scholarly journals Optimisation of the PCR method for the detection of Campylobacter jejuni and Campylobacter coli in samples of ready-to-eat chicken meals

2008 ◽  
Vol 26 (No. 4) ◽  
pp. 291-297
Author(s):  
Z. Šabatková ◽  
K. Demnerová ◽  
J. Pázlarová
Keyword(s):  
Campylobacter Jejuni ◽  
Internal Control ◽  
Fast Food ◽  
Campylobacter Coli ◽  
Duplex Pcr ◽  
Chain Reaction ◽  
Negative Results ◽  
Pcr Product ◽  
Pcr Method ◽  
Polymerase Chain

This work compared the use of polymerase chain reaction (PCR) and the conventional CSN/ISO/10272 culture-based methods in the detection of <I>Campylobacter</I> species in ready-to-eat meals made from chicken meat. PCR was carried out with the primers specific to <I>C. jejuni, C. coli, C. lari</I>, and was modified with an internal control. The detection of campylobacters by PCR was performed on both untreated and spiked samples of real food purchased in local stores. For PCR, the detection limit was 2 CFU/g after 48 h enrichment in Park and Sanders broth. Duplex PCR proved to be highly reliable in the detection of campylobacters in different food types. Without extra spiking, samples from a global fast food chain exhibited positive amplification of the PCR product while but negative results were obtained from the cultivation of the same samples.

scholarly journals Suitability of different tissue fixatives for subsequent PCR analysis of Cysticercus ovis

2012 ◽  
Vol 49 (2) ◽  
pp. 67-70 ◽  
Author(s):  
M. Kolesárová ◽  
R. Herich ◽  
M. Levkut ◽  
J. Čurlík ◽  
M. Levkut
Keyword(s):  
Retrospective Studies ◽  
Pcr Amplification ◽  
Pcr Analysis ◽  
Chain Reaction ◽  
Fresh Tissue ◽  
Carnoy’S Solution ◽  
Negative Results ◽  
Pcr Product ◽  
Polymerase Chain ◽  
Cysticercus Ovis

AbstractPCR amplification of specific DNA regions is a powerful tool for retrospective studies, but not all preservation or fixation methods render DNA that is suitable for subsequent amplification. Several factors affect sensitivity of polymerase chain reaction (PCR) amplification. There were reported the effects of commonly used fixation solutions — 10 % neutral buffered formalin, 20 % neutral buffered formalin and Carnoy’s solution and the efficiency of PCR amplification in fresh tissue and paraffin (or wax) embedded samples of Cysticercus ovis. DNA from samples was isolated and PCR product of 1300 bp was amplified. Results indicated that the samples fixed in Carnoy’s solution produced reliable amplification of desired fragments. The samples that were fixed in 10 % and 20 % neutral buffered formalin brought negative results.

scholarly journals Deteksi Plasmodium falciparum dengan menggunakan metode real-time polymerase chain reaction di daerah Likupang dan Bitung

2016 ◽  
Vol 4 (1) ◽  
Author(s):  
David C. Tooy ◽  
Janno B. Bernadus ◽  
Angle Sorisi
Keyword(s):  
Polymerase Chain Reaction ◽  
Plasmodium Falciparum ◽  
Real Time ◽  
Real Time Pcr ◽  
18S Rrna ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Negative Results ◽  
Pcr Method ◽  
Polymerase Chain

Abstract: Malaria is one of the most important parasitic disease which is caused by Plasmodium spp. There are approximately 1,2 billion people in the world with high risk of getting malaria. Plasmodium falciparum (P. falciparum) is the cause of tropical malaria or falciparum malaria, and is responsible for most of the mortality rate. Currently, real-time polymerase chain reaction (RT-PCR) is being studied as an alterative of conventional malarian examination. Mangold et al reported that RT-PCR have 94.1% sensitivity and 100% specificity compared to microscopic examination in detecting P. falciparum. The aim of this research is to detect the presence of P. falciparum using RT-PCR in Likupang and Bitung region. This research were using descriptive design to find out the capability of real-time PCR method to detect P. falciparum in Likupang dan Bitung region. The researcher have examined 71 samples which are fulfill the research sample’s criteria. Postive results of P. falciparum found in 18 samples (25,3%) and negative results in 53 samples (74,6%) of total 71 samples with using RT-PCR. No positive results were found in samples from Likupang. There are positive result of P. falciparum in samples from Bitung. It is concluded that RT-PCR method can detect the presence of P. falciparum from the samples obtained from Likupang and Bitung based on the presence of its DNA. This detection efford is done by using 18S rRNA as target gene and ajust specific temperature on the RT-PCR instrument.Keywords: Plasmodium falciparum, Real-time Polymerase Chain Reaction (PCR), DetectionAbstrak: Malaria merupakan salah satu penyakit penting yang disebabkan oleh parasit Plasmodium spp. Kira-kira 1,2 miliar penduduk dunia memiliki risiko tinggi untuk mendapat malaria. Di Indonesia sendiri, terdapat 343.527 kasus terkonfirmasi dan 45 kematian karena malaria. Plasmodium falciparum (P. Falciparum) merupakan penyebab dari malaria tropika atau malaria falsiparum, dan bertanggung jawab atas sebagian besar angka mortalitas. Saat ini Real-Time Polymerase Chain Reaction (RT-PCR) telah banyak diteliti sebagai alternatif dari pemeriksaan malaria. Mangold dkk melaporkan bahwa real-time PCR memiliki nilai sensitivitas 94,1% dan nilai spesifisitas 100% terhadap pemeriksaan mikroskopis dalam mendeteksi P. falciparum. Penelitian bertujuan untuk mendeteksi P. falciparum dengan menggunakan RT-PCR di daerah Likupang dan Bitung. Penelitian ini menggunakan rancangan penelitian deskriptif untuk mengetahui kemampuan metode real-time PCR dalam mendeteksi P. falciparum di daerah Likupang dan Bitung. Tujuan penelitian ini ialah untuk mendeteksi keberadaan P. falciparum dengan menggunakan metode real-time PCR di daerah Likupang dan Bitung. Peneliti memeriksa 71 sampel darah yang memenuhi kriteria sampel penelitian. Hasil positif P. falciparum ditemukan pada 18 sampel (25,3 %) dan hasil negatif pada 53 sampel (74,6 %) dari total 71 sampel dengan menggunakan RT-PCR. Tidak ditemukannya hasil positif P. falciparum pada sampel dari Likupang. Ditemukan hasil positif P. falciparum pada sampel dari Bitung. Simpulan: Metode RT-PCR dapat mendeteksi P. falciparum berdasarkan keberadaan DNA-nya pada sampel yang diperoleh dari daerah Likupang dan Bitung. Deteksi ini berhasil dilakukan dengan menggunakan 18S rRNA sebagai gen target dan pengaturan suhu tertentu pada instrument RT-PCR.Kata kunci: P. falciparum, Real-time Polymerase Chain Reaction (PCR), Detection

scholarly journals Species-Specific Polymerase Chain Reaction Amplification of Camel (Camelus) DNA Extracts

2005 ◽  
Vol 88 (5) ◽  
pp. 1394-1398 ◽  
Author(s):  
Ying Chen ◽  
Ya-Jun Wu ◽  
Bao-Liang Xu ◽  
Jing Wan ◽  
Zeng-Ming Qian
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Amplification ◽  
Chain Reaction ◽  
Pcr Product ◽  
Mitochondrial Cytochrome B ◽  
Base Pair Fragment ◽  
Sensitive Polymerase Chain Reaction ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Species Specific

Abstract A sensitive polymerase chain reaction (PCR) method based on amplification of a specific DNA fragment was established for the identification of camel (Camelus) materials. The species-specific primer pair L183/H372 was designed based on the nucleotide sequence of the mitochondrial cytochrome b gene, and its specificity was confirmed by amplification of 3 camel (domestic double-humped camel, wild double-humped camel, wild one-humped camel) samples and 11 non-Camelus animal (sheep, goat, pig, chicken, cattle, fish, dog, horse, donkey, deer, and rabbit) materials. An expected 208 base pair fragment was amplified from camel materials; no cross-reactive or additional fragments were generated from other animal materials. Taq I restriction endonuclease digestion of the unpurified PCR product can be used routinely to confirm the camel origin of the amplified sequence.

scholarly journals Survey of chicken abattoir for the presence of Campylobacter jejuni and Campylobacter coli

2006 ◽  
Vol 48 (6) ◽  
pp. 307-310 ◽  
Author(s):  
Ana L.L. Cortez ◽  
Angela C.F.B. Carvalho ◽  
Eliana Scarcelli ◽  
Simone Miyashiro ◽  
Ana M.C. Vidal-Martins ◽  
...  
Keyword(s):  
Public Health ◽  
Polymerase Chain Reaction ◽  
Campylobacter Jejuni ◽  
Intestinal Tract ◽  
Campylobacter Coli ◽  
Improve Quality ◽  
Chain Reaction ◽  
Paulo State ◽  
Polymerase Chain ◽  
Campylobacter Spp

The genus Campylobacter is of great importance to public health because it includes several species that may cause diarrhea. These species may be found in water, food and in the intestinal tract of chickens. This study investigated the presence of Campylobacter jejuni and Campylobacter coli in chicken abattoirs in São Paulo State, Brazil. A total of 288 samples of feces, feathers, scald water, evisceration water, chiller water, and the rinse water of eviscerated, not eviscerated and chilled carcasses were collected in six chicken abattoirs. Polymerase Chain Reaction (PCR) was performed in Campylobacter spp.-positive isolates using the gene HIP, specific for hippuricase enzyme from Campylobacter jejuni and aspartokinase gene, specific to detect Campylobacter coli. The percentage of positive isolates of Campylobacter jejuni was 4.9% (14/288). Isolation was greater in feces samples (22%, 8/36). One sample was positive for the species C. coli. In conclusion, the results indicate that it is necessary to improve quality control for Campylobacter spp. in chicken abattoirs.

Detection of Campylobacter jejuni and Campylobacter coli in Foods by Enrichment Culture and Polymerase Chain Reaction Enzyme-Linked Immunosorbent Assay

2002 ◽  
Vol 65 (5) ◽  
pp. 760-767 ◽  
Author(s):  
F. J. BOLTON ◽  
A. D. SAILS ◽  
A. J. FOX ◽  
D. R. A. WAREING ◽  
D. L. A. GREENWAY
Keyword(s):  
Polymerase Chain Reaction ◽  
Campylobacter Jejuni ◽  
Predictive Value ◽  
Enrichment Culture ◽  
Food Samples ◽  
Campylobacter Coli ◽  
Chain Reaction ◽  
Enrichment Cultures ◽  
Selective Agar ◽  
Polymerase Chain

A polymerase chain reaction (PCR) assay based on a solution hybridization format with colorimetric end-point detection (PCR ELISA) was investigated for the specific detection of Campylobacter jejuni and Campylobacter coli in food samples following enrichment culture. One hundred fifteen samples of raw meat and offal (poultry, porcine, ovine, and bovine), raw shellfish, and artificially contaminated milk were enriched in blood-free Campylobacter Enrichment Broth for 48 h. Enrichment cultures were subcultured to Campylobacter blood-free selective agar plates, and presumptive isolates were identified by phenotypic methods. DNA was extracted from 1-ml aliquots of the enrichment cultures using a rapid extraction method, and the DNA was used as the template in a PCR ELISA. A comparison of the PCR ELISA with the enrichment culture and subculture to selective agar method showed that the results of 112 of the 115 samples tested were in agreement by both methods. Seventy-one of the various food samples were positive in the PCR ELISA, and 70 samples were positive by culture. The PCR ELISA had a sensitivity of 99% and a specificity of 96%, with a positive predictive value of 97% and a negative predictive value of 98%. The PCR ELISA is a rapid, sensitive, and specific method for the detection of C. jejuni and C. coli in foods following enrichment culture and significantly reduces the time required for their detection.

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