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Papers by Ivo Pavlík

Czech Journal of Food Sciences, Dec 31, 2011

We assessed the survival of Mycobacterium avium subsp. hominissuis (MAH) in artificially contamin... more We assessed the survival of Mycobacterium avium subsp. hominissuis (MAH) in artificially contaminated homemade smoked sausages prepared from pork meat according to traditional recipes, the effect of storage of such sausages at -20°C for three months on MAH viability and to compare assessment of MAH viability/presence by culture and qPCR. Three isolates of MAH were inoculated into the sausage mixture at concentrations of 10 6 CFU per gram of meat and cold smoked at 40°C for 12 h or hot smoked at 70°C for 6 hours. MAH survived the cold smoking procedure without any significant decrease in viable MAH CFU counts; no viable MAH were detected in the hot smoked sausages. The storage of sausages at -20°C caused a decrease in viable MAH counts of about 1 to 3 log 10 . Absolute MAH counts determined by qPCR were not significantly reduced by the storage or smoking. The presence of viable MAH in sausages after the cold smoking should be considered as a risk for immunodeficient individuals and children.

Food Technology and Biotechnology, Jun 9, 2009

The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, originally f... more The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, originally found in seeds of amaranth (Amaranthus caudatus), into the genome of potatoes (Solanum tuberosum). The purpose of the present study is to develop a PCR method for the detection of the mentioned genetically modified potatoes in various foodstuffs. The method was used to test twenty different potato-based products; none of them was positive for the genetic modification 'ac2'. The European Union legislation requires labelling of products made of or containing more than 0.9 % of genetically modified organisms. The genetic modification 'ac2' is not allowed on the European Union market. For that reason it is suitable to have detection methods, not only for the approved genetic modifications, but also for the 'unknown' ones, which could still occur in foodstuffs.

The "atypical mycobacteria" are widely distributed in the natural environment and can produce inf... more The "atypical mycobacteria" are widely distributed in the natural environment and can produce infection in farm and wild animals bred in captivity. The objective of the study was to analyze the variety of mycobacterial species in the environment of breeding facilities, tanks and aquariums, as well as in samples of peat used as a feed supplement in the Czech Republic over the 2003-2004 period. A total of 1389 samples from environment were examined, collected from 29 sites throughout the Czech Republic. The samples were decontaminated and cultured at 25 °C and 37 °C in three culture media: Stonebrink's medium, Herrold's egg yolk medium and Sula's medium. Mycobacterial isolates grown were identified by the PCR method and growth and biochemical tests. Of the 1389 environmental samples, mycobacteria were demonstrated by culture in 400 (28.8%) of them originating from different substances. In the samples of farm environment and in peat, mycobacteria were isolated in 185 of 1064 (17.4%) samples, and in 201 of 325 (61.8%) samples from aquarium and breeding tank environment. The results show significant difference between the findings from the stable environment (17.4%) and those from the aquarium environment (61.8%). A wide range of species was found in all constituents of the environment. The following species were most frequently isolated from the stable environment: M. avium (6.5%), M. fortuitum (3.2%), M. flavescens (3.2%), and the aquarium environment harboured most frequently M. fortuitum (13.9%), M. marinum (8.0%) and M. gordonae (4.5%). The results confirm the frequent occurrence of various mycobacterial species in the environment, predominantly in aquariums, some of which can produce infection in animals and in man following contact with the infected environment.

Veterinární medicína

In 2001, genetically modified potatoes carrying the ac2 gene were developed. This gene was origin... more In 2001, genetically modified potatoes carrying the ac2 gene were developed. This gene was originally found in amaranth seeds (Amaranthus caudatus) and is expressed into the fungicidal peptide Ac-AMP2. The purpose of the present study was to test 105 potato cultivars (Solanum tuberosum) registered in the Czech Republic for the presence of the above mentioned genetic modification "ac2" with the use of a previously published method . The method was based on the simultaneous detection of the ac2 gene from amaranth seeds as well as the StTS1 gene from potatoes as an internal amplification control. The results showed none of tested potatoes cultivars were positive for the genetic modification "ac2". These results confirmed the currently valid legislative in the Czech Republic and the European Union, where the use of genetically modified potatoes carrying the gene for the fungicidal peptide from amaranth is not allowed, was respected.

Veterinární medicína, 2012

Genetically modified organisms (GMO) become a real constituent of our lives and nowadays, they ar... more Genetically modified organisms (GMO) become a real constituent of our lives and nowadays, they are commonly introduced into the food chain of people and animals in some states. Among higher organisms, plants are used above all for genetic modifications; potatoes are a suitable model plants for this purpose. Nowadays, a number of various genetic modifications of potato plants are available, particularly those with increased resistance to biological agents and factors of the external environment or with improved nutritional value. Plants that produce proteins of the immune system of man or animals or substances that may be used as vaccines in human or veterinary medicine are highly important. Modified potato plants that produce biomaterials for potential applications in the industry are a significant category.

Veterinární medicína, 2008

The hepatitis E virus (HEV), the causative agent of hepatitis E, is a non-enveloped RNA virus. Th... more The hepatitis E virus (HEV), the causative agent of hepatitis E, is a non-enveloped RNA virus. The HEV genome is formed by a non-segmented positive-sense RNA chain. The 3´end of the chain is polyadenylated and the 5´end is structurally characterised by the so called “capping”. According to currently accepted taxonomy, HEV is classified in the genus Hepevirus, the only member of the Hepeviridae family. HE is usually transmitted via the faecal-oral route due to the fact that drinking water or water for industrial purposes is contaminated due to poor sanitation. This spread of HEV has been reported in developing countries of Asia, Africa, South and Central America. However, cases in countries with the sporadic occurrence of HEV have been associated with travelling to countries with an increased risk of infection (developing countries in Asia, Africa and America). HEV infections have subsequently been described in people who have not travelled to endemic countries. Further studies of th...

Veterinární medicína, 2012

Viruses cause many diseases in plants, animals, and humans. They are strict intracellular parasit... more Viruses cause many diseases in plants, animals, and humans. They are strict intracellular parasites with cellular specificity. Viral particles can be transmitted by different routes, such as contaminated food and water. People usually get infected orally, after ingestion of products contaminated during processing or subsequent handling or preparation. This review article is focused on the most severe foodborne viruses specific for humans, of the following genera: Norovirus, Enterovirus, Hepatovirus, Astrovirus, and some others. Methods for detecting viruses in food and strategies for preventing virus transmission via food are also discussed.

The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, ori-... more The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, ori-ginally found in seeds of amaranth (Amaranthus caudatus), into the genome of potatoes (So-lanum tuberosum). The purpose of the present study is to develop a PCR method for the detection of the mentioned genetically modified potatoes in various foodstuffs. The meth-od was used to test twenty different potato-based products; none of them was positive for the genetic modification 'ac2'. The European Union legislation requires labelling of pro-ducts made of or containing more than 0.9 % of genetically modified organisms. The gene-tic modification 'ac2' is not allowed on the European Union market. For that reason it is suitable to have detection methods, not only for the approved genetic modifications, but also for the 'unknown' ones, which could still occur in foodstuffs.

The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, orig... more The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, originally found in seeds of amaranth (Amaranthus caudatus), into the genome of potatoes (Solanum tuberosum). The purpose of the present study is to develop a PCR method for the detection of the mentioned genetically modified potatoes in various foodstuffs. The method was used to test twenty different potato-based products; none of them was positive for the genetic modification 'ac2'. The European Union legislation requires labelling of products made of or containing more than 0.9 % of genetically modified organisms. The genetic modification 'ac2' is not allowed on the European Union market. For that reason it is suitable to have detection methods, not only for the approved genetic modifications, but also for the 'unknown' ones, which could still occur in foodstuffs.

Journal of dairy science, 2012

The aim of this study was to assess the suitability of real-time quantitative PCR (qPCR) for the ... more The aim of this study was to assess the suitability of real-time quantitative PCR (qPCR) for the detection of Mycobacterium avium ssp. paratuberculosis (MAP) in milk filters as a herd level indicator of paratuberculosis infection. Seventy-nine samples from textile or metal milk filters from 15 herds with defined MAP prevalence (infection status = noninfected, 0-5%, 5-10%, or >10% of animals with clinically confirmed paratuberculosis) were analyzed. The MAP DNA was isolated by a modified commercially available protocol for feces, and detection and quantification of the pathogen was performed by the IS900 qPCR. Mycobacterium avium ssp. paratuberculosis DNA was detected in 63 (79.7%) samples. Determination of MAP infection established by fecal and tissue culture was correctly confirmed by the analysis of milk filters on 11 of 12 infected farms; MAP was not detected in filters from 3 farms where paratuberculosis was never diagnosed. Statistical analysis of the data supports the evide...

Food and Environmental Virology, 2010

Viruses are the causative agents of an estimated 60% of human infections worldwide. The most comm... more Viruses are the causative agents of an estimated 60% of human infections worldwide. The most common viral illnesses are produced by enteric and respiratory viruses. Transmission of these viruses from an infected person or animal to a new host can occur via several routes. Existing studies strongly suggest that contaminated fomites or surfaces play an important role in the spreading of viral diseases. The potential of viral spreading via contaminated surfaces depends particularly on the ability of the virus to maintain infectivity whilst it is in the environment. This is affected by a combination of biological, physical and chemical factors. This review summarises current knowledge about the influence of environmental factors on the survival and spread of viruses via contaminated surfaces.

Real-Time Quantitative PCR Detection of Mycobacterium avium Subspecies in Meat Products

Journal of Food Protection, 2011

The aim of this work was to examine various purchased meat products and to find out if any traces... more The aim of this work was to examine various purchased meat products and to find out if any traces of Mycobacterium avium subsp. avium, M. avium subsp. hominissuis, and M. avium subsp. paratuberculosis could be detected in these samples. Analysis of the meat products (raw, cooked, and fermented) was performed using a real-time quantitative PCR (qPCR) method for the detection of specific insertion sequences: duplex qPCR for the detection of IS900 specific for M. avium subsp. paratuberculosis, and triplex qPCR for the detection of IS901 specific for Mycobacterium avium subsp. avium and IS1245 specific for M. avium subsp. hominissuis. Of the 77 analyzed meat samples, 17 (22%) were found to contain M. avium subsp. paratuberculosis DNA, 4 (5%) samples contained Mycobacterium avium subsp. avium DNA, and in 12 (16%) samples M. avium subsp. hominissuis DNA was detected. The concentration of M. avium subsp. paratuberculosis and M. avium subsp. hominissuis DNA in some meat products exceeded 10...

International Journal of Food Microbiology, 2013

In recent years, numerous foodborne outbreaks due to consumption of berry fruit contaminated by h... more In recent years, numerous foodborne outbreaks due to consumption of berry fruit contaminated by human enteric viruses have been reported. This European multinational study investigated possible contamination routes by monitoring the entire food chain for a panel of human and animal enteric viruses. A total of 785 samples were collected throughout the food production chain of four European countries (Czech Republic, Finland, Poland and Serbia) during two growing seasons. Samples were taken during the production phase, the processing phase, and at point-of-sale. Samples included irrigation water, animal faeces, food handlers' hand swabs, swabs from toilets on farms, from conveyor belts at processing plants, and of raspberries or strawberries at points-of-sale; all were subjected to virus analysis. The samples were analysed by real-time (reverse transcription, RT)-PCR, primarily for human adenoviruses (hAdV) to demonstrate that a route of contamination existed from infected persons to the food supply chain. The analyses also included testing for the presence of selected human (norovirus, NoV GI, NoV GII and hepatitis A virus, HAV), animal (porcine adenovirus, pAdV and bovine polyomavirus, bPyV) and zoonotic (hepatitis E virus, HEV) viruses. At berry production, hAdV was found in 9.5%, 5.8% and 9.1% of samples of irrigation water, food handlers' hands and toilets, respectively. At the processing plants, hAdV was detected in one (2.0%) swab from a food handler's hand. At point-of-sale, the prevalence of hAdV in fresh raspberries, frozen raspberries and fresh strawberries, was 0.7%, 3.2% and 2.0%, respectively. Of the human pathogenic viruses, NoV GII was detected in two (3.6%) water samples at berry production, but no HAV was detected in any of the samples. HEV-contaminated frozen raspberries were found once (2.6%). Animal faecal contamination was evidenced by positive pAdV and bPyV assay results. At berry production, one water sample contained both viruses, and at point-of-sale 5.7% and 1.3% of fresh and frozen berries tested positive for pAdV. At berry production hAdV was found both in irrigation water and on food handler's hands, which indicated that these may be important vehicles by which human pathogenic viruses enter the berry fruit chain. Moreover, both zoonotic and animal enteric viruses could be detected on the end products. This study gives insight into viral sources and transmission routes and emphasizes the necessity for thorough compliance with good agricultural and hygienic practice at the farms to help protect the public from viral infections.

BMC Research Notes, 2012

Background: Different approaches are used for determining the number of Mycobacterium avium subsp... more Background: Different approaches are used for determining the number of Mycobacterium avium subsp. paratuberculosis (MAP) cells in a suspension. The majority of them are based upon culture (determination of CFU) or visual/instrumental direct counting of MAP cells. In this study, we have compared the culture method with a previously published F57 based quantitative real-time PCR (F57qPCR) method, to determine their relative abilities to count the number of three different MAP isolates in suspensions with the same optical densities (OD). McFarland turbidity standards were also compared with F57qPCR and culture, due to its frequent inclusion and use in MAP studies. Findings: The numbers of MAP in two-fold serial dilutions of isolates with respective OD measurements were determined by F57qPCR and culture. It was found that culture provided lower MAP CFU counts by approximately two log 10 , compared to F57qPCR. The McFarland standards (as defined for E. coli) showed an almost perfect fit with the enumeration of MAP performed by F57qPCR. Conclusions: It is recommended to use culture and/or qPCR estimations of MAP numbers in experiments where all subsequent counts are performed using the same method. It is certainly not recommended the use of culture as the standard for qPCR experiments and vice versa.

Biologia, 2008

Using primers to amplify the gene AMP2 in Amaranthus caudatus, we found the gene to be present in... more Using primers to amplify the gene AMP2 in Amaranthus caudatus, we found the gene to be present in seven other species of the Amaranthus genus (A. albus, A. cruentus, A. blitum, A. hybridus, A. hypochondriacus, A. retroflexus and A. tricolor), in which it had not been described previously. The PCR products were sequenced and it was established that all the sequences were identical, except for two polymorphisms. These single nucleotide polymorphisms occurred at nucleotide positions 45 and 246. This exchange of one nucleotide for another was manifested in an amino acid change in both cases. Due to the fact that both polymorphisms lay outside the region encoding the chitin-binding peptide domain, which is crucial for antimicrobial peptide function, they will not likely affect the proper functioning of the peptide. With the exception of the above-mentioned polymorphisms, all sequences were identical to the sequence of the AMP2 gene that codes for the A. caudatus Ac-AMP2 (antimicrobial pe...

Veterinary Microbiology, 2008

Mycobacterium avium subsp. avium (MAA) of serotype 2 and genotype IS901+ and IS1245+ was cultured... more Mycobacterium avium subsp. avium (MAA) of serotype 2 and genotype IS901+ and IS1245+ was cultured from 21 naturally infected hens (Gallus domesticus) from one smallholder aviary. From a total of 330 samples taken from hens, 124 mycobacteria were detected. Out of which MAA was detected in 103 (35.7%) of 288 tissues, in 4 (19.0%) of 21 swabs of cloacae and in 9 (42.9%) of 21 faeces samples, 8 other conditionally pathogenic mycobacterial species were also isolated. Tuberculous (TB) lesions were found in the liver, spleen and intestinal organs of 7 hens. The isolates of MAA (n = 58) from 16 infected hens ( with TB lesions and 9 without TB lesions) were found to be of three IS901 RFLP types AE (n = 48), AD (n = 4) and E (n = 6), where these MAA isolates are highly virulent to hens. Mixed infections with IS901 RFLP types (AE and AD) and (AE and E) were also evident in 7 hens. From a total of 35 examined environmental samples, 23 mycobacterial isolates were detected. Out of which, 4 (17.4%) MAA isolates of IS901 RFLP type AE and 19 (82.6%) other isolates of conditionally pathogenic mycobacteria were detected. The finding of identical IS901 RFLP types from both tissues and faecal isolates confirms that infected domestic hens are the principal source of infection for other susceptible hosts and lead to the contamination of the surrounding environment. The presence of different IS901 RFLP types in tissue isolates may indicate the repeated incidence of MAA infection and the occurrence of polyclonal infection.

Applied and Environmental Microbiology, 2012

The aim of this study was to monitor the persistence of Mycobacterium avium subsp. paratuberculos... more The aim of this study was to monitor the persistence of Mycobacterium avium subsp. paratuberculosis in environmental samples taken from a Holstein farm with a long history of clinical paratuberculosis. A herd of 606 head was eradicated, and mechanical cleaning and disinfection with chloramine B with ammonium (4%) was carried out on the farm; in the surrounding areas (on the field and field midden) lime was applied. Environmental samples were collected before and over a period of 24 months after destocking. Only one sample out of 48 (2%) examined on the farm (originating from a waste pit and collected before destocking) was positive for M. avium subsp. paratuberculosis by cultivation on solid medium (Herrold's egg yolk medium). The results using real-time quantitative PCR (qPCR) showed that a total of 81% of environmental samples with an average mean M. avium subsp. paratuberculosis cell number of 3.09 × 10 3 were positive for M. avium subsp. paratuberculosis before destocking co...

Veterinární medicína, 2011

This study reports on the transmission of Mycobacterium tuberculosis of the same IS6110 RFLP type... more This study reports on the transmission of Mycobacterium tuberculosis of the same IS6110 RFLP type between two acquaintances with open pulmonary tuberculosis and a five-year-old Doberman bitch. No clinical signs, gross lesions at necropsy or histopathological lesions were observed in the infected lungs and gastrointestinal tract of the dog, although M. tuberculosis was directly detected by IS6110 PCR and culture examinations in the respiratory and gastrointestinal tracts. IS6110 PCR positivity in the faeces and blood of the dog poses a risk of M. tuberculosis transmission between the dog and humans.

Veterinární medicína, 2010

A five-year old female brown caiman (Caiman crocodilus fuscus) was admitted to a veterinary clini... more A five-year old female brown caiman (Caiman crocodilus fuscus) was admitted to a veterinary clinic because of anorexia and lethargy. Chronic deterioration of the patient’s condition together with the formation of slushy stools coloured from brown to red was observed during the previous eight weeks. Physical examination showed significant apathy and cachexia. Radiographic examinations of chest and abdomen revealed no pathological findings. Initial blood tests revealed decreased hematocrit and low levels of haemoglobin. Despite treatment with enrofloxacin and intensive supportive therapy with amino acids, vitamins and mineral matter, the animal died 14 days after admission to the clinic. Post mortem examination revealed splenomegaly with a total destruction of inner organ structure together with multiple granulomas in liver and lungs. Ziehl-Neelsen staining of tissue samples from liver, lungs and spleen revealed numerous acid-fast bacilli consistent with Mycobacterium spp. Identificat...

Veterinární medicína, 2012

Peat is an easily available natural material and a source of biologically active substances widel... more Peat is an easily available natural material and a source of biologically active substances widely used, not only in agriculture but in human and animal medicine as well. In recent years, interest in the use of peat as a feed supplement has increased, particularly due to its capability to prevent enteric diseases and to stimulate growth in piglets and pigs. The purpose of this review was to compare the advantages and risks associated with the use of peat for animal nutrition based on the literature available. Beneficial effects of various peat preparations on digestion, growth and the immune systems of animals as well as the absorbent and detoxifying capabilities are associated with the high content of favourable humic substances. One disadvantage of using peat preparations is the considerable diversity of the various types of peat caused by different biological, chemical and geological conditions during formation. Biological activity of various peat preparations is associated not o...

Czech Journal of Food Sciences, Dec 31, 2011

We assessed the survival of Mycobacterium avium subsp. hominissuis (MAH) in artificially contamin... more We assessed the survival of Mycobacterium avium subsp. hominissuis (MAH) in artificially contaminated homemade smoked sausages prepared from pork meat according to traditional recipes, the effect of storage of such sausages at -20°C for three months on MAH viability and to compare assessment of MAH viability/presence by culture and qPCR. Three isolates of MAH were inoculated into the sausage mixture at concentrations of 10 6 CFU per gram of meat and cold smoked at 40°C for 12 h or hot smoked at 70°C for 6 hours. MAH survived the cold smoking procedure without any significant decrease in viable MAH CFU counts; no viable MAH were detected in the hot smoked sausages. The storage of sausages at -20°C caused a decrease in viable MAH counts of about 1 to 3 log 10 . Absolute MAH counts determined by qPCR were not significantly reduced by the storage or smoking. The presence of viable MAH in sausages after the cold smoking should be considered as a risk for immunodeficient individuals and children.

Food Technology and Biotechnology, Jun 9, 2009

The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, originally f... more The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, originally found in seeds of amaranth (Amaranthus caudatus), into the genome of potatoes (Solanum tuberosum). The purpose of the present study is to develop a PCR method for the detection of the mentioned genetically modified potatoes in various foodstuffs. The method was used to test twenty different potato-based products; none of them was positive for the genetic modification 'ac2'. The European Union legislation requires labelling of products made of or containing more than 0.9 % of genetically modified organisms. The genetic modification 'ac2' is not allowed on the European Union market. For that reason it is suitable to have detection methods, not only for the approved genetic modifications, but also for the 'unknown' ones, which could still occur in foodstuffs.

The "atypical mycobacteria" are widely distributed in the natural environment and can produce inf... more The "atypical mycobacteria" are widely distributed in the natural environment and can produce infection in farm and wild animals bred in captivity. The objective of the study was to analyze the variety of mycobacterial species in the environment of breeding facilities, tanks and aquariums, as well as in samples of peat used as a feed supplement in the Czech Republic over the 2003-2004 period. A total of 1389 samples from environment were examined, collected from 29 sites throughout the Czech Republic. The samples were decontaminated and cultured at 25 °C and 37 °C in three culture media: Stonebrink's medium, Herrold's egg yolk medium and Sula's medium. Mycobacterial isolates grown were identified by the PCR method and growth and biochemical tests. Of the 1389 environmental samples, mycobacteria were demonstrated by culture in 400 (28.8%) of them originating from different substances. In the samples of farm environment and in peat, mycobacteria were isolated in 185 of 1064 (17.4%) samples, and in 201 of 325 (61.8%) samples from aquarium and breeding tank environment. The results show significant difference between the findings from the stable environment (17.4%) and those from the aquarium environment (61.8%). A wide range of species was found in all constituents of the environment. The following species were most frequently isolated from the stable environment: M. avium (6.5%), M. fortuitum (3.2%), M. flavescens (3.2%), and the aquarium environment harboured most frequently M. fortuitum (13.9%), M. marinum (8.0%) and M. gordonae (4.5%). The results confirm the frequent occurrence of various mycobacterial species in the environment, predominantly in aquariums, some of which can produce infection in animals and in man following contact with the infected environment.

Veterinární medicína

In 2001, genetically modified potatoes carrying the ac2 gene were developed. This gene was origin... more In 2001, genetically modified potatoes carrying the ac2 gene were developed. This gene was originally found in amaranth seeds (Amaranthus caudatus) and is expressed into the fungicidal peptide Ac-AMP2. The purpose of the present study was to test 105 potato cultivars (Solanum tuberosum) registered in the Czech Republic for the presence of the above mentioned genetic modification "ac2" with the use of a previously published method . The method was based on the simultaneous detection of the ac2 gene from amaranth seeds as well as the StTS1 gene from potatoes as an internal amplification control. The results showed none of tested potatoes cultivars were positive for the genetic modification "ac2". These results confirmed the currently valid legislative in the Czech Republic and the European Union, where the use of genetically modified potatoes carrying the gene for the fungicidal peptide from amaranth is not allowed, was respected.

Veterinární medicína, 2012

Genetically modified organisms (GMO) become a real constituent of our lives and nowadays, they ar... more Genetically modified organisms (GMO) become a real constituent of our lives and nowadays, they are commonly introduced into the food chain of people and animals in some states. Among higher organisms, plants are used above all for genetic modifications; potatoes are a suitable model plants for this purpose. Nowadays, a number of various genetic modifications of potato plants are available, particularly those with increased resistance to biological agents and factors of the external environment or with improved nutritional value. Plants that produce proteins of the immune system of man or animals or substances that may be used as vaccines in human or veterinary medicine are highly important. Modified potato plants that produce biomaterials for potential applications in the industry are a significant category.

Veterinární medicína, 2008

The hepatitis E virus (HEV), the causative agent of hepatitis E, is a non-enveloped RNA virus. Th... more The hepatitis E virus (HEV), the causative agent of hepatitis E, is a non-enveloped RNA virus. The HEV genome is formed by a non-segmented positive-sense RNA chain. The 3´end of the chain is polyadenylated and the 5´end is structurally characterised by the so called “capping”. According to currently accepted taxonomy, HEV is classified in the genus Hepevirus, the only member of the Hepeviridae family. HE is usually transmitted via the faecal-oral route due to the fact that drinking water or water for industrial purposes is contaminated due to poor sanitation. This spread of HEV has been reported in developing countries of Asia, Africa, South and Central America. However, cases in countries with the sporadic occurrence of HEV have been associated with travelling to countries with an increased risk of infection (developing countries in Asia, Africa and America). HEV infections have subsequently been described in people who have not travelled to endemic countries. Further studies of th...

Veterinární medicína, 2012

Viruses cause many diseases in plants, animals, and humans. They are strict intracellular parasit... more Viruses cause many diseases in plants, animals, and humans. They are strict intracellular parasites with cellular specificity. Viral particles can be transmitted by different routes, such as contaminated food and water. People usually get infected orally, after ingestion of products contaminated during processing or subsequent handling or preparation. This review article is focused on the most severe foodborne viruses specific for humans, of the following genera: Norovirus, Enterovirus, Hepatovirus, Astrovirus, and some others. Methods for detecting viruses in food and strategies for preventing virus transmission via food are also discussed.

The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, ori-... more The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, ori-ginally found in seeds of amaranth (Amaranthus caudatus), into the genome of potatoes (So-lanum tuberosum). The purpose of the present study is to develop a PCR method for the detection of the mentioned genetically modified potatoes in various foodstuffs. The meth-od was used to test twenty different potato-based products; none of them was positive for the genetic modification 'ac2'. The European Union legislation requires labelling of pro-ducts made of or containing more than 0.9 % of genetically modified organisms. The gene-tic modification 'ac2' is not allowed on the European Union market. For that reason it is suitable to have detection methods, not only for the approved genetic modifications, but also for the 'unknown' ones, which could still occur in foodstuffs.

The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, orig... more The genetic modification 'ac2' is based on the insertion and expression of ac2 gene, originally found in seeds of amaranth (Amaranthus caudatus), into the genome of potatoes (Solanum tuberosum). The purpose of the present study is to develop a PCR method for the detection of the mentioned genetically modified potatoes in various foodstuffs. The method was used to test twenty different potato-based products; none of them was positive for the genetic modification 'ac2'. The European Union legislation requires labelling of products made of or containing more than 0.9 % of genetically modified organisms. The genetic modification 'ac2' is not allowed on the European Union market. For that reason it is suitable to have detection methods, not only for the approved genetic modifications, but also for the 'unknown' ones, which could still occur in foodstuffs.

Journal of dairy science, 2012

The aim of this study was to assess the suitability of real-time quantitative PCR (qPCR) for the ... more The aim of this study was to assess the suitability of real-time quantitative PCR (qPCR) for the detection of Mycobacterium avium ssp. paratuberculosis (MAP) in milk filters as a herd level indicator of paratuberculosis infection. Seventy-nine samples from textile or metal milk filters from 15 herds with defined MAP prevalence (infection status = noninfected, 0-5%, 5-10%, or >10% of animals with clinically confirmed paratuberculosis) were analyzed. The MAP DNA was isolated by a modified commercially available protocol for feces, and detection and quantification of the pathogen was performed by the IS900 qPCR. Mycobacterium avium ssp. paratuberculosis DNA was detected in 63 (79.7%) samples. Determination of MAP infection established by fecal and tissue culture was correctly confirmed by the analysis of milk filters on 11 of 12 infected farms; MAP was not detected in filters from 3 farms where paratuberculosis was never diagnosed. Statistical analysis of the data supports the evide...

Food and Environmental Virology, 2010

Viruses are the causative agents of an estimated 60% of human infections worldwide. The most comm... more Viruses are the causative agents of an estimated 60% of human infections worldwide. The most common viral illnesses are produced by enteric and respiratory viruses. Transmission of these viruses from an infected person or animal to a new host can occur via several routes. Existing studies strongly suggest that contaminated fomites or surfaces play an important role in the spreading of viral diseases. The potential of viral spreading via contaminated surfaces depends particularly on the ability of the virus to maintain infectivity whilst it is in the environment. This is affected by a combination of biological, physical and chemical factors. This review summarises current knowledge about the influence of environmental factors on the survival and spread of viruses via contaminated surfaces.

Real-Time Quantitative PCR Detection of Mycobacterium avium Subspecies in Meat Products

Journal of Food Protection, 2011

The aim of this work was to examine various purchased meat products and to find out if any traces... more The aim of this work was to examine various purchased meat products and to find out if any traces of Mycobacterium avium subsp. avium, M. avium subsp. hominissuis, and M. avium subsp. paratuberculosis could be detected in these samples. Analysis of the meat products (raw, cooked, and fermented) was performed using a real-time quantitative PCR (qPCR) method for the detection of specific insertion sequences: duplex qPCR for the detection of IS900 specific for M. avium subsp. paratuberculosis, and triplex qPCR for the detection of IS901 specific for Mycobacterium avium subsp. avium and IS1245 specific for M. avium subsp. hominissuis. Of the 77 analyzed meat samples, 17 (22%) were found to contain M. avium subsp. paratuberculosis DNA, 4 (5%) samples contained Mycobacterium avium subsp. avium DNA, and in 12 (16%) samples M. avium subsp. hominissuis DNA was detected. The concentration of M. avium subsp. paratuberculosis and M. avium subsp. hominissuis DNA in some meat products exceeded 10...

International Journal of Food Microbiology, 2013

In recent years, numerous foodborne outbreaks due to consumption of berry fruit contaminated by h... more In recent years, numerous foodborne outbreaks due to consumption of berry fruit contaminated by human enteric viruses have been reported. This European multinational study investigated possible contamination routes by monitoring the entire food chain for a panel of human and animal enteric viruses. A total of 785 samples were collected throughout the food production chain of four European countries (Czech Republic, Finland, Poland and Serbia) during two growing seasons. Samples were taken during the production phase, the processing phase, and at point-of-sale. Samples included irrigation water, animal faeces, food handlers' hand swabs, swabs from toilets on farms, from conveyor belts at processing plants, and of raspberries or strawberries at points-of-sale; all were subjected to virus analysis. The samples were analysed by real-time (reverse transcription, RT)-PCR, primarily for human adenoviruses (hAdV) to demonstrate that a route of contamination existed from infected persons to the food supply chain. The analyses also included testing for the presence of selected human (norovirus, NoV GI, NoV GII and hepatitis A virus, HAV), animal (porcine adenovirus, pAdV and bovine polyomavirus, bPyV) and zoonotic (hepatitis E virus, HEV) viruses. At berry production, hAdV was found in 9.5%, 5.8% and 9.1% of samples of irrigation water, food handlers' hands and toilets, respectively. At the processing plants, hAdV was detected in one (2.0%) swab from a food handler's hand. At point-of-sale, the prevalence of hAdV in fresh raspberries, frozen raspberries and fresh strawberries, was 0.7%, 3.2% and 2.0%, respectively. Of the human pathogenic viruses, NoV GII was detected in two (3.6%) water samples at berry production, but no HAV was detected in any of the samples. HEV-contaminated frozen raspberries were found once (2.6%). Animal faecal contamination was evidenced by positive pAdV and bPyV assay results. At berry production, one water sample contained both viruses, and at point-of-sale 5.7% and 1.3% of fresh and frozen berries tested positive for pAdV. At berry production hAdV was found both in irrigation water and on food handler's hands, which indicated that these may be important vehicles by which human pathogenic viruses enter the berry fruit chain. Moreover, both zoonotic and animal enteric viruses could be detected on the end products. This study gives insight into viral sources and transmission routes and emphasizes the necessity for thorough compliance with good agricultural and hygienic practice at the farms to help protect the public from viral infections.

BMC Research Notes, 2012

Background: Different approaches are used for determining the number of Mycobacterium avium subsp... more Background: Different approaches are used for determining the number of Mycobacterium avium subsp. paratuberculosis (MAP) cells in a suspension. The majority of them are based upon culture (determination of CFU) or visual/instrumental direct counting of MAP cells. In this study, we have compared the culture method with a previously published F57 based quantitative real-time PCR (F57qPCR) method, to determine their relative abilities to count the number of three different MAP isolates in suspensions with the same optical densities (OD). McFarland turbidity standards were also compared with F57qPCR and culture, due to its frequent inclusion and use in MAP studies. Findings: The numbers of MAP in two-fold serial dilutions of isolates with respective OD measurements were determined by F57qPCR and culture. It was found that culture provided lower MAP CFU counts by approximately two log 10 , compared to F57qPCR. The McFarland standards (as defined for E. coli) showed an almost perfect fit with the enumeration of MAP performed by F57qPCR. Conclusions: It is recommended to use culture and/or qPCR estimations of MAP numbers in experiments where all subsequent counts are performed using the same method. It is certainly not recommended the use of culture as the standard for qPCR experiments and vice versa.

Biologia, 2008

Using primers to amplify the gene AMP2 in Amaranthus caudatus, we found the gene to be present in... more Using primers to amplify the gene AMP2 in Amaranthus caudatus, we found the gene to be present in seven other species of the Amaranthus genus (A. albus, A. cruentus, A. blitum, A. hybridus, A. hypochondriacus, A. retroflexus and A. tricolor), in which it had not been described previously. The PCR products were sequenced and it was established that all the sequences were identical, except for two polymorphisms. These single nucleotide polymorphisms occurred at nucleotide positions 45 and 246. This exchange of one nucleotide for another was manifested in an amino acid change in both cases. Due to the fact that both polymorphisms lay outside the region encoding the chitin-binding peptide domain, which is crucial for antimicrobial peptide function, they will not likely affect the proper functioning of the peptide. With the exception of the above-mentioned polymorphisms, all sequences were identical to the sequence of the AMP2 gene that codes for the A. caudatus Ac-AMP2 (antimicrobial pe...

Veterinary Microbiology, 2008

Mycobacterium avium subsp. avium (MAA) of serotype 2 and genotype IS901+ and IS1245+ was cultured... more Mycobacterium avium subsp. avium (MAA) of serotype 2 and genotype IS901+ and IS1245+ was cultured from 21 naturally infected hens (Gallus domesticus) from one smallholder aviary. From a total of 330 samples taken from hens, 124 mycobacteria were detected. Out of which MAA was detected in 103 (35.7%) of 288 tissues, in 4 (19.0%) of 21 swabs of cloacae and in 9 (42.9%) of 21 faeces samples, 8 other conditionally pathogenic mycobacterial species were also isolated. Tuberculous (TB) lesions were found in the liver, spleen and intestinal organs of 7 hens. The isolates of MAA (n = 58) from 16 infected hens ( with TB lesions and 9 without TB lesions) were found to be of three IS901 RFLP types AE (n = 48), AD (n = 4) and E (n = 6), where these MAA isolates are highly virulent to hens. Mixed infections with IS901 RFLP types (AE and AD) and (AE and E) were also evident in 7 hens. From a total of 35 examined environmental samples, 23 mycobacterial isolates were detected. Out of which, 4 (17.4%) MAA isolates of IS901 RFLP type AE and 19 (82.6%) other isolates of conditionally pathogenic mycobacteria were detected. The finding of identical IS901 RFLP types from both tissues and faecal isolates confirms that infected domestic hens are the principal source of infection for other susceptible hosts and lead to the contamination of the surrounding environment. The presence of different IS901 RFLP types in tissue isolates may indicate the repeated incidence of MAA infection and the occurrence of polyclonal infection.

Applied and Environmental Microbiology, 2012

The aim of this study was to monitor the persistence of Mycobacterium avium subsp. paratuberculos... more The aim of this study was to monitor the persistence of Mycobacterium avium subsp. paratuberculosis in environmental samples taken from a Holstein farm with a long history of clinical paratuberculosis. A herd of 606 head was eradicated, and mechanical cleaning and disinfection with chloramine B with ammonium (4%) was carried out on the farm; in the surrounding areas (on the field and field midden) lime was applied. Environmental samples were collected before and over a period of 24 months after destocking. Only one sample out of 48 (2%) examined on the farm (originating from a waste pit and collected before destocking) was positive for M. avium subsp. paratuberculosis by cultivation on solid medium (Herrold's egg yolk medium). The results using real-time quantitative PCR (qPCR) showed that a total of 81% of environmental samples with an average mean M. avium subsp. paratuberculosis cell number of 3.09 × 10 3 were positive for M. avium subsp. paratuberculosis before destocking co...

Veterinární medicína, 2011

This study reports on the transmission of Mycobacterium tuberculosis of the same IS6110 RFLP type... more This study reports on the transmission of Mycobacterium tuberculosis of the same IS6110 RFLP type between two acquaintances with open pulmonary tuberculosis and a five-year-old Doberman bitch. No clinical signs, gross lesions at necropsy or histopathological lesions were observed in the infected lungs and gastrointestinal tract of the dog, although M. tuberculosis was directly detected by IS6110 PCR and culture examinations in the respiratory and gastrointestinal tracts. IS6110 PCR positivity in the faeces and blood of the dog poses a risk of M. tuberculosis transmission between the dog and humans.

Veterinární medicína, 2010

A five-year old female brown caiman (Caiman crocodilus fuscus) was admitted to a veterinary clini... more A five-year old female brown caiman (Caiman crocodilus fuscus) was admitted to a veterinary clinic because of anorexia and lethargy. Chronic deterioration of the patient’s condition together with the formation of slushy stools coloured from brown to red was observed during the previous eight weeks. Physical examination showed significant apathy and cachexia. Radiographic examinations of chest and abdomen revealed no pathological findings. Initial blood tests revealed decreased hematocrit and low levels of haemoglobin. Despite treatment with enrofloxacin and intensive supportive therapy with amino acids, vitamins and mineral matter, the animal died 14 days after admission to the clinic. Post mortem examination revealed splenomegaly with a total destruction of inner organ structure together with multiple granulomas in liver and lungs. Ziehl-Neelsen staining of tissue samples from liver, lungs and spleen revealed numerous acid-fast bacilli consistent with Mycobacterium spp. Identificat...

Veterinární medicína, 2012

Peat is an easily available natural material and a source of biologically active substances widel... more Peat is an easily available natural material and a source of biologically active substances widely used, not only in agriculture but in human and animal medicine as well. In recent years, interest in the use of peat as a feed supplement has increased, particularly due to its capability to prevent enteric diseases and to stimulate growth in piglets and pigs. The purpose of this review was to compare the advantages and risks associated with the use of peat for animal nutrition based on the literature available. Beneficial effects of various peat preparations on digestion, growth and the immune systems of animals as well as the absorbent and detoxifying capabilities are associated with the high content of favourable humic substances. One disadvantage of using peat preparations is the considerable diversity of the various types of peat caused by different biological, chemical and geological conditions during formation. Biological activity of various peat preparations is associated not o...