Polish Strains Research Articles

Intensified Circulation of Echovirus 11 after the COVID-19 Pandemic in Poland: Detection of a Highly Pathogenic Virus Variant.

After the first phase of the COVID-19 pandemic in Europe, a new highly pathogenic variant of echovirus 11 (E11) was detected. The aim of this study was to analyze the genetic diversity of Polish E11 environmental and clinical strains circulating between 2017 and 2023 as well as compare them with E11 strains isolated from severe neonatal sepsis cases reported in Europe between 2022 and 2023. Additionally, the study explores the effectiveness of environmental monitoring in tracking the spread of new variants. For this purpose, the complete sequences of the VP1 capsid protein gene were determined for 266 E11 strains isolated in Poland from 2017 to 2023, and phylogenetic analysis was performed. In the years 2017-2023, a significant increase in the detection of E11 strains was observed in both environmental and clinical samples in Poland. The Polish E11 strains represented three different genotypes, C3, D5 and E, and were characterized by a high diversity. In Poland, the intensive circulation of the new variant E11, responsible for severe neonatal infections with a high mortality in Europe, was detected in the years 2022-2023. This investigation demonstrates the important role of environmental surveillance in the tracking of enteroviruses circulation, especially in settings with limited clinical surveillance.

Prevalence of coronaviruses in European bison (Bison bonasus) in Poland

Coronaviruses have been confirmed to infect a variety of species, but only one case of associated winter dysentery of European bison has been described. The study aimed to analyze the prevalence, and define the impact on the species conservation, the source of coronavirus infection, and the role of the European bison in the transmission of the pathogen in Poland. Molecular and serological screening was performed on 409 European bison from 6 free-ranging and 14 captive herds over the period of 6 years (2017–2023). Presence of coronavirus was confirmed in one nasal swab by pancoronavirus RT-PCR and in 3 nasal swab samples by bovine coronavirus (BCoV) specific real time RT-PCR. The detected virus showed high (> 98%) homology in both RdRp and Spike genes to BCoV strains characterised recently in Polish cattle and strains isolated from wild cervids in Italy. Antibodies specific to BCoV were found in 6.4% of tested samples, all originating from free-ranging animals. Seroprevalence was higher in adult animals over 5 years of age (p = 0.0015) and in females (p = 0.09). Our results suggest that European bison play only a limited role as reservoirs of bovine-like coronaviruses. Although the most probable source of infections in the European bison population in Poland is cattle, other wild ruminants could also be involved. In addition, the zoonotic potential of bovine coronaviruses is quite low.

The Role of Bacteria in Acute Oak Decline in South-West Poland.

The process of multifactorial oak disease has been of interest to scientists from all over the world for many years. Recently, a new phenomenon has been added to the model related to oak decline, acute oak dieback, which causes oak decline and was first described in the UK. This study presents research on this phenomenon in the area of the largest oak stand complex in Poland, the so-called Krotoszyn Plateau. This work was carried out in two stages. In the first stage, 54 trees were tested for the presence of bacteria using molecular biology (real-time PCR). Subsequently, a tissue fragment was taken for inoculation from the trees in which the presence of B. goodwinii and G. quercinecans bacteria was confirmed. The isolates obtained were used to test Koch's postulates and for biochemical analyses for Polish strains. In addition, the results obtained were also compared with the presence of A. biguttatus, which is considered a bacterial vector, which, in turn, confirmed that the bacteria responsible for the AOD phenomenon can also be present in trees not inhabited by this insect. The results obtained confirmed the presence of bacteria and their potential to cause necrosis in oaks, which fits into the model of the spiral disease that has been causing mass mortality of oaks in this naturally and economically valuable area since the 1980s.

Genetic Diversity of the LTR Region of Polish SRLVs and Its Impact on the Transcriptional Activity of Viral Promoters.

A long terminal repeat (LTR) plays an indispensable role in small ruminant lentivirus (SRLV) gene expression. In this study, we present the LTR sequence of Polish SRLVs representing different subtypes, and analyzed their impact on SRLV promoter activity, as measured in transient transfection assays. Although certain nucleotide motifs (AML(vis), TATA box and the polyadenylation site (AATAAA)) were conserved across sequences, numerous mutations within the LTR sequences have been identified. Single nucleotide polymorphisms (SNPs) were detected in both regulatory (AP-1, AP-4, Stat and Gas) and non-regulatory sequences, and subtype-specific genetic diversity in the LTR region of Polish SRLVs was observed. In vitro assays demonstrated subtype-specific functional differences between the LTR regions of distinct SRLV subtypes. Our results revealed that the promoter activity of Polish strains was lower (1.64-10.8-fold) than that noted for the K1514 reference strain; however, the differences in most cases were not statistically significant. The lowest promoter activity was observed for strains representing subtype A5 (mean 69.067) while the highest promoter activity was observed for strain K1514 representing subtype A1 (mean 373.48). The mean LTR activities of strains representing subtypes A12, A17, A23, A18 and A24 were 91.22, 137.21, 178.41, 187.05 and 236.836, respectively. The results of the inter-subtype difference analysis showed that the promoter activity of strains belonging to subtype A5 was significantly lower than that for subtype A12 strains (1.32-fold; p < 0.00). The promoter activities of the A5 strain were 1.98-fold and 2.58-fold less active than that of the A17 and A23 strains, and the promoter activities of A12 strains were 1.955 and 1.5 times lower than the promoter activity of A23 and A17 strains, respectively. Furthermore, the promoter activity of A17 strains was 1.3 lower than the promoter activity of A23 strains. Our findings suggest that subtype-specific genetic diversity, mainly in the transcription factor's binding sites, has an impact on their transcriptional activity, producing a distinct activity pattern for the subtypes. This study provides new information that is important for better understanding the function of the SRLV LTR. However, further research including more strains and subtypes as well as other cell lines is needed to confirm these findings.

Phylogenetic Analysis of Small Ruminant Lentiviruses Originating from Naturally Infected Sheep and Goats from Poland Based on the Long Terminal Repeat Sequences.

Previous gag and env sequence studies placed Polish small ruminant lentiviruses (SRLVs) isolated from sheep and goats in subtypes B1, B2, A1, A5, A12, A13, A16-A18, A23, A24 and A27. This study extended the genetic/phylogenetic analysis of previously identified Polish SRLV strains by contributing long terminal repeat (LTR) sequences. A total of 112 samples were analysed. Phylogenetic analyses were carried out on the LTR fragment using the neighbour-joining, maximum likelihood, and unweighted pair group method with arithmetic mean methods. Polish caprine and ovine LTR sequences clustered within group A and grouped in at least 10 clusters (subtypes A1, A5, A12, A13, A16-A18, A23, A24 and A27). Most of the Polish strains (78%) belonged to the same subtype by the indication of the gag, env and LTR genomic regions. Discrepancies in affiliation depending on the particular sequence were observed in 24 (21%) strains, most of which came from mixed-species flocks where more than one SRLV genotype circulated. Sequences of the LTR reflected subtype-specific patterns. Several subtype-specific markers were identified, e.g. a unique substitution of T to A in the fifth position of the TATA box in A17, A27, A20 and B3. This study provides valuable insights into the genetic diversity of SRLV field strains in Poland, their phylogenetic relationships and their position in the recently established SRLV classification. Our results confirmed the existence of the ten subtypes listed and the readier emergence of new SRLV variants in mixed-species flocks.

Characterisation of ORF3, M, N and E Gene Sequences of Porcine Epidemic Diarrhoea Virus from Domestic Pigs in Poland.

Porcine epidemic diarrhoea virus (PEDV) is an enteric pathogen causing porcine epidemic diarrhoea and acute gastroenteritis in pigs of all ages. Previous analysis of the viral genome of PEDV in Poland was only based on the spike protein (S) gene sequences and no analysis of other genes has been performed. The aim of this study was to analyse the envelope (E), membrane (M) and nucleocapsid (N) protein and open reading frame 3 (ORF3) gene sequences. Viral RNA from 18 Polish pig faecal samples that were quantitative reverse transcription PCR-positive for PEDV was analysed in four genomic regions (E, M, N and ORF3). Phylogenetic analysis based on these regions' sequences revealed that Polish PEDV isolates were highly related and were clustered into group G2a across the four genes compared. Moreover, the Polish strains were located in distinct subclusters on the phylogenetic trees, which suggests the presence of at least three independently evolving PEDV genetic lines circulating in Poland. The occurrence of unique mutations in the sequences of Polish PEDV strains suggests that PEDV continues to undergo evolutionary processes, accumulating the mutations necessary for viral fitness in its natural hosts. The Polish PEDV strains differed genetically from the CV777 vaccine strain, suggesting the risk of relatively low vaccine efficacy if this strain is used. Our results promote a better understanding of the genetic diversity of PEDV field isolates in Poland and highlight the importance of molecular characterisation of PEDV field strains for the development of an effective vaccine against PEDV.

Avian Bornaviruses in Wild Aquatic Birds of the Anseriformes Order in Poland.

Bornaviruses are a diverse family of viruses infecting various hosts, including birds. Aquatic bird bornavirus 1 (ABBV-1) and aquatic bird bornavirus 2 (ABBV-2) have been found in wild waterfowl but data on their prevalence are scarce. To gain knowledge on the occurrence of ABBVs in Poland, samples originating from dead birds of the Anseriformes order collected in 2016–2021 were tested with a real time RT-PCR method targeting the ABBVs genome. A total of 514 birds were examined, including 401 swans, 96 ducks and 17 geese. The presence of ABBV-1 RNA was detected in 52 swans (10.1% of all tested birds) from 40 different locations. No positive results were obtained for ducks and geese. Sequences of about 2300 bases were generated for 18 viruses and phylogenetic analysis was performed. A relatively low genetic diversity of the examined ABBV-1 strains was observed as all were gathered in a single cluster in the phylogenetic tree and the minimum nucleotide identity was 99.14%. The Polish strains were closely related to ABBV-1 identified previously in Denmark and Germany, but a limited number of sequences from Europe hinders the drawing of conclusions about interconnections between Polish and other European ABBVs. The results of the present study provide new insights into the distribution and genetic characteristics of ABBVs in wild birds in Europe.

Molecular characterization of porcine epidemic diarrhoea virus (PEDV) in Poland reveals the presence of swine enteric coronavirus (SeCoV) sequence in S gene.

Porcine epidemic diarrhoea (PED) is a highly contagious enteric viral disease of pigs with a high morbidity and mortality rate, which ultimately results in huge economic losses in the pig production sector. The etiological agent of this disease is the porcine epidemic diarrhoea virus (PEDV) which is an enveloped, positive single-stranded RNA virus. The aim of this study was to perform molecular characterization of PEDV to identify the strains circulating in Poland. In this study, 662 faecal samples from 2015 to 2021 were tested with reverse transcription quantitative real-time PCR (RT-qPCR) and the results showed that 3.8% of the tested samples revealed a positive result for PEDV. A phylogenetic analysis of the complete genome and complete S gene sequences showed that Polish PEDV strains belonged to the G1b (S-INDEL) subgroup and were closely related to the European PEDV strains isolated from 2014 to 2019. Furthermore, RDP4 analysis revealed that the Polish PEDV strains harboured a recombinant fragment of ~400 nt in the 5’ end of S gene with PEDV and swine enteric coronavirus (SeCoV) being the major and minor parents, respectively. Antigenic analysis showed that the aa sequences of neutralizing epitopes were conserved among the Polish PEDV strains. Only one strain, #0100/5P, had a unique substitution in the COE epitope. However, Polish PEDV strains showed several substitutions, especially in the COE antigen, as compared to the classical strain CV777. To the best of our knowledge, this is the first report concerning the molecular characterization of porcine epidemic diarrhoea virus strains, as well as the first phylogenetic analysis for PEDV in Poland.

Phylogenetic Analysis of European Brown Hare Syndrome Virus Strains from Poland (1992-2004).

European brown hare syndrome (EBHS) is lethal to several species of free-living hares worldwide. The genetic characterization of its virus (EBHSV) strains in European circulation and epidemiological knowledge of EBHSV infections is not yet complete. The study determined the nucleotide sequences of the genomes of EBHSV strains from Poland and analyzed their genetic and phylogenetic relationships to a group of hare lagoviruses. The genome of five virus strains detected in Poland between 1992 and 2004 was obtained by RT-PCR and sequencing of the obtained amplicons. The genetic relationships of the EBHSV strains were analyzed using the full genome and VP60 gene sequences. Additionally, the amino acid sequence of the VP60 gene was analyzed to identify mutations specific to recognized EBHSV subgroups. Partial amplification of the virus open reading frame (ORF)1 and ORF2 regions obtained nearly complete nucleotide genome sequences of the EBHSV strains. Phylogenetic analysis placed them in a GII.1 cluster with other European strains related to nonpathogenic hare caliciviruses. VP60 gene analysis allocated these EBHSV strains to the G1.2, G2.2–2.3 or G3 virus genetic groups. The amino acid sequence differences in the entire genome ranged from 1.1 to 2.6%. Compared to a reference French EBHSV-GD strain, 22 variable amino acid sites were identified in the VP60 region of the Polish strains, but only six were in VP10. Single amino acid changes appeared in different sequence positions among Polish and other European virus strains from different genetic groups, as well as in VP10 sequences of nonpathogenic hare caliciviruses. The results of the study showed a high genetic homogeneity of EBHSV strains from Poland despite their different location occurrence and initial detection times. These strains are also phylogenetically closely related to other EBHSV strains circulating in Europe, likely confirming the slow evolutionary dynamics of this lagovirus species.

The first detection and characterization of goose parvovirus (GPV) in Turkey.

Derzsy's disease, which is seen in goslings (Anser anser domestica) and Muscovy ducks (Cairina moschata), progresses to high mortality and causes significant yield losses. The disease agent is goose parvovirus (GPV), which is common in countries with waterfowl production. It has not previously been reported in Turkey. Using qPCR and sequencing of the VP3 protein-encoding gene, GPV is identified as the causative agent of high mortality among geese between 2018 and 2019. The VP3 sequences were also compared with the similar GenBank sequences phylogenetically. All the sequences were found to be most similar (98.90%) with Polish and Taiwan GPV strains. Phylogenetic analysis of the VP3 gene in strains in Turkey and comparison with strains from other countries demonstrated that the Turkish strains are native to the geography and circulated locally. This study detected the presence of the GPV gene for the first time in Turkey and demonstrated the importance of comparing the vaccine strain and wild type.

The Spillover of African Swine Fever in Western Poland Revealed Its Estimated Origin on the Basis of O174L, K145R, MGF 505-5R and IGR I73R/I329L Genomic Sequences.

The African swine fever epidemic occurred in Poland at the beginning of 2014 and, up to date, the disease has been spreading mainly in the eastern part of the country. Unexpectedly, in November 2019 an infected wild boar case was confirmed in Lubuskie voivodship in western Poland. During the following weeks, several dozen African swine fever virus (ASFV)-positive animals were notified in the neighboring area, causing severe concern regarding further spread of the disease to the mostly pig-dense region in Poland, namely, Wielkopolskie voivodship. Moreover, almost a year after, several infected wild boar cases were confirmed for the first time in Germany, just beyond the Polish border, sending out a shock wave through the global pig market. The whole genome sequence of ASFV, isolated from the first case of ASF in western Poland, and three selected viruses from other affected areas, revealed the tandem repeat and single nucleotide polymorphism (SNP) variations in reference to the Georgia 2007/1 strain. These data, supported by the conventional sequencing of selected genomic regions from a total of 154 virus samples isolated between 2017 and 2020 in Poland, shed a new light on pathogen epidemiology. The sequence variations within the O174L gene detected in this study showed that cases identified in western Poland might be originating from the so-called southern Warsaw cluster. Moreover, the viruses originating from the northern Warsaw cluster do not possess single nucleotide polymorphism (SNP) mutations within the K145R and MGF 505-5R genes, which are specific to all of the other Polish ASFV strains. These results led to a conclusion of their distinct origin. Supporting these results, the nucleotide sequencing of I73R/I329L intergenic region revealed its new, previously undescribed variant, called IGR IV, with an additional three tandem repeats of 10 nucleotides in comparison to the reference sequence of the Georgia 2007/1 strain.

Genomic background of the Klebsiella pneumoniae NDM-1 outbreak in Poland, 2012-18.

To characterize genomes of Klebsiella pneumoniae ST11 NDM-1 responsible for a countrywide outbreak in Poland and compare them phylogenetically with other Polish and international ST11 strains. Seventy-one carbapenemase-producing K. pneumoniae ST11 isolates from Poland, including 66 representatives of the NDM-1 epidemic from 2012-18, were sequenced using Illumina MiSeq. Additionally, three outbreak isolates were also sequenced using MinION. The clonality and phylogenetic analysis was done by core-genome MLST and SNP approaches. Resistomes, virulomes, K/O antigens and plasmid replicons were screened for. The detailed plasmid analysis was based on full assemblies using Oxford Nanopore Technologies data. Chromosomes of the outbreak isolates formed an essentially homogeneous cluster (though accumulating SNPs gradually with time), differing remarkably from other Polish NDM-1/-5-, KPC-2- or OXA-48-producing K. pneumoniae ST11 strains. The cluster belonged to a clade with 72 additional isolates identified worldwide, including closely related NDM-1 producers from several countries, including organisms from Bulgaria and Greece. All these had KL24 and O2v1 antigens and the chromosomal yersiniabactin locus YbST230 residing in the ICEKp11 element. The specific blaNDM-1-carrying Tn125 transposon derivative, named Tn125A, was located in IncFII/pKPX-1- and/or IncR-like plasmids; however, the IncRs rearranged extensively during the outbreak, contributing to highly dynamic plasmid profiles and resistomes. The K. pneumoniae ST11 NDM-1 genotype that has been expanding in Poland since 2012 is largely monoclonal and represents a novel international high-risk lineage that is also spreading in other countries.

Different Gene Expression Response of Polish and Australian Raphidiopsis raciborskii Strains to the Chill/Light Stress

R. raciborskii is known for growing under wide ranges of temperature and light. In temperate regions, however, low temperature and high light may serve as a stressful condition for invading tropical populations. The genetic basis of R. raciborskii’s adaptation to this combination of stresses are unknown. In this study, the growth rate and the expression of genes that may be crucial in the response to the chill/light stress of two R. raciborskii strains (differing in their climatic origin and toxicity) exposed to low temperature and high light were examined. Results showed that AMU-DH-30, a non CYN (cylindrospermopsin) producing strain isolated from the temperate region, exhibited under stress the upregulation of genes involved in the protein translation (rbp1, nusG, hflX), membrane fluidity (desA), photosynthetic activity (ccr2 and ftsH), and the accumulation of compatible solutes (asd). In contrast, a CYN producing Australian strain CS-505 was not able to adapt quickly and to continue growth during stress conditions. Intriguingly, CS-505 and AMU-DH-30 had a similar ability to resume growth when the stress conditions subsided. Moreover, in strain CS-505 the cyrB gene was significantly upregulated under the stress conditions. The presented results shed new light on the possible mechanisms involved in the response of R.raciborskii to chill/light stress.

Emergence and expansion of novel pathogenic reassortant strains of infectious bursal disease virus causing acute outbreaks of the disease in Europe.

Infectious bursal disease virus (IBDV) is the aetiological agent of a highly contagious chicken immunodeficiency disorder known as Gumboro disease, which cause severe economic loses to the poultry worldwide. The emergence of very virulent IBDV strains (vvIBDV) during the late 80s resulted in drastic changes to the epidemiology of IBDV with a dramatic increase in the mortality of the animals affected. Molecular studies determined that the emergence of the vvIBDV was a consequence of a genomic reorganization of IBDV known as reassortant event by which the virus combined two emergent genetic background vvIBDV for segment A and vvIBDV for segment B. In the current study, a retrospective analysis was conducted, and samples collected during acute outbreaks of Gumboro disease in Poland during 1992-2015 were submitted to sequencing and further molecular and phylogenetic analyses. The results obtained not only revealed a high genetic diversity for Polish IBDV strains but a new population of IBDV was identified. These novel reassortant strains with a unique genetic background contain the segment A from very virulent strains and segment B from an unidentified source, phylogenetically segregated and classified as 'transitionallineage'. The results obtained also showed the presence of this new lineage in Finland, evidencing the expansion of this new genomic reorganized viral strain in Europe representing an additional threat to the global situation of IBDV.

Molecular epidemiology of infectious bronchitis virus in Poland from 1980 to 2017

The presence of infectious bronchitis virus (IBV) was identified for the first time in the poultry population in Poland at the end of the 1960s. From this time a few waves of epidemics caused by different IBV variants spread across the country. In order to gain more insight into the molecular epidemiology of IBV in Poland, in the present study the S1 coding region of 34 IBV isolates and nearly whole genome of 10 strains collected over a period of 38 years was characterized. Phylogenetic analysis showed that these strains belonged to five recently established IBV lineages: GI-1, GI-12, GI-13, GI-19 and GI-23. Additionally, two strains from 1989 and 1997 formed a separate branch of the phylogenetic tree categorized as unique early Polish variants, and one strain was revealed to be the recombinant of these and GI-1 lineage viruses. Irrespective of year of isolation and S1-dependent genotype, the genome sequences of Polish IBV strains showed the presence of six genes and 13 ORFs: 5′UTR-1a-1b-S-3a-3b-E-M-4b-4c-5a-5b-N-6b-3′UTR, however their individual genes and putative proteins had different lengths. The phylogenetic analyses performed on the genome of ten Polish IBV strains revealed that they cluster into different groups. The Polish GI-1, GI-19 and GI-23 strains cluster with other similar viruses of these lineages, with the exception of the two strains from 1989 and 1997 which are different. It seems that in Poland in the 1980s and 1990s IBV strains with a unique genome backbone circulated in the field, which were then replaced by other strains belonging to other IBV lineages with a genome backbone specific to these lineages. The recombination analysis showed that some Polish strains resulted from a recombination event involving different IBV lineages, most frequently GI-13 and GI-19.

Pectobacterium polonicum sp. nov. isolated from vegetable fields.

Gram-stain-negative, rod-shaped pectinolytic bacteria strains designated as DPMP315T, DPMP316, DPMP317 and DPMP318 isolated from groundwater sampled from a vegetable field in the North of Poland, were subjected to the polyphasic analyses. Multilocus sequence analyses based on five housekeeping genes (gyrA, recA, recN, rpoA and rpoS) revealed their distinctiveness from the other species of the genus, simultaneously indicating that the newly described species, Pectobacterium punjabense, as well as Pectobacterium parmentieri and P. wasabiae, to be the closest relatives. In silico DNA-DNA hybridization (<43.1 %) and average nucleotide identity (<92.5 %) values of strain DPMP315T with other type strains of species of the genus Pectobacterium supported the delineation of the novel strain as representing a novel species. The phenotypic comparisons, fatty acid methyl esters compositions, genetic rep PCR fingerprint and detailed whole-cell MALDI-TOF mass spectrometry proteomic profiles permitted the differentiation of Polish strains from the type strains of all other known species of the genus Pectobacterium. The results of polyphasic analyses performed for four Polish strains are the basis for the distinction of the novel species. Here, we propose to establish DPMP315T as a type strain (=PCM3006T=LMG 31077T) with the name Pectobacterium polonicum sp. nov.

New Insight into Genotypic and Phenotypic Relatedness of Staphylococcus aureus Strains from Human Infections or Animal Reservoirs.

Staphylococcus aureus is a common human and livestock opportunistic pathogen, and there is evidence of animal to human transmission. This paper aimed to recognize properties of the isolates from collections of human and livestock S. aureus strains and to estimate compatibility of results based on phenotypic tests, microarrays and the spa typing methods. The second goal was to study differences between human and animal isolates in terms of specificity of their hosts and the strain transmission among various hosts. Most strains showed multi-susceptible profiles and produced enzymes on a high level, and they were phenotypically and genetically similar. However, in contrast to the Polish bovine mastitis strains, the Slovakian strains were multi-resistant. In this research, the strains showed significant differences in terms of their phenotypic manifestations and the presence of hemolysins genes; however, other enzyme-encoding genes correlated to a higher extent with the microarrays results. Interestingly, there was a lack of enterotoxin genes in human Poultry-like protein A+ strains in comparison to other human strains. Our study showed that differences between virulence profiles of the human and animal strains correlated with their origin rather than their hosts, and any trait allowed clearly distinguishing between them based on the microarray results.

Is the presence of Central European strains of Raphidiopsis (Cylindrospermopsis) raciborskii a threat to a freshwater fish? An in vitro toxicological study in common carp cells

As yet European strains of Raphidiopsis raciborskii (previously Cylindrospermopsis raciborskii) have not been found to produce known cyanotoxins although their extracts have caused adverse effects in mammals, as shown using in vitro and in vivo experimental models. The present study investigated whether R. raciborskii isolated from Western Poland and Ukraine can affect fish cells using in vitro exposures of hepatocytes and red blood cells (RBC), and brain homogenates obtained from common carp (Cyprinus carpio) to 1.0% and 0.1% extracts of 7 strains. The studied extracts evoked different responses of catalase activity in hepatocytes with both increase and decrease observed under low and high concentrations. The cellular thiol pool was also altered with most extracts inducing a decrease in the activity of glutathione-S-transferase, and Ukrainian strains leading to an increase in glutathione level and a decrease in metallothionein content. All the studied extracts induced comparable reactive oxygen species formation, lipid peroxidation, protein carbonylation and DNA fragmentation in hepatocytes, and all but one increased the activity of caspase-3. Only one extract caused lysosomal membrane destabilization as measured by neutral red retention in RBC. In contrast to extracts of Ukrainian isolates, exposure of brain homogenates to extracts of Polish strains induced an increase in acetylcholinesterase activity suggesting the neurotoxic action of their exudates. The results indicate that both Polish and Ukrainian strains of R. raciborskii may pose a toxicological risk to freshwater fish, and further, that Polish strains may produce compound(s) evoking neurotoxic effects.

Comparison of Schmallenberg virus sequences isolated from mammal host and arthropod vector

Schmallenberg virus (SBV) is the member of Peribunyaviridae family, which comprises pathogens of importance for human and veterinary medicine. The virus is transmitted only between animals and mainly by biting midges of the genus Culicoides. This study was performed in order to determine SBV genetic diversity and elucidate the host–vector adaptation. All three viral segments were analysed for sequence variability and phylogenetic relations. The Polish SBV strains obtained from acute infections of cattle, congenital cases in sheep, and from Culicoides midges were sequenced using Sanger and next-generation sequencing (NGS) methods. The obtained sequences were genetically similar (99.2–100% identity) to the first-detected strain BH80/11—4 from German cattle. The sampling year and origin of Polish sequences had no effect on molecular diversity of SBV. Considering all analysed Polish as well as European sequences, ovine-derived sequences were the most variable, while the midge ones were more conserved and encompassed unique substitutions located mainly in nonstructural protein S. SBV sequences isolated from Culicoides are the first submitted to GenBank and reported.

Molecular analysis of the chromosomal 16S rRNA gene and vapA plasmid gene of Polish field strains of R. equi.

Rhodococcus equi (R. hoagii) is an opportunistic pathogen commonly found in foals up to 6 months old and animal environment. The R. equi genome contains genetically stable chromosomal DNA and an 80–90 kb plasmid containing vapA gene, responsible for virulence. Most reports from around the world focus on the determination of R. equi plasmid profiles. Few studies have attempted to determine differences in nucleotide sequences between virulent strains of R. equi isolated from foals and breeding environment. The aim of the study was to perform a molecular analysis of a fragment of the chromosomal gene encoding the 16S rRNA subunit and the vapA plasmid gene of virulent R. equi strains isolated on Polish studs from foals and from the breeding environment of horses. The sequencing method was used to compare the primary structure of fragments of the chromosomal and plasmid DNA of the virulent R. equi strains. The sequences of 22 clinical and 18 environmental R. equi isolates were compared with the sequences of the gene fragments of reference strains available in the NCBI GenBank database. All sequenced 16S rRNA amplicons of Polish field strains were identical and showed 99.5% similarity to the four randomly selected sequences of this gene fragment in the GenBank database. The results confirm that fragments of the 16S rRNA gene of R. equi strains are highly conserved and do not undergo variation in field conditions. Analysis of the sequencing results for the vapA gene fragment of the strains used in our study revealed two polymorphic variants and clear differences between the sequences of strains isolated from foals and from soil samples. Presumably, R. equi strains present in the breeding environment are more exposed than clinical strains to adverse external factors. This may result in changes in the DNA sequence due to natural selection.