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Toxoplasma gondii is a common zoonotic intracellular parasite in livestock raised forhuman consumption and is a public health concern. The mode of transmission is ingestion,and meat is considered to be a major vehicle for human and animal infection. AsT. gondii is environmentally transmissible, other important vehicles in particular foranimals include vegetation, soil and water. The seroprevalence of T. gondii infection inpigs is Serbia has been determined in several studies over the past two decades. It hasbeen established that it varies considerably, primarily based on husbandry, with strictlyto mostly indoor animals having a lower prevalence (below 20%) than animals raisedoutdoors, where prevalence exceeds 60%. Experimental data suggests that differentgenotypes of the parasite vary in virulence, but the significance of virulence in terms ofpathology and disease manifestations is still being investigated. Genotypes of T. gondiiisolated from pig tissues in Serbia to date are ToxoDB#1 (archetype II) and ToxoDB#2(archetype III). Archetype II is predominant and, based on historical reports and recentfindings, low to intermediately virulent. The virulence phenotype and mechanisms ofarchetype III, however, have not been extensively studied, but recent data suggests thatits virulence may vary considerably. This review will also summarize the current knowledgeregarding the virulence of archetypes II and III and evaluate it in the context ofthe pig host.

As pork is an important source for Toxoplasma gondii infection, we have analyzed T. gondii genotypes and toxoplasmosis prevalence in pigs in Serbia in the context of production statistics and economics to assess the specific risk to public health. Genotyping was performed using MnPCR-RFLP; T. gondii-specific IgG antibodies were detected using a modified agglutination test (MAT); and statistical data were extracted from official records and provided by government authorities. The results indicate that, from 2006 to 2021, the median number of annually slaughtered pigs was 5.6 million, yet only 36.1% were processed by abattoirs. The remainder were “backyard pigs” slaughtered on family farms and homesteads. Toxoplasmosis seroprevalence in market-weight (MW) pigs prior to 2006 was 15.2%, and was 15.1% in 2019. The seroprevalence in owned city cats, likely infected by livestock meat, was 33.2%. ToxoDB#1 was identified in pig tissues. The results indicate that backyard pigs are the backbone of the industry and provide as much as 60% of the pork in Serbia. The seroprevalence in pigs and city cats shows that farms are reservoirs for the parasite. Thus, innovative means of reducing T. gondii infection designed with backyard farmers in mind are needed to reduce the risk to public health.

Parasites are highly significant pathogens that are transmitted through food. Their specificlife cycles, transmission routes, and usually a lengthy period between infection and the firstsymptoms of the disease make them a substantial risk to public health. Additionally, thereare challenges in detection, diagnosis, and treatment. Toxoplasmosis is considered the mostwidespread parasitic infection on a global scale. It is caused by the protozoan Toxoplasmagondii, one of the most successful parasites of animals and humans due to its ability to parasitizewithin the nuclei of a wide range of hosts. Because of its importance in both veterinaryand human medicine, T. gondii is one of the most extensively studied parasites. Existing datashow seroprevalences differ across continents, countries, and even within states and amongspecific communities. Consuming undercooked meat presents one of the greatest risk factorsfor human infection with the T. gondii parasite, with pork being recognized as a dominantsource of infection.

In Europe, Toxoplasma gondii lineage II is dominant, and ToxoDB#1 the most frequently occurring genotype. The abundance of lineage III genotypes varies geographically and lineage I are rare, yet present in several regions of the continent. Data on the T. gondii population structure in southeastern Europe (SEE) are scarce, yet necessary to appreciate the diversity of the species in Europe. To help fill this gap, we genotyped 67 strains from nine species of intermediate hosts in Serbia by MnPCR-RFLP, determined the population structure, and identified the genotypes using ToxoDB. A neighbor-joining tree was also constructed from the isolates genotyped on nine loci. While 42% of the total genotype population consisted of ToxoDB#1 and ToxoDB#2, variant genotypes of both lineages comprised 46% of the population in wildlife and 28% in domestic animals and humans. One genotype of Africa 4 lineage was detected in a human sample. Interestingly, the findings include one lineage III variant and one II/III recombinant isolate with intercontinental distribution, which appear to be moderately related to South American genotypes. Based on these findings, SEE is a region of underappreciated T. gondii genetic diversity and possible strain exchange between Europe and Africa.

Toxoplasma gondii archetypes II and III are mildly virulent, yet virulence of variant strains is largely unknown. While lineage II dominates in humans in Europe, lineage III strains are present in various intermediate hosts. In Serbia, lineage III represents 24% of the population structure and occurs most frequently in domestic animals, implying a significant presence in the human food web. In this study, the virulence of four genetically distinct lineage III variants was assessed in vivo and in vitro. In vivo, two strains were shown to be intermediately virulent and two mildly virulent, with cumulative mortalities of 69.4%, 38.8%, 10.7%, and 6.8%, respectively. The strain with the highest mortality has previously been isolated in Europe and may be endemic; the strain with the lowest mortality matches ToxoDB#54, while the remaining two represent novel genotypes. Identical alleles were detected at ROP5, ROP16, ROP18, and GRA15. A set of in vitro analyses revealed proliferation and plaque formation as virulence factors. Higher levels of expression of ENO2 in intermediately virulent strains point to enhanced metabolism as the underlying mechanism. The results suggest that metabolic attenuation, and possibly stage conversion, may be delayed in virulent strains.

To what extent did the Covid-19 pandemic affect the tools, priorities and organisation of cultural policies? And did the pandemic enhance the digital aspect of these policies? This paper compares pandemic cultural policy measures in seven European countries to answer these questions. The countries all installed a plurality of mitigating measures, combining grants and subsidies, compensation of lost income, income support and financial flexibility, creating a tendency towards cultural policy turning into economic policy, fiscal policy, and labour market policy. Cultural policies have not been fundamentally challenged by the pandemic, in the sense that it has affected the essential political tools, divisions of labour, or core goals. The responses have confirmed an existing policy structure or enhanced existing developments. The importance of a state-centred or a federalist cultural policy system has not been challenged in a substantial way. Secondly there is little evidence to show a general acceleration of national digital cultural policies.

Toxoplasma gondii is one of the most successful parasites in the world. Its life form found in the environment, where it may stay viable for a year or more, is the oocyst, excreted by members of the family Felidae as the definite host. During the past decades, several waterborne epidemics of toxoplasmosis worldwide indicated the importance of water as a source of T. gondii infection. In Serbia, detection of oocysts in water has not been previously performed and, therefore, there are no data regarding the presence of T. gondii in Serbian rivers. To fill this gap, surface water samples (n=20) were collected from four major rivers and their tributaries throughout Serbia. The samples were concentrated by filtration and total DNA was extracted from the pellet using a commercial kit. Next, the samples were screened for T. gondii DNA by amplification of the 529 bp repeat fragment by conventional PCR. Three positive samples were detected and all three were confirmed by amplification of GRA6 and SAG2 genes. This first detection of T. gondii in river waters in Serbia suggests that the examined rivers are contaminated with this protozoan, which makes them a potential source of infection. In addition, this study has shown that the used PCR methodology can successfully detect T. gondii in water samples.

Toxoplasma gondiiis an obligate intracellular parasite infecting up to one third of the human population. The central event in the pathogenesis of toxoplasmosis is the conversion of tachyzoites into encysted bradyzoites. A novel approach to analyze the structure ofin vivo-derived tissue cysts may be the increasingly used computational image analysis. The objective of this study was to quantify the geometrical complexity ofT.gondiicysts by morphological, particle, and fractal analysis, as well as to determine if it is impacted by parasite strain, cyst age, and host type. A total of 31 images ofT.gondiibrain cysts of four type-2 strains (Me49, and local isolates BGD1, BGD14, and BGD26) was analyzed using ImageJ software. The parameters of interest included diameter, circularity, packing density (PD), fractal dimension (FD), and lacunarity. Although cyst diameter varied widely, its negative correlation with PD was observed. Circularity was remarkably close to 1, indicating a perfectly round shape of the cysts. PD and FD did not vary among cysts of different strains, age, and derived from mice of different genetic background. Conversely, lacunarity, which is a measure of heterogeneity, was significantly lower for BGD1 strain vs. all other strains, and higher for Me49 vs. BGD14 and BGD26, but did not differ among Me49 cysts of different age, or those derived from genetically different mice. The results indicate a highly uniform structure and occupancy of the differentT.gondiitissue cysts. This study furthers the use of image analysis in describing the structural complexity ofT.gondiicyst morphology, and presents the first application of fractal analysis for this purpose. The presented results show that use of a freely available software is a cost-effective approach to advance automated image scoring forT.gondiicysts.

Toxoplasma gondiiis an obligate intracellular parasite infecting up to one third of the human population. The central event in the pathogenesis of toxoplasmosis is the conversion of tachyzoites into encysted bradyzoites. A novel approach to analyze the structure ofin vivo-derived tissue cysts may be the increasingly used computational image analysis. The objective of this study was to quantify the geometrical complexity ofT.gondiicysts by morphological, particle, and fractal analysis, as well as to determine if it is impacted by parasite strain, cyst age, and host type. A total of 31 images ofT.gondiibrain cysts of four type-2 strains (Me49, and local isolates BGD1, BGD14, and BGD26) was analyzed using ImageJ software. The parameters of interest included diameter, circularity, packing density (PD), fractal dimension (FD), and lacunarity. Although cyst diameter varied widely, its negative correlation with PD was observed. Circularity was remarkably close to 1, indicating a perfectly round shape of the cysts. PD and FD did not vary among cysts of different strains, age, and derived from mice of different genetic background. Conversely, lacunarity, which is a measure of heterogeneity, was significantly lower for BGD1 strain vs. all other strains, and higher for Me49 vs. BGD14 and BGD26, but did not differ among Me49 cysts of different age, or those derived from genetically different mice. The results indicate a highly uniform structure and occupancy of the differentT.gondiitissue cysts. This study furthers the use of image analysis in describing the structural complexity ofT.gondiicyst morphology, and presents the first application of fractal analysis for this purpose. The presented results show that use of a freely available software is a cost-effective approach to advance automated image scoring forT.gondiicysts.

Consumption of undercooked or raw pork is considered a significant risk factor for human infection with Toxoplasma gondii. In this study, we investigated the genetic structure of 18 T. gondii strains obtained from slaughter pigs from Northern Serbia (mainly Vojvodina). The examined samples originated from eight pigs from large commercial farms, six backyard pigs and four free-range Mangalica pigs, all found to be positive for either viable T. gondii or T. gondii DNA. Genotyping was attempted from both pig tissues and mouse brains from the bioassays using a multiplex multilocus nested polymerase chain reaction-restriction fragment length polymorphism (Mn-PCR-RFLP) method with seven markers (GRA6,.lt. SAG2, PK-1, BTUB, C22-8, CS3 and Apico). Identification was achieved for nine T. gondii isolates. Seven isolates were classified as type II and two as type III. These results are consistent with previous studies on animal isolates from Serbia as well as with previous reports that type III is more frequently found in samples from Southern Europe than in those from other parts of the continent.

Toxoplasma gondii is one of the most successful parasites on Earth, infecting a wide array of mammals including one third of the global human population. The obligate intracellular protozoon is not capable of synthesizing cholesterol (Chl), and thus depends on uptake of host Chl for its own development. To explore the genetic regulation of previously observed lipid metabolism alterations during acute murine T. gondii infection, we here assessed total Chl and its fractions in serum and selected tissues at the pathophysiological and molecular level, and integrated the observed gene expression of selected molecules relevant for Chl metabolism, including its biosynthetic and export KEGG pathways, with the results of published transcriptomes obtained in similar murine models of T. gondii infection. The serum lipid status as well as the transcript levels of relevant genes in the brain and the liver were assessed in experimental models of acute and chronic toxoplasmosis in wild-type mice. The results showed that acute infection was associated with a decrease in Chl content in both the liver and periphery (brain, peripheral lymphocytes), and a decrease in Chl reverse transport. In contrast, in chronic infection, a return to normal levels of Chl metabolism has been noted. These changes corresponded to the brain and liver gene expression results as well as to data obtained via mining. We propose that the observed changes in Chl metabolism are part of the host defense response. Further insight into the lipid metabolism in T. gondii infection may provide novel targets for therapeutic agents.