Identification of Bovine-Specific DNA in Feedstuffs

2001 ◽  
Vol 64 (1) ◽  
pp. 117-119 ◽  
Author(s):  
PAVEL KRČMÁŘ ◽  
EVA RENČOVÁ
Keyword(s):  
Polymerase Chain Reaction ◽  
Mitochondrial Dna ◽  
Czech Republic ◽  
Dna Sequences ◽  
Fish Meal ◽  
Spongiform Encephalopathy ◽  
Vertebrate Species ◽  
Chain Reaction ◽  
The Czech Republic ◽  
Polymerase Chain

Considering the menace of transmission of bovine spongiform encephalopathy, feed components intended for cattle nutrition must be checked for the presence of bovine-derived materials. We have been using a method based on polymerase chain reaction for the identification of bovine-specific mitochondrial DNA sequences for this purpose. The specificity of the primers for polymerase chain reaction has been tested using samples of DNA of other vertebrate species, which may also be present in rendering plant products. The method allows the detection in concentrate mixtures of 0.125% of bovine-derived material. Bovine DNA at concentrations corresponding to less than 0.5% of bovine-derived material was detected in 3 of the 30 samples of concentrate mixtures collected from distributors' stores all over the Czech Republic. All 44 samples of fish meal collected from the same sources were free of bovine-derived material.

scholarly journals Preliminary Study of Sars-Cov-2 Occurrence in Wastewater in the Czech Republic

2020 ◽  
Vol 17 (15) ◽  
pp. 5508 ◽  
Author(s):  
Hana Mlejnkova ◽  
Katerina Sovova ◽  
Petra Vasickova ◽  
Vera Ocenaskova ◽  
Lucie Jasikova ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Wastewater Treatment ◽  
Czech Republic ◽  
Wastewater Treatment Plants ◽  
Chain Reaction ◽  
The Czech Republic ◽  
Untreated Wastewater ◽  
Preliminary Study ◽  
Polymerase Chain

The virus SARS-CoV-2, which has caused the recent COVID-19 pandemic, may be present in the stools of COVID-19 patients. Therefore, we aimed to detect SARS-CoV-2 in wastewater for surveillance of SARS-CoV-2 in the population. Samples of untreated wastewater were collected from 33 wastewater treatment plants (WWTPs) of different sizes within the Czech Republic. SARS-CoV-2 RNA was concentrated from wastewater and viral RNA was determined using real-time reverse transcription polymerase chain reaction (RT-qPCR). SARS-CoV-2 RNA was detected in 11.6% of samples and more than 27.3% of WWTPs; in some of them, SARS-CoV-2 was detected repeatedly. Our preliminary results indicate that an epidemiology approach that focuses on the determination of SARS-CoV-2 in wastewater could be suitable for SARS-CoV-2 surveillance in the population.

scholarly journals PCR detection of Bartonella spp. in the dog

2014 ◽  
Vol 83 (2) ◽  
pp. 79-82 ◽  
Author(s):  
Jarmila Konvalinová ◽  
Vlasta Svobodová ◽  
Dobromila Molinková ◽  
Miroslav Svoboda
Keyword(s):  
Polymerase Chain Reaction ◽  
Czech Republic ◽  
Clinical Symptoms ◽  
Bartonella Henselae ◽  
Chain Reaction ◽  
The Czech Republic ◽  
Two Samples ◽  
Healthy Dogs ◽  
Polymerase Chain ◽  
First Time

Our study aimed at using PCR to identify the incidence ofBartonellaspp. in blood of dogs. Altogether 286 dogs of 92 breeds aged 3 month to 17 years were tested from October 2008 to December 2009. Healthy dogs as well as dogs with various clinical symptoms of disease were included in the group. Samples were tested by polymerase chain reaction (PCR) specific for the presence ofBartonellaspp. Following the DNA examination in 286 dogs by PCR and subsequent sequencing, two samples were identified asBartonella henselae(0.7%). Other species ofBartonellawere not found. It was the first time in the Czech Republic when incidence ofBartonellaspp. was determined in dogs.

scholarly journals Detection of selected antibiotic resistance genes using multiplex PCR assay in mastitis pathogens in the Czech Republic

2017 ◽  
Vol 86 (2) ◽  
pp. 167-174 ◽  
Author(s):  
Vladimir Pyatov ◽  
Irena Vrtková ◽  
Aleš Knoll
Keyword(s):  
Polymerase Chain Reaction ◽  
Antibiotic Resistance ◽  
Czech Republic ◽  
Resistance Genes ◽  
Multiplex Polymerase Chain Reaction ◽  
Antibiotic Resistance Genes ◽  
Chain Reaction ◽  
The Czech Republic ◽  
Polymerase Chain ◽  
Mastitis Pathogens

The aim of this research was to develop multiplex polymerase chain reaction assays for the detection of aminoglycoside (strA, strB), sulphonamide (sulI, sulII), tetracycline (tetA, tetB, tetK, tetM, tetO), macrolide and lincosamide (msrA, ermA, ermB, ermC, mefA/E) genes of resistance in mastitis pathogens (Escherichia coli, Staphylococcus aureus, Streptococcus uberis, Streptococcus agalactiae and Streptococcus dysgalactiae). Applying the established assays, we investigated the distribution of antibiotic resistance genes in the above mentioned species isolated from milk samples in the Czech Republic. Each assay consisted of seven pairs of primers. Six of them amplified fragments of antibiotic resistance genes and one pair a fragment of a species specific gene. Polymerase chain reaction conditions were optimized to amplify seven gene fragments simultaneously in one reaction. In total, 249 isolates were used, among which 111 were positive for E. coli, 52 for S. aureus and 86 for Streptococcus spp. The majority (60.2%) of bacteria carried at least one antibiotic resistance gene and 44.6% were multidrug-resistant. The designed multiplex polymerase chain reaction assays may be applied as diagnostic method to replace or complement standard techniques of antibiotic susceptibility testing in the mentioned pathogens.

Comparison of mitochondrial DNA sequences of seven morphospecies of black flies (Diptera: Simuliidae)

Genome ◽  
1991 ◽  
Vol 34 (2) ◽  
pp. 306-311 ◽  
Author(s):  
Bai Xiong ◽  
Thomas D. Kocher
Keyword(s):  
Polymerase Chain Reaction ◽  
Mitochondrial Dna ◽  
Dna Sequences ◽  
Polytene Chromosomes ◽  
Universal Primers ◽  
Homologous Region ◽  
Black Flies ◽  
Chain Reaction ◽  
Nucleotide Substitutions ◽  
Polymerase Chain

Universal primers constructed from the 16S ribosomal RNA gene in the Drosophila yakuba mitochondrial genome were successfully used to amplify, via the polymerase chain reaction, the homologous region of mitochondrial DNA from seven black fly morphospecies. Amplification was achieved from single larval salivary glands and from single adults preserved in Carnoy's fixative (ethanol – acetic acid, 3:1), allowing DNA sequences and polytene chromosome banding pattern data to be gathered from the same individuals. Nucleotide sequences of the amplified DNA segment (347 base pairs) were obtained from all the species examined. As in Drosophila, the nucleotide base composition of the sequenced segment from black flies had a high adenine (A) and thymine (T) content (A + T on average comprised 77% of all nucleotides.). Nucleotide differences among the seven species were observed at 59 positions (55 nucleotide substitutions and 4 deletions). There were more transversion differences than transition differences both among and within genera; the proportion of transversions was higher between genera than within genera. Most transversion differences were A ↔ T type, comprising 79% of all transversion differences and 50% of all sequence differences. Phylogenetic inference based strictly on transversion differences confirmed traditional generic and tribal groupings, i.e., Prosimulium fuscum (Syme &Davies) is close to Prosimulium magnum (Dyar &Shannon); Simulium decorum (Walker), Simulium venustum s.l. (Say), and Simulium vittatum s.l. (Zetterstedt) are close to each other; Stegopterna mutata (Malloch) and Cnephia dacotensis (Dyar &Shannon), which belong to the tribe Cnephiini, are grouped together.Key words: polymerase chain reaction, mitochondrial DNA, polytene chromosomes, phylogeny, black flies.

scholarly journals Distribution of Two Formae Speciales of Polymyxa Graminis in the Czech Republic

2017 ◽  
Vol 48 (2) ◽  
pp. 54-62
Author(s):  
L. Grimová ◽  
L. Winkowska ◽  
B. Špuláková ◽  
P. Růžičková ◽  
P. Ryšánek
Keyword(s):  
Polymerase Chain Reaction ◽  
Czech Republic ◽  
Soil Samples ◽  
Chain Reaction ◽  
The Czech Republic ◽  
Polymyxa Graminis ◽  
Formae Speciales ◽  
Crop Fields ◽  
Polymerase Chain ◽  
Mixed Populations

Abstract It has been shown that two formae speciales of P. graminis, namely f. sp. temperata (ribotype Pg-I) and f. sp. tepida (ribotype Pg-II), are widely distributed throughout temperate areas of Europe. In this study, the presence of both forms of the temperate Polymyxa spp. was identified in soil samples from different locations of the Czech Republic during a survey performed in 2012 and 2013. Based on polymerase chain reaction results, of the total 58 tested samples, 67.2% contained at least one monitored forma specialis. Specifically, P. graminis f. sp. temperata was detected in 48.3% of soil samples, while P. graminis f. sp. tepida was detected in 44.8% of samples. Mixed populations were found in 25.9% of the tested areas. This plasmodiophorid was confirmed not only in crop fields but also in meadows and forests in all explored regions. Our results extend the knowledge on the distribution of both ribotypes of P. graminis and provide the first evidence of f. sp. tepida within the Czech Republic.

scholarly journals Diagnostic efficacy of molecular assays for the viral haemorrhagic septicaemia virus isolates from the Czech Republic

2017 ◽  
Vol 86 (3) ◽  
pp. 207-212 ◽  
Author(s):  
Ľubomír Pojezdal ◽  
Dagmar Pokorová ◽  
Stanislava Reschová ◽  
Miroslava Palíková ◽  
Monika Vícenová ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Czech Republic ◽  
Real Time ◽  
Reverse Transcription ◽  
Chain Reaction ◽  
The Czech Republic ◽  
Haemorrhagic Septicaemia ◽  
Viral Haemorrhagic Septicaemia Virus ◽  
Polymerase Chain ◽  
The One

The diagnostic properties of the one-step real-time reverse-transcription polymerase chain reaction assay for viral haemorrhagic septicaemia virus detection were compared to methods currently in use in the Czech Republic, namely, virus isolation using the cell culture and conventional reverse-transcription polymerase chain reaction followed by the nested polymerase chain reaction. The assays were tested on a panel of 25 archived viral haemorrhagic septicaemia isolates and 8 archived infectious haematopoietic necrosis isolates obtained from monitoring and/or outbreaks of the diseases among farmed salmonids in the Czech Republic. The ability to detect the presence of the virus in the tissues of fish was tested on additional 32 field samples collected from the rainbow trout (Oncorhynchus mykiss), brown trout (Salmo trutta) and brook trout (Salvelinus fontinalis). The real-time assay showed the highest analytic sensitivity by detecting the presence of viral nucleic acid in samples with 10-7 dilution, whereas the sensitivity of the conventional polymerase chain reaction peaked at 10-5. Diagnostic specificity of both molecular assays was confirmed by absence of cross-reactivity with the infectious haematopoietic necrosis virus isolates. This, along with consistent results in the detection of the virus in the fish tissues, confirms that the one-step real-time reverse-transcription polymerase chain reaction is currently an optimal stand-alone diagnostic method for the detection of the viral haemorrhagic septicaemia virus.

Polymerase Chain Reaction–Based Analysis To Detect Terrestrial Animal Protein in Fish Meal

2003 ◽  
Vol 66 (4) ◽  
pp. 682-685 ◽  
Author(s):  
FEDERICA BELLAGAMBA ◽  
FRANCO VALFRÉ ◽  
SARA PANSERI ◽  
VITTORIO M. MORETTI
Keyword(s):  
Polymerase Chain Reaction ◽  
Fish Meal ◽  
Control Measures ◽  
Spongiform Encephalopathy ◽  
Chain Reaction ◽  
Monitoring Method ◽  
Meat Meal ◽  
Multiplex Pcr Assay ◽  
Nucleotide Sequence Variation ◽  
Polymerase Chain

The recent European bovine spongiform encephalopathy crisis has focused attention on the importance of adopting stringent control measures to avoid the risk of the diffusion of mad cow disease through meat meal–based animal feedstuffs. Potential adulteration of such feedstuffs with bone particles from terrestrial animals is determined by microscopic examination by law before the release of these feedstuffs for free circulation in the European Community. This study describes a DNA monitoring method to examine fish meal for contamination with mammalian and poultry products. A polymerase chain reaction (PCR) method based on the nucleotide sequence variation in the 12S ribosomal RNA gene of mitochondrial DNA was developed and evaluated. Three species-specific primer pairs were designed for the identification of ruminant, pig, and poultry DNA. The specificity of the primers used in the PCR was tested by comparison with DNA samples for several vertebrate species and confirmed. The PCR specifically detected mammalian and poultry adulteration in fish meals containing 0.125% beef, 0.125% sheep, 0.125% pig, 0.125% chicken, and 0.5% goat. A multiplex PCR assay for ruminant and pig adulteration was optimized and had a detection limit of 0.25%.

Analysis of ancient bone DNA: techniques and applications

1991 ◽  
Vol 333 (1268) ◽  
pp. 399-407 ◽  
Keyword(s):  
Polymerase Chain Reaction ◽  
Mitochondrial Dna ◽  
Dna Sequences ◽  
Genetic Information ◽  
Chain Reaction ◽  
Base Pairs ◽  
Dna Recovery ◽  
Human Femur ◽  
Ancient Bone ◽  
Polymerase Chain

The analysis of DNA from ancient bone has numerous applications in archaeology and molecular evolution. Significant amounts of genetic information can be recovered from ancient bone: mitochondrial DNA sequences of 800 base pairs have been amplified from a 750-year-old human femur by using the polymerase chain reaction. DNA recovery varies considerably between bone samples and is not dependent on the age of the specimen. We present the results of a study on a small number of bones from a mediaeval and a 17th-century cemetery in Abingdon showing the relation between gross preservation, microscopic preservation and DNA recovery.

Using a Polymerase Chain Reaction as a Complementary Test to Improve the Detection of Dermatophyte Fungus in Nails

2014 ◽  
Vol 104 (3) ◽  
pp. 233-237 ◽  
Author(s):  
María José Iglesias Sánchez ◽  
Ana María Pérez Pico ◽  
Félix Marcos Tejedor ◽  
María Jesús Iglesias Sánchez ◽  
Raquel Mayordomo Acevedo
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Sequences ◽  
Classical Method ◽  
Clinical Manifestations ◽  
Culture Method ◽  
Clinical Samples ◽  
Chain Reaction ◽  
Nested Polymerase Chain Reaction ◽  
Traditional Procedure ◽  
Polymerase Chain

Background Dermatomycoses are a group of pathologic abnormalities frequently seen in clinical practice, and their prevalence has increased in recent decades. Diagnostic confirmation of mycotic infection in nails is essential because there are several pathologic conditions with similar clinical manifestations. The classical method for confirming the presence of fungus in nail is microbiological culture and the identification of morphological structures by microscopy. Methods We devised a nested polymerase chain reaction (PCR) that amplifies specific DNA sequences of dermatophyte fungus that is notably faster than the 3 to 4 weeks that the traditional procedure takes. We compared this new technique and the conventional plate culture method in 225 nail samples. The results were subjected to statistical analysis. Results We found concordance in 78.2% of the samples analyzed by the two methods and increased sensitivity when simultaneously using the two methods to analyze clinical samples. Now we can confirm the presence of dermatophyte fungus in most of the positive samples in just 24 hours, and we have to wait for the result of culture only in negative PCR cases. Conclusions Although this PCR cannot, at present, substitute for the traditional culture method in the detection of dermatophyte infection of the nails, it can be used as a complementary technique because its main advantage lies in the significant reduction of time used for diagnosis, in addition to higher sensitivity.

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