scholarly journals Specific PCR Identification and Differentiation of the Thermophilic Campylobacters, Campylobacter jejuni, C. coli, C. lari, and C. upsaliensis

1999 ◽  
Vol 37 (10) ◽  
pp. 3370-3373 ◽  
Author(s):  
Christian Fermér ◽  
Eva Olsson Engvall
Keyword(s):  
Campylobacter Jejuni ◽  
Present Method ◽  
Restriction Enzymes ◽  
Species Differentiation ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Detection And Identification ◽  
Pcr Identification ◽  
Pcr Products

A sensitive PCR assay that detects the thermophilic campylobactersC. jejuni, C. coli, C. lari, andC. upsaliensis is reported. Furthermore, by digestion of the PCR products with two restriction enzymes, species differentiation was demonstrated. Thus, the present method has the potential to be used for both detection and identification of thermophilicCampylobacter species.

scholarly journals Development of a multiplex PCR assay for identification of Campylobacter coli, Campylobacter fetus, Campylobacter hyointestinalis subsp. hyointestinalis, Campylobacter jejuni, Campylobacter lari and Campylobacter upsaliensis

2007 ◽  
Vol 56 (11) ◽  
pp. 1467-1473 ◽  
Author(s):  
Wataru Yamazaki-Matsune ◽  
Masumi Taguchi ◽  
Kazuko Seto ◽  
Ryuji Kawahara ◽  
Kentaro Kawatsu ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Campylobacter Jejuni ◽  
Campylobacter Coli ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Campylobacter Fetus ◽  
Multiplex Pcr Assay ◽  
Pcr Products ◽  
Campylobacter Lari ◽  
Campylobacter Upsaliensis

A multiplex PCR assay has been developed for the identification of the six common Campylobacter taxa associated with human gastroenteritis and/or septicaemia, namely Campylobacter coli, Campylobacter fetus, Campylobacter hyointestinalis subsp. hyointestinalis, Campylobacter jejuni, Campylobacter lari and Campylobacter upsaliensis. The assay was developed using a combination of newly designed and published primers. It provided a specific PCR product for each of the five Campylobacter species and the one subspecies, and each of the PCR products was sufficiently distinguished by a difference in size by agarose gel electrophoresis. On evaluation of efficacy with 142 Campylobacter strains, the assay correctly identified all strains as 1 of the 6 Campylobacter taxa. This multiplex PCR assay is a rapid, simple and practical tool for identification of the six Campylobacter taxa commonly associated with gastroenteritis and/or septicaemia in humans, and offers an effective alternative to conventional biochemical-based assays.

scholarly journals Specific PCR detection of Peptostreptococcus magnus

2003 ◽  
Vol 52 (4) ◽  
pp. 309-313 ◽  
Author(s):  
M.P. Riggio ◽  
A. Lennon
Keyword(s):  
Pcr Primers ◽  
Oral Bacteria ◽  
Rrna Gene ◽  
Subgingival Plaque ◽  
Pcr Assay ◽  
Clinical Specimens ◽  
Specific Pcr ◽  
Wide Range ◽  
Pcr Products ◽  
Peptostreptococcus Magnus

Peptostreptococcus magnus is the most pathogenic and one of the most common Gram-positive anaerobic cocci found in human clinical specimens. The organism has been isolated in pure culture from a range of serious infections, including meningitis and endocarditis. However, isolation of Peptostreptococcus magnus from the oral cavity has rarely been attempted. Identification of Peptostreptococcus magnus in clinical specimens is reliant upon microbiological culture and biochemical methods, which often give ambiguous results. The aim of this study was to develop a PCR assay for the specific detection of Peptostreptococcus magnus in oral clinical specimens. PCR primers specific for Peptostreptococcus magnus DNA were derived by comparison of 16S rRNA gene sequences and selection of primers that demonstrated specificity at their 3′ ends for Peptostreptococcus magnus. PCR positivity for Peptostreptococcus magnus DNA was indicated by the amplification of a 553 bp product. The PCR assay was then used to attempt detection of Peptostreptococcus magnus DNA in subgingival plaque samples from adult periodontitis patients and pus aspirates from subjects with acute dento-alveolar abscesses. The PCR assay was demonstrated to be highly specific for Peptostreptococcus magnus DNA, since no PCR products were obtained when genomic DNA from a wide range of other oral bacteria, including closely related Peptostreptococcus species, was used in the PCR assay. Confirmation of specific amplification of Peptostreptococcus magnus DNA was obtained by digestion of PCR products with the restriction endonuclease RsaI, which gives a unique restriction profile for Peptostreptococcus magnus. Of the 33 subgingival plaque samples analysed, 2 (6 %) were positive for Peptostreptococcus magnus DNA. None of the 60 pus aspirates analysed was positive for Peptostreptococcus magnus DNA. It is concluded that Peptostreptococcus magnus is not a major pathogen in adult periodontitis or dento-alveolar abscesses. The PCR assay provides a more rapid, specific and sensitive alternative to conventional methods for identification of Peptostreptococcus magnus in clinical specimens.

scholarly journals Utilization of DNA microarrays for detection and identification of selected Fusarium species from the Czech Republic

2012 ◽  
Vol 29 (Special Issue) ◽  
pp. S93-S101
Author(s):  
L. Pavlátová ◽  
D. Novotný ◽  
J. Hodek ◽  
J. Chrpová ◽  
J. Ovesná
Keyword(s):  
Dna Microarrays ◽  
Fusarium Species ◽  
Elongation Factor ◽  
Translation Elongation ◽  
Specific Pcr ◽  
Detection And Identification ◽  
Pcr Products ◽  
Contaminated Food ◽  
Food And Feed ◽  
Template Dna

Fusarium is a serious phytopathogenic fungal genus with producting of many kinds of highly toxic secondary metabolites – mycotoxins. The consumption of Fusarium contaminated food and feed can cause dangerous mycotoxicoses both in humans and animals, therefore the detection of a wider range of Fusarium species in the samples of crops is very important. The aim of our work was to test the reliability of detection and identification of three Fusarium species in infected wheat grains by DNA microarrays versus classical mycological methods and by specific PCR. The in-house DNA microarrays for the detection and identification of the selected Fusarium species by using oligonucleotides probes were prepared. For hybridisation on DNA microarrays fluorescent labelled PCR products were used of part of the translation elongation factor 1 alpha. The conditions of hybridisation were optimised on fungal template DNA. The method of DNA microarrays was verified on artificially infected samples of wheat and tested on unknown infected wheat samples with simultaneous analysis by classical mycological methods and by specific PCR.

scholarly journals Molecular Cloning and Sequencing of thearoA Gene from Actinobacillus pleuropneumoniaeand Its Use in a PCR Assay for Rapid Identification

1999 ◽  
Vol 37 (5) ◽  
pp. 1575-1578 ◽  
Author(s):  
Carmen Hernanz Moral ◽  
Alberto Cascón Soriano ◽  
María Sánchez Salazar ◽  
Javier Yugueros Marcos ◽  
Susana Suárez Ramos ◽  
...  
Keyword(s):  
Rapid Identification ◽  
Chromosomal Dna ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Specificity And Sensitivity ◽  
Pcr Products ◽  
Dna Fragment ◽  
Specific Pcr Assay ◽  
Dna Specificity ◽  
K 12

The gene (aroA) of Actinobacillus pleuropneumoniae, serotype 2, encoding 5-enolpyruvylshikimate-3-phosphate synthase was cloned by complementation of the aroA mutation in Escherichia coli K-12 strain AB2829, and the nucleotide sequence was determined. A pair of primers from the 5′ and 3′ termini were selected to be the basis for development of a specific PCR assay. A DNA fragment of 1,025 bp was amplified from lysed A. pleuropneumoniaeserotypes 1 to 12 of biovar 1 or from isolated DNA. No PCR products were detected when chromosomal DNAs from other genera were used as target DNAs; however, a 1,025-bp DNA fragment was amplified whenActinobacillus equuli chromosomal DNA was used as a target, which could be easily differentiated by its NAD independence. The PCR assay developed was very sensitive, with lower detection limits of 12 CFU with A. pleuropneumoniae cells and 0.8 pg with extracted DNA. Specificity and sensitivity make this PCR assay a useful method for the rapid identification and diagnosis of A. pleuropneumoniae infections.

scholarly journals Species-Specific Plasmid Sequences for PCR Identification of the Three Species of Borrelia burgdorferiSensu Lato Involved in Lyme Disease

1998 ◽  
Vol 36 (1) ◽  
pp. 269-272 ◽  
Author(s):  
Marie-Christine Misonne ◽  
Philippe Pierre Hoet
Keyword(s):  
Lyme Disease ◽  
Pcr Amplification ◽  
Sensu Stricto ◽  
Pcr Assay ◽  
Borrelia Garinii ◽  
Specific Pcr ◽  
Pcr Identification ◽  
Identification Of Species ◽  
Species Specific ◽  
Specific Sequences

Species-specific sequences were shown to be carried by plasmids of the three main species of Borrelia burgdorferi sensu lato involved in Lyme disease. Libraries of the 16-, 33-, and 25-kb plasmids of B. burgdorferi sensu stricto, Borrelia garinii, and Borrelia afzelii, respectively, were then built and used to isolate species-specific sequences. After sequencing of the cloned inserts, three sets of primers were designed. They were shown to determine species-specific PCR amplification products. The sensitivities of the PCR assay with these primers were 100 spirochetes for B. burgdorferi sensu stricto and 1,000 spirochetes for B. garinii and B. afzelii. The usefulness of these primers for the identification of species in biological samples (tick, serum, and cerebrospinal fluid samples) was ascertained.

Molecular approaches in species identification of Sarcocysts in Indian buffalo meat

2018 ◽  
Author(s):  
C. Ramakrishn ◽  
S. Vaithiyanathan ◽  
M. Muthukumar ◽  
L., P. Lavanya and V.V. Kulkarni. R. Chatlod ◽  
P. Lavanya ◽  
...  
Keyword(s):  
Restriction Enzymes ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Restriction Enzyme Digestion ◽  
Pcr Assay ◽  
Naked Eye ◽  
Molecular Approaches ◽  
Pcr Product ◽  
Pcr Products ◽  
Eye Examination

DNA was extracted from sarcocysts (visible on naked eye examination) collected from 168 buffaloes belonging to Mumbai (60), Hyderabad (54) and Kolkata (54) cities of India. They were subjected to PCR assay using 18S rRNA gene primer. All the PCR amplicons of about 900 bp were subjected to restriction enzyme digestion with four different restriction enzymes (BslI, DraI, FokI and RsaI). PCR amplicons showed two different patterns (Pattern A and Pattern B) on RFLP. Twenty one PCR products from Pattern A and one PCR product from Pattern B were subjected to DNA sequencing. S. fusiformis and S. taeniata were identified from Pattern A and S. buffalonis was identified from Pattern B on sequencing.

Evaluation of a PCR assay for the detection and identification of Campylobacter jejuni and Campylobacter coli in retail poultry products

2005 ◽  
Vol 156 (4) ◽  
pp. 568-574 ◽  
Author(s):  
Estibaliz Mateo ◽  
Jose Cárcamo ◽  
Maria Urquijo ◽  
Ildefonso Perales ◽  
Aurora Fernández-Astorga
Keyword(s):  
Campylobacter Jejuni ◽  
Campylobacter Coli ◽  
Pcr Assay ◽  
Poultry Products ◽  
Detection And Identification

scholarly journals Arcobacter trophiarum sp. nov., isolated from fattening pigs

2011 ◽  
Vol 61 (2) ◽  
pp. 356-361 ◽  
Author(s):  
Sarah De Smet ◽  
Peter Vandamme ◽  
Lieven De Zutter ◽  
Stephen L. W. On ◽  
Laid Douidah ◽  
...  
Keyword(s):  
Sodium Deoxycholate ◽  
Cell Protein ◽  
Marker Genes ◽  
Polymorphism Analysis ◽  
Pcr Assay ◽  
Biochemical Tests ◽  
Specific Pcr ◽  
Detection And Identification ◽  
Species Specific ◽  
Dna Hybridizations

In the course of a longitudinal study elucidating the dynamics of Arcobacter populations in pigs, 16 isolates of Gram-reaction-negative, rod-shaped, slightly curved, non-spore-forming bacteria were grouped by amplified fragment length polymorphism analysis into a distinct phenon within the genus Arcobacter. Fragments were generated for all isolates in a genus-specific PCR assay, but no amplicon was obtained in a species-specific multiplex-PCR test. Numerical analysis of the whole-cell protein profiles also showed that all isolates clustered in a single group that was distinct from related members of the genus Arcobacter. DNA–DNA hybridizations between two representative strains, designated 64T and 122, of the isolates obtained exhibited a mean DNA–DNA relatedness of 72 %. DNA–DNA hybridizations between strains 64T and 122 and reference strains of other animal-related bacteria of the genus Arcobacter revealed binding values of 47 % or less. The DNA G+C contents of the two representative strains were 28.5 and 28.4 mol%, respectively, and analysis of three marker genes identified Arcobacter cryaerophilus, A. thereius, A. cibarius and A. skirrowii as their closest phylogenetic neighbours. Strains 64T and 122 could be distinguished from other members of the genus Arcobacter by means of biochemical tests for catalase and urease activities, nitrate reduction, indoxyl acetate hydrolysis, lack of growth at 37 °C, growth in 2 % (w/v) NaCl, growth on 0.1 % sodium deoxycholate and non-supplemented Campylobacter charcoal-deoxycholate base medium and resistance to cephalothin (32 mg l−1) and cefoperazone (64 mg l−1). Additionally, a PCR assay was developed for the detection and identification of strains 64T and 122, which represent a novel species of the genus Arcobacter, for which the name Arcobacter trophiarum sp. nov. is proposed. The type strain is strain 64T (=LMG 25534T =CCUG 59229T).

Rapid and reliable identification of rice genomes by RFLP analysis of PCR-amplified Adh genes

Genome ◽  
2001 ◽  
Vol 44 (6) ◽  
pp. 1136-1142 ◽  
Author(s):  
Song Ge ◽  
Tao Sang ◽  
Bao-rong Lu ◽  
De-yuan Hong
Keyword(s):  
Rflp Analysis ◽  
Restriction Enzymes ◽  
Pcr Primers ◽  
Specific Pcr ◽  
Adh Genes ◽  
Pcr Rflp ◽  
Restriction Sites ◽  
Pcr Products ◽  
Adh Gene ◽  
Genus Oryza

The rice genus (Oryza L.) consists of 24 species with 10 recognized genome types. With the realization of many useful genes in species of wild rice, continuous efforts have been made to understand their genomic composition and relationships. However, the identification of rice genomes has often been difficult owing to complex morphological variation and formation of allotetraploids. Here we propose a rapid and reliable method for identifying rice genomes based on the restriction sites of PCR-amplified Adh genes. The experimental procedure was as follows: (i) amplify a portion of Adh1 and Adh2 genes with the locus-specific PCR primers; (ii) digest PCR products with restriction enzymes that distinguish different genomes; and (iii) run the digested products on 1.4% agarose gel, and photograph. Using various combinations of restriction digestion of the two Adh genes, all of the rice genomes can be identified.Key words: Adh gene, genome, identification, Oryza L., PCR–RFLP.

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