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The study has been taken up to collect more data on Yersinia enterocolitica isolated from pigs as the main reservoir and source of infection with strains pathogenic for humans. Bacteriological examinations, bio- and serotyping were conducted on 616 rectal swabs, taken from 308 fattening pigs. Two samples were taken from each animal to determine the ability of Y. enterocolitica to grow under different temperature conditions (warm and cold culture). It has been proven that low temperature constitutes a suitable culture condition. 138 Yersinia enterocolitica strains were isolated (22.40%), most of which (65.22%) were obtained in cold culture and 99.28% included in biotype 3 (one strain belonged to biotype 2). Serotyping yielded a positive result in 107 strains with the diagnostic serum for antigen O:3, in 18 - with the serum for antigen O:9, and 13 strains were determined to be non-typable. The results indicated that asymptomatic infections with Y. enterocolitica strains of the biotypes and serotypes pathogenic for humans are common in pig population.
The purpose of the study was to evaluate the occurrence of genes directly connected with pathogenicity in Yersinia enterocolitica strains isolated from fattening pigs. Multiplex PCR, used for ystA, ystB, ystC, and ymoA gene detection, was optimised in order to determine the existence of the genes in one reaction. Material for the study consisted of 138 strains of Y. enterocolitica, which were preliminary examined by the bacteriological, sero- and biotyping methods, then bacterial DNA was isolated and multiplex PCR was performed. The presence of the products of length corresponding to the ystA and ymoA gene fragments was found in each of the examined strains. The ystB and ystC genes were not detected in any of the tested samples. The molecularly confirmed existence of Y. enterocolitica in fattening pigs indicates the carrier state and a possibility of shedding the microorganism into the environment, and shows that pigs are an important reservoir of the bacteria and a potential source of infection for humans.
The objective of this study was to compare the effectiveness of the multiplex PCR method and traditional light microscopy in identifying and discriminating the species of Nosema spp. spores in worker bees from winter hive debris in the Province of Warmia and Mazury (NE Poland). A total of 1000 beesdead after from the bottom of the hive from bee colonies were analyzed. Spores were identified with the use of a light microscope (400-600x magnification). Spores were assigned to species by the multiplex PCR method. The microscopic evaluation revealed the presence of Nosema spp. spores in 803 samples (80.3%). Nosema ceranae spores were observed in 353 positive samples (43.96%), Nosema apis spores were found in 300 samples (37.35%), while 150 samples (19.67%) showed signs of a mixed infection. A multiplex PCR analysis revealed that 806 samples were infested with Nosema spp., of which 206 were affected only by Nosema ceranae, 600 showed signs of mixed invasion, while no samples were infected solely by Nosema apis parasites. In two cases, the presence of spores detected under a light microscope was not confirmed by the PCR analysis. The results of the study indicate that Nosema ceranae is the predominant parasitic species found in post-winter worker bees from the bottom of the hive in the region of Warmia and Mazury.
Yersinia (Y.) enterocolitica is the third etiological agent of human diarrhea in terms of the number of confirmed clinical cases. One of the important virulence markers is the yst gene which encodes the production of enterotoxins Yst (Yersinia stable toxins). However, not all strains with yst genes produce enterotoxins, what seems to be caused by the ymoA gene encoding the production of the YmoA protein inhibiting the expression of various genes. The purpose of our study was to evaluate the distribution of the ymoA and ystA genes and Yst production by Y. enterocolitica isolated from humans and pigs. All the studied strains obtained from pigs had the ystA gene which indicates that they belong to the group of strains commonly regarded as pathogenic, but the ability to produce YstA was detected in only 14 out of 96 examined strains. The fragments of ystA gene were also detected in all Y.enterocolitica strains isolated from human cases of diarrhea. Amplification of a fragment of the ymoA gene was detected in all the studied strains, both from humans and pigs, based on the presence of a 330 bp band. Thus no correlation was identified between the occurrence of the ymoA and ystA genes and the production of a specific type of enterotoxin.
Enterotoxic capacities of 95 Yersinia enterocolitica strains isolated from fattening pigs by the use of the suckling mouse bioassay were investigated. A positive result was received for 14 strains (14.74%), doubtful for 33 strains (34.74%), while the rest of 48 strains (50.52%) did not induce the production of enterotoxins. The received results show that in the pig population there are almost 15% of Yersinia enterocolitica strains, which are directly dangerous to public health because of their enterotoxic capacity. It is also worth paying attention to those strains, which in the suckling mouse bioassay provided doubtful results (around 35%) because their ability to produce enterotoxins was not explicitly declared as impossible.
In order to perform a phylogenetic analysis of Nosema spp. spores, samples of 10 worker bees each were collected from 30 infected colonies (the presence of spores was confirmed by light microscopy) kept in 15 apiaries in north-eastern Poland. Both N. apis and N. ceranae are common in this region (mixed infection N. apis/ N. ceranae – 60%, N. ceranae – 37%, N. apis – 3%). The DNA samples of N. apis were 100% identical with the N. apis sequences deposited in the GenBank database in Queensland, Australia, Spain, New Zealand, Lithuania and Tasmania in Australia. The DNA samples of N. ceranae were found to be 99.5%-100% identical with the N. ceranae sequences previously published in Italy, Germany, Switzerland and Austria.
The aim of the study was to identify the bovine papillomavirus (BPV) types associated with equine sarcoids observed in Polish horses, during 2001-2004. The samples of skin tumours were formalin-fixed and paraffin-embedded. Out of 14 tumours obtained from 10 horses, 11 were diagnosed as fibroblastic sarcoid, two as verrucose sarcoid, and one as epidermal cystoid. Using PCR, the presence of E5 open reading frames (ORFs) fragment BPV-1/BPV-2 were shown in all specimens obtained from archival tissues. Sequence variants of E5 ORFs confirm that European variants of BPV-1 occur in fibroblastic and verrucose equine sarcoids in Poland.
Free-living animals are an important environmental reservoir of pathogens dangerous for other animal species and humans. One of those is Yersinia (Y.) enterocolitica, the causative agent of yersiniosis - foodborne, enzootic disease, significant for public health. The purpose of the study was to identify bioserotypes and virulence markers of Y. enterocolitica strains isolated from roe deer (Capreolus capreolus) and red deer (Cervus elaphus) obtained during the 2010/2011 hunting season in north-eastern Poland. From among 48 rectal swabs obtained from 24 roe deer, two strains of Y. enterocolitica from one animal were isolated. Although both belonged to biotype 1A they were identified as different serotypes. The strain obtained from cold culture (PSB) belonged to serotype O:5, while the strain isolated from warm culture (ITC) was regarded as nonidentified (NI), what may suggest mixed infection in that animal. The presence of ystB gene, coding for YstB enterotoxin, directly related to Y. enterocolitica pathogenicity was detected in both strains using triplex PCR. The effect of the examination of 32 swabs obtained from 16 red deer was the isolation of two Y. enterocolitica strains from two different animals. Both belonged to biotype 1A with NI serotype, but were originated from different types of culture. They gave positive results in case of products of a size corresponding to the ystB gene. No amplicons corresponding to ail and ystA genes were found. Roe deer and red deer may carry and shed Y. enterocolitica, what seems to be important in aspect of an environmental reservoir of this pathogen. The Y. enterocolitica strains isolated from wild ruminants had the amplicons of the ystB gene, what suggest they can be potential source of Y. enterocolitica infection for humans.
The aim of the study was to analyse a part of the sequence of the E5 gene of bovine papil-lomaviruses (BPV) associated with equine sarcoids in Polish horses. Samples of 40 skin lesions obtained from 29 horses were collected for molecular examination. The PCR amplicons of BPV DNA were detected in 38 specimens. After phylogenetic analysis 37 specimens were recognized as BPV-1 and one as BPV-2. Phylogenetic analysis has allowed the classification of the amplicons into two phylogenetic groups (A1,) and four separate isolates (2, 10, 16, 17).
The purpose of the studies was to determine, by means of bacteriological and molecular methods, the occurrence of Y. enterocolitica in the faeces of dogs, as well as to assess the degree of their spread in the urban environment. The faeces were collected from the lawns of six large districts of Olsztyn. In order to isolate Y. enterocolitica, "warm culture" (ITC/CIN) and "cold culture" (PSB/CIN) were used in parallel, together with biochemical tests. Next, genomic DNA of Y. enterocolitica was isolated and αil, ystA, and ystB genes were detected with the use of the multiplex PCR method. A relatively frequent occurrence of Y. enterocolitica strains in canine faeces contaminating the urban lawns of Olsztyn was demonstrated. The greatest percentage of contaminated faecal samples was found in the area of large housing estates. By means of molecular tests, the presence of ystB gene only, in the absence of αil and ystA genes, was demonstrated in the Y enterocolitica genome, which could indicate a potentially pathogenic biotype 1A. Therefore, it seems important to keep monitoring the changes, which occur within this species of microorganisms, the epidemiological situation of yersiniosis in human and animal populations, as well as to continue the studies on the epidemiology of Y enterocolitica infections also in the context of a reservoir of animals accompanying a man.
The study aimed at the evaluation of the occurrence of genes directly connected with pathogenicity in Yersinia enterocolitica (Y. enterocolitica) strains isolated from wild boars, obtained in the 2007/2008 hunting season in the North-East territory of Poland. Multiplex PCR, used for αil, ystA, and ystB gene detection, was optimised in order to determine the existence of the genes in one reaction. Forty-six palatine tonsils, gained from 46 wild boars of various ages, were preliminary examined from the bacteriological, sero- and biotyping points of view; then bacterial DNA was isolated and multiplex PCR was performed. The presence of the product of volume corresponding to the ystB gene fragment was found in two cases, which constituted 4.35% of the examined samples. The ail and ystA genes were not detected in any of the tested samples. The molecularly confirmed existence of Y. enterocolitica in wild boar palatine tonsils, indicates the carrier state and possibly shedding of the microorganism into the environment, and suggests that wild boars, likewise pigs to a lesser extent, may constitute a reservoir of the bacteria and a potential source of infection for the man.
Equine sarcoids have been recognized for centuries, but were first characterized by Jackson in 1936. The term sarcoid was originally used to emphasize the clinical and pathological differences from papilloma, fibroma and fibrosarcoma. Sarcoids are the most common skin tumors in horses. They tend to be locally aggressive but they do not metastasize. They affect horses of all ages, types and hair colours without obvious sex predilection and they are commonly encountered worldwide. Sarcoids are recognized as having six different clinical types: verrucose, fibroblastic, occult, nodular, mixed and malignant. All of them can occur at any skin site. The mechanisms of equine sarcoid development are not clear, but papillomaviruses (PV) - BPV-1 and less commonly BPV-2, which belong to the Papillomaviridae family - participate in etiopathogenesis. All PV are strictly species-specific and, even in experimental conditions, do not infect other species. The only known case of cross-species infection is the infection of horses and other Equidae by BPV-1 and BPV-2.
Y. enterocolitica infection of pigs is concerned with a common carrier and shedding states in this species. The aim of the study was to determine the influence of the experimental immunization of pigs with a selected Y. enterocolitica strains suspension on the controled infection course, the antibody level formation, as well as the duration of pathogen shedding. The immunization of pigs was enforced with a suspension of Y. enterocolitica strains previously inactivated with formol, suspended in PBS and demonstrated in vitro high immunogenicity in Respiratory Burst Activity/Potential Killing Activity (RBA/PKA) and Mitogen Transformation Test (MTT). The study was performed on 15 pigs divided into 3 groups. The animals from groups I and II received an immunizate with a density of 2.7 × 10⁹ cfu/ml administered subcutaneously in doses 2 ml and 5 ml, respectively, twice in a 2-week interval. The third group (control) was administered PBS in an analogous scheme. The evaluation in vivo was conducted after a per os challenge with a pathogenic strain of Y. enterocolitica O:3. The first and booster immunization had no effect on the clinical picture and body weight gains of the immunized animals. The fastest and strongest immune response to the per os challenge with Y. enterocolitica O:3 was observed in the control group, already in the first week post infection (wpi). Higher antibody levels were found in the group of animals where 5 ml 2.7 × 10⁹ cfu/cm³ subcutaneously were administered. In the first wpi bacterial shedding was observed in all animals that belonged to group I and the control group which persisted up to 3 wpi. Among the pigs from group II the shedding was observed in 3 out of 5 animals, and which finished in 6 days post infection. Applied experimental immunization against per os challenge with a pathogenic strain of Y. enterocolitica O:3 did not prevent pathogen shedding, but merely limited its intensity and duration.
Pseudocloacal swabs and palatine tonsils from beavers have been examined for the Yersinia enterocolitica presence. Thirty-six samples from 24 beavers were collected and subjected to bacteriological examinations including sero- and biotypisation. Amplicons confirmed by PCR as Y. enterocolitica were sequenced. Positive samples originated from 4 out of the 24 beavers (16.7 %) and all the strains belonged to biotype 1A. The study suggested that Y. enterocolitica could be isolated from beavers, which may therefore be treated as a reservoir, a significant factor of water contamination and a vector of the Y. enterocolitica.
Papillomaviruses (PV) are small, nonenveloped, DNA viruses, which had originally been grouped together with the Polyomaviruses in one family, Papovaviridae. In the year 2004 the International Committee on the Taxonomy of Viruses officially recognized two separate families: Papillomaviridae and Polyomaviridae. PV are pathogens of skin and mucosa in animals and humans, and they are very species-specific. The only known case of cross-species infection is the infection of horses by bovine papillomaviruses (BPV) type 1 and 2. Infection by high-risk types of human papillomaviruses (HPV) such as HPV type 16 and 18 is directly related to the subsequent development of cervical carcinoma in women. In the year 1995 The International Agency for Research on Cancer officially declared, that HPV-16 and HPV-18 are carcinogenic for humans. Animal PV cause various diseases in both farm and companion animals, e.g. skin and teat papillomatosis in cattle, canine oral papillomatosis, oesophageal papillomas and carcinoma in cattle and equine sarcoids. The mechanisms of carcinogenesis caused by PV were initially established using animal models and specifically chosen PV, particularly cottontail rabbit papillomavirus (CRPV), BPV and canine oral papillomavirus (COPV). In the paper the organization and structure of the PV genome, the characteristics of early and late regions, enzymatic and regulatory proteins, encoded by specific open reading frames and engaged in virus replication process, as well as structural proteins that take part in virus-cell interaction have been discussed. The replication process of PV and mechanisms of carcinogenic transformation of cells infected with PV were also described. The possibility of the implementation of specific immunoprophylaxis and the necessity of improvement of diagnostic methods, as well as conducting molecular comparative studies of human and animal PV, important for the protection of animal health and public health, have been indicated.
The aim of the study was to evaluate a novel diagnostic scheme which combines quantitative PCR and High-Resolution Melting (qPCR-HRM) curve analysis for rapid differentiation based on E5 partial CDS of bovine papillomavirus type 1 or 2 (BPV-1 or BPV-2), and to perform a phylogenetic analysis of the complete CDS of the E5 gene of BPV detected in equine sarcoids. Samples of 38 skin lesions obtained from 27 horses were collected for molecular examinations. All lesions were clinically diagnosed as sarcoids, but results of histopathological examinations did not always corroborate the clinical diagnosis. Although all the samples were positive for the presence of BPV DNA, after qPCR-HRM analysis 6 (16%) specimens were recognized as BPV-1 “wild”, 24 (63%) as BPV-1 “European” and 8 (21%) as a “variant” of BPV E5 ORF partial CDS. Phylogenetic analysis based on nucleotide sequences of E2 ORF partial CDS and E5 ORF complete CDS was conducted on 7 specimens, whose sequences were published in GenBank and recognized as: 2PL (Accession Number - Acc. No. KC684939) - “variant” BPV-1, 7aPL (Acc. No. KC684940) - “European” BPV-1, 10PL (Acc. No. KC693480) - “variant” BPV-1, 16PL (Acc. No. KC693484) - “variant” BPV-2, 17PL (Acc. No. KC693481) - “variant” BPV-1, 20aPL (Acc. No. KC693482) - “European” BPV-1 and 20cPL (Acc. No. KC693483) - “wild” BPV-1. Amino acid (aa) sequences of E5 ORF complete CDS were also analyzed. The E5 variant of aa sequences found in isolate 10PL (protein identification - ID: AGM 20700) is a novel variant of E5 ORF complete CDS of BPV-1 detected in equine sarcoid in Poland.
The aim of the study was to evaluate the prevalence of seroreagents to Aleutian mink disease virus (AMDV) in minks. Sera samples from 1233 minks were tested by CIEP. The antibodies against AMDV were not found in the mink farms which continued the AMD eradication program. In farms where the eradication program had not been implemented a bad epizootic situation was detected. The percentage of seroreagents between 33.3% and 97.8% was noticed: in females 33.3% to 91.8% whereas in males it ranged from 61.5% to 97.8%. The PCR method was used in 101 spleen samples to detect the presence of 11 new nucleotide sequences of VP2 gene, encoding the capsid protein of AMDV. All 11 amplicons are new variants of the VP2 AMDV gene isolated from minks in Poland.
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