strains differing by year and source of isolation. Strains isolated in 2001–2005 were characterized by higher phylogenetic diversity and greater presentation of ECs and CCs non-typical for natural and anthropogenic environments of the European part of Russia comparatively to isolates obtained in 2019–2020.Closing of the Russian market in 2019–2020 for imported food might be responsible for these differences.

Susceptibility of 117 L. monocytogenes strains isolated during three time periods (1950–1980; 2000–2005, and 2018–2021) to 23 antibiotics was tested by the disk diffusion method. All strains were sensitive to aminoglycosides (gentamicin, kanamycin, neomycin, streptomycin), glycopeptides (vancomycin and teicoplanin), clarithromycin, levofloxacin, amoxicillin/clavulanic acid, and trimethoprim/sulfamethoxazole. Resistance to clindamycin was observed in 35.5% of strains. Resistance to carbapenems, imipenem and meropenem was found in 4% and 5% of strains, respectively. Resistance to erythromycin, penicillin G, trimethoprim, and ciprofloxacin was found in 4%, 3%, 3%, and 2.5% of strains, respectively. Resistance to tylosin, ampicillin, enrofloxacin, linezolid, chloramphenicol, and tetracycline was found in less than 2%. Three strains with multiple antibiotic resistance and 12 strains with resistance to two antibiotics were revealed. Comparison of strains isolated in different time periods showed that the percentage of resistant strains was the lowest among strains isolated before 1980, and no strains with multiple antibiotic resistance were found among them. Statistical analysis demonstrated that the temporal evolution of resistance in L. monocytogenes has an antibiotic-specific character. While resistance to some antibiotics such as ampicillin and penicillin G has gradually decreased in the population, resistance to other antibiotics acquired by particular strains in recent years has not been accompanied by changes in resistance of other strains.

This article summarizes the study results on the generation of attenuated strains of African swine fever virus (ASFV) of seroimmunotypes I–VIII and the creation of live vaccines for temporary protection of pigs during a period of epizootics in the surveillance zone (a zone adjacent to the area of outbreak). These studies were initiated at the Federal Research Center for Virology and Microbiology (FRCVM, formerly VNIIVViM) at the time of introduction of the pathogen to the Iberian Peninsula in the middle of the 20th century. The developed experimental vaccines against ASFV seroimmunotypes I–V provided protection against virulent strains of homologous seroimmunotypes by day 14 after vaccination, lasting at least four months.

This review presents comparative results of simultaneously conducted studies on proteins responsible for the haemadsorption and serotype-specific properties of African swine fever virus (ASFV). An ASFV gene EP402R (or LMW8-DR) encoding protein CD2v homologous to murine, human or porcine T-cell adhesive receptor was found. The CD2v was shown to be directly involved into a haemadsorption process, and expressed in ASFV-infected cells as a glycoprotein with a molecular weight of approximately 105-110 kDa. In the presence of a glycosylation inhibitor, tunicamycin, its molecular weight is about 42 kDa. In ASFV-infected cells labeled with 3H-glucosamine or 14C-sodium acetate, a virus-specific major glycoprotein with a molecular weight of 110-140 kDa (gp 110-140) was identified using radioimmunoprecipitation assay. Using ASFV reference strains belonging to seroimmunotypes I-IV and the corresponding antisera active in haemadsorption inhibition assay (HADIA), we determined that gp 110-140 defines the serotype specificity. Genotyping on the basis of the genetic locus encoding the CD2v and a C-type lectin protein also showed a concurrence with the grouping of ASFV isolates and strains based on their seroimmunotypes. Immunization of pigs with the gp 110-140 within liposomes, or a recombinant haemagglutinin (CD2v) protected 67 to 100% of animals from death due to their subsequent infection with homologous virulent ASFV strains. Based on the physico-chemical and biological characteristics of the gp 110-140 and CD2v it is suggested that they are one and the same virus-specific glycoprotein crucial for induction of the immunological protection against ASF. Key words: ASFV; seroimmunotypes; serotype; glycoproteins; gp 110-140; CD2v; protectivity SEROTIPNO-SPECIFIČEN IN HEMADSORPCIJSKI PROTEIN VIRUSA AFRIŠKE PRAŠIČJE KUGE A.D Povzetek: Pregledni članek predstavlja primerjavo rezultatov sočasno izvedenih raziskav o beljakovinah, ki so pomembne za hemadsorpcijo in serotipno specifične lastnosti virusa afriške prašičje kuge (ASFV; iz angl. african swine fever virus). Pri virusu ASFV je bil odkrit gen EP402R (imenovan tudi LMW8-DR), ki kodira beljakovino CD2v, ki je homologna glodavskemu, človeškemu in prašičjemu T-celičnemu adhezivnemu receptorju. Pokazalo se je, da je CD2v neposredno vpletena v proces hemadsorpcije in je izražena v celicah, okuženih z ASFV kot glikoprotein z molekulsko maso okrog 105-110 kDa. V prisotnosti zaviralca glikozilacije tunicamicina je njegova molekulska masa približno 42 kDa. V celicah, okuženih z ASFV, označenih s 3H-glukozaminom ali 14C-natrijevim acetatom, je bil s testom radioimunoprecipitacije ugotovljen virusno specifični osrednji glikoprotein z molekulsko maso 110-140 kDa (gp 110-140). Z uporabo referenčnih sevov ASFV, ki pripadajo seroimunotipom I-IV, in ustreznim antiserumom, ki so bili aktivni pri preizkusu zaviranja hemadsorpcije (HADIA), smo ugotovili, da gp 110-140 določa specifičnost serotipa. Genotipizacija na osnovi genskega lokusa, ki kodira CD2v in C-tip lektinske beljakovine, je pokazala soizražanje s skupino izolatov in sevov ASFV na podlagi njihovih seroimunotipov. Imunizacija prašičev z gp 110-140 v liposomih ali z rekombinantnim hemaglutininom (CD2v) je zaščitila od 67 do 100 odstotkov živali pred smrtjo zaradi njihove naknadne okužbe z virulentnimi sevi ASFV. Na podlagi fizikalno-kemičnih in bioloških značilnosti beljakovin gp 110-140 in CD2v menimo, da gre za isti virusni glikoprotein, ki je ključnega pomena za vzpodbuditev imunološke zaščite pred ASF. Ključne besede: ASFV; seroimunotipi; serotip; glikoproteini; gp 110-140; CD2v; zaščita

Listeriosis is one of the most significant humans and animals foodborne infectious diseases. Here, we characterized 48 Listeria monocytogenes strains isolated in the territory of inner Eurasia during the second half of the 20th century. A total of 23 strains (52.3%) were susceptible to the nine antibiotics tested, 30.43%, 15.22%, and 8.7% were resistant penicillin G, ampicillin, and enrofloxacin, respectively. We applied the multilocus sequence typing (MLST) scheme to determine the phylogenetic positions of the strains. All but one strain belonged to the II phylogenetic lineage, and the majority of the strains belonged to one of the previously described clonal complexes (CCs). More than 60% of the strains belonged to the clonal complex CC7 that prevailed among all sources, including cattle (58%), small ruminants (64%), rodents (71%), and humans (50%). Further, CC7, CC101, and CC124 were found among human isolates. The MLST scheme was supplemented with virulence gene analysis. In total, eight inlA, six inlB, and six inlC allelic variants were found, and all but one strain carried one of the two inlE alleles. Most strains (62.5%) belonged to the same multivirulence locus sequence typing (MvLST) type, which includes CC7, inlA allele 4, inlB allele 14, inlC allele 6, and inlE allele 8.

African swine fever (ASF) is an infectious disease of domestic and wild pigs of all breeds and ages, with the acute form of the disease being characterized by high fever, hemorrhages in the reticuloendothelial system and a high mortality rate. Registered safe and efficacious ASF vaccines are not available. The development of experimental ASF vaccines, particularly live attenuated, have considerably intensified in the last years. There is much variability in experimental approaches undertaken by laboratories attempting to develop first generation vaccines, rendering it difficult to interpret and make comparisons across trials. ASF virus (ASFV) genotyping does not fully correlate with available cross-protection data and may be of limited value in predicting cross-protective vaccine efficacy. Recently, ASFV strains were assigned to a respective nine groups by seroimmunotype (from I to IX): in vivo the grouping is based on results of cross protection of pigs survived after their infection with a virulent strain (bioassay), while in vitro this grouping is based on hemadsorption inhibition assay (HADIA) data. Here we demonstrate the antigenic and protective properties of two attenuated ASFV strains MK200 and FK-32/135. Pronounced differences in the HADIA and in immunological test in animals allow us to consider them and the corresponding reference virulent strains of the ASFV of Mozambique-78 (seroimmunotype III, genotype V) and France-32 (seroimmunotype IV, genotype I) as useful models for studying the mechanisms of protective immunity and evaluation of the candidate vaccines.