scholarly journals Multiplex PCR Assay for Identifi cation and Differentiation of Campylobacter jejuni and Campylobacter coli Isolates

Folia Medica ◽  
2016 ◽  
Vol 58 (2) ◽  
pp. 95-100 ◽  
Author(s):  
Maria R. Pavlova ◽  
Elina G. Dobreva ◽  
Katucha I. Ivanova ◽  
Galina D. Asseva ◽  
Ivan N. Ivanov ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Campylobacter Jejuni ◽  
Accurate Method ◽  
Clinical Isolates ◽  
Campylobacter Coli ◽  
Pcr Assay ◽  
Biochemical Tests ◽  
Multiplex Pcr Assay ◽  
Hydrolysis Of ◽  
Campylobacter Spp

AbstractIntroduction: Campylobacter spp. are important causative agents of gastrointestinal infections in humans. The most frequently isolated strains of this bacterial genus are Campylobacter jejuni and Campylobacter coli. To date, genetic methods for bacterial identification have not been used in Bulgaria. We optimized the multiplex PSR assay to identify Campylobacter spp. and differentiate C. jejuni from C. coli in clinical isolates. We also compared this method with the routinely used biochemical methods.Aim: To identify Campylobacter spp. and discriminate C. coli from C. jejuni in clinical isolates using multiplex PCR assay.Materials and methods: Between February 2014 and January 2015 we studied 93 stool samples taken from patients with diarrheal syndrome and identified 40 species of Campylobacter spp. in them. The clinical material was cultured in microaerophilic atmosphere, the isolated strains being biochemically diff erentiated (hydrolysis of sodium hippurate for C. jejuni, and hydrolysis of indoxyl acetate for C. coli). DNA was isolated from the strains using QiaAmp MiniKit (QIAGEN, Germany). Twenty strains were tested with multiplex PCR for the presence of these genes: cadF, characteristic for Campylobacter spp., hipO for C. jejuni and asp for C. coli.Results and discussion: The biochemical tests identified 16 strains of C. jejuni, 3 strains of C. coli, and 1 strain of C. upsaliensis. After the multiplex PCR assay the capillary gel electrophoresis confirmed 16 strains of C. jejuni, 2 strains of C. coli and 2 strains of Campylobacter spp. - because of the presence of the gene cadF. C. jejuni has the gene hipO, and it is possible that this gene may not be expressed in the biochemical differentiation yielding a negative reaction as a result. In comparison, we can conclude that the genetic differentiation is a more accurate method than the biochemical tests.Conclusion: The multiplex PCR assay is a fast, accurate method for identifi cation of Campylobacter spp. which makes it quite necessary in the clinical diagnostic practice.

scholarly journals Evaluation of a Multiplex PCR Assay for the Identification of Campylobacter jejuni and Campylobacter coli

2017 ◽  
Vol 3 (1) ◽  
pp. 6-8
Author(s):  
Saeed Shams ◽  
Mehdi Ghorbanalizadgan ◽  
Somayeh Haj Mahmmodi ◽  
Alessandra Piccirillo ◽  
◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Campylobacter Jejuni ◽  
Campylobacter Coli ◽  
Pcr Assay ◽  
Multiplex Pcr Assay

scholarly journals Evaluation of a Multiplex PCR Assay for the Identification of Campylobacter jejuni and Campylobacter coli

2017 ◽  
Vol 3 (1) ◽  
pp. 6-8 ◽  
Author(s):  
Saeed Shams ◽  
◽  
Mehdi Ghorbanalizadgan ◽  
Somayeh Haj Mahmmodi ◽  
Alessandra Piccirillo ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Campylobacter Jejuni ◽  
Campylobacter Coli ◽  
Pcr Assay ◽  
Multiplex Pcr Assay

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.

Rapid Detection of Campylobacter jejuni, Campylobacter coli, and Campylobacter lari in Fresh Chicken Meat and By-Products in Bangkok, Thailand, Using Modified Multiplex PCR

2015 ◽  
Vol 78 (7) ◽  
pp. 1363-1369 ◽  
Author(s):  
S. SAIYUDTHONG ◽  
K. PHUSRI ◽  
S. BUATES
Keyword(s):  
Multiplex Pcr ◽  
Campylobacter Jejuni ◽  
Predictive Value ◽  
Chicken Meat ◽  
Campylobacter Coli ◽  
Fresh Market ◽  
Pcr Assay ◽  
Multiplex Pcr Assay ◽  
Campylobacter Lari ◽  
By Products

A multiplex PCR assay for simultaneous detection and differentiation of Campylobacter jejuni, Campylobacter coli, and Campylobacter lari was developed and validated to assess the occurrence of these bacteria in fresh chicken meat and by-products in Bangkok, Thailand, by using a new combination of four previously published PCR primers for C. jejuni, C. coli, C. lari, and a universal 16S rDNA gene as an internal control. The specificity was determined by using 13 strains of other bacteria. With pure culture DNA, the detection limit was 0.017 ng/PCR for C. jejuni and C. coli and was 0.016 ng/PCR for C. lari. It can detect 10 CFU of C. jejuni, C. coli, and C. lari in 2 g of chicken meat within a 16-h enrichment time. Our multiplex PCR assay was applied for identification of Campylobacter spp. in 122 supermarket samples and 108 fresh market samples. Of the 230 samples evaluated by multiplex PCR, 54.0, 3.3, and 10.7% of supermarket samples were positive for C. jejuni, C. coli, and mixed C. jejuni and C. coli, respectively, and 56.5 and 33.3% of fresh market samples were positive for C. jejuni and mixed C. jejuni and C. coli, respectively. No sample was positive for C. lari. Fresh market samples had significantly higher C. jejuni and C. coli contamination than those from supermarkets (relative risk: 1.3; P = 0.0001). Compared with the culture method (a gold standard), the sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy of multiplex PCR were 97.7, 86.8, 96.1, 92.0, and 95.2%, respectively. No significant difference was observed between results from two methods (P = 0.55). Therefore, the established multiplex PCR was not only rapid and easy to perform but had a high sensitivity and specificity to distinguish between C. jejuni, C. coli, and C. lari, even in samples containing mixed contamination. Our study indicated that fresh chicken meat and by-products from fresh markets were significantly less hygienic than those from supermarkets.

scholarly journals Multiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groups

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Woo Jun Bang ◽  
Heung Chul Kim ◽  
Jihun Ryu ◽  
Hyeon Seung Lee ◽  
So Youn Lee ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Molecular Identification ◽  
Malaria Infection ◽  
Diagnostic System ◽  
Accurate Method ◽  
Its2 Region ◽  
Infection Rates ◽  
Pcr Assay ◽  
Multiplex Pcr Assay ◽  
The Republic

Abstract Background Genus Anopheles mosquitoes are the primary vectors of human malaria, which is a serious threat to public health worldwide. To reduce the spread of malaria and identify the malaria infection rates in mosquitoes, accurate species identification is needed. Malaria re-emerged in 1993 in the Republic of Korea (ROK), with numbers peaking in 2004 before decreasing to current levels. Eight Anopheles species (Anopheles sinensis, Anopheles pullus, Anopheles belenrae, Anopheles lesteri, Anopheles kleini, Anopheles sineroides, Anopheles koreicus, Anopheles lindesayi) are distributed throughout Korea. Members of the Anopheles Hyrcanus group currently cannot be identified morphologically. The other species of Anopheles can be identified morphologically, except when specimens are damaged in traps. The purpose of this study was to develop a rapid and accurate method for simultaneous molecular identification of the eight Anopheles species present in the ROK. Methods Anopheles spp. used in this study were collected near/in the demilitarized zone in ROK, where most malaria cases are reported. DNA from 165 of the Anopheles specimens was used to develop a multiplex PCR assay. The internal transcribed spacer 2 (ITS2) region of each species was sequenced and analysed for molecular identification. Results DNA from a total of 165 Anopheles specimens was identified to species using a multiplex diagnostic system. These included: 20 An. sinensis, 21 An. koreicus, 17 An. lindesayi, 25 An. kleini, 11 An. lesteri, 22 An. sineroides, 23 An. belenrae, and 26 An. pullus. Each species was clearly distinguished by electrophoresis as follows: 1,112 bp for An. sinensis; 925 bp for An. koreicus; 650 bp for An. lindesayi; 527 bp for An. kleini; 436 bp for An. lesteri; 315 bp for An. sineroides; 260 bp for An. belenrae; and, 157 bp for An. pullus. Conclusion A multiplex PCR assay was developed to identify Anopheles spp. distributed in ROK. This method can be used to accurately identify Anopheles species that are difficult to identify morphologically to determine species distributions and malaria infection rates.

scholarly journals Identification through Culture and Molecular Methods of Campylobacter jejuni, Campylobacter coli and Campylobacter fetus in Surface Waters in Rasht

2019 ◽  
pp. 1-7
Author(s):  
Keyvan Roshanjo ◽  
Nematallah Jonaidi Jafari ◽  
Leila Asadpour ◽  
Reza Ranjbar ◽  
Davoud Afshar ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Campylobacter Jejuni ◽  
Surface Waters ◽  
Disease Transmission ◽  
Cross Sectional Study ◽  
Campylobacter Coli ◽  
Infectious Agents ◽  
Biochemical Tests ◽  
Cross Sectional ◽  
Campylobacter Fetus

Backgrounds: As zoonotic infectious agents, Campylobacter spp. are important factors causing gastroenteritis in humans. Surveys show that the three strains; Campylobacter jejuni, Campylobacter coli and Campylobacter fetus play a major role in human infections. Identification of these infectious agents is valuable for sanitary control of disease transmission through water resources. Objectives: The aim of this study was identification and molecular diagnosis of Campylobacter jejuni, Campylobacter coli and Campylobacter fetus in surface waters in Rasht. Materials and Methods: This cross-sectional study was conducted on 45 samples of surface water in Rasht collected according to water health guidelines. After culture and biochemical tests on collected samples, detection and identification of Campylobacter jejuni, Campylobacter coli and Campylobacter fetus was done using sequence-specific amplification by Multiplex PCR. The results were subjected to statistical analysis using SPSS software. Results: Out of 45 samples tested, 6 were positive in culture, four of which were identified as Campylobacter jejuni after biochemical tests. Using Multiplex PCR, 8 samples were positive, from which 3 were Campylobacter jejuni, 1 Campylobacter coli and 4 were positive for both Campylobacter jejuni and Campylobacter coli. All the samples did not yield C. fetus. Conclusions: Multiplex PCR is regarded a diagnostic method with higher sensitivity and specificity than compared to methods for Campylobacter. The prevalence of Campylobacter jejuni and Campylobacter coli in surface waters in Rasht is considerable. Therefore, public health measures for the control of these organisms are recommended.

Development of multiplex PCR for rapid detection of metallo-β-lactamase genes in clinical isolates of Acinetobacter baumannii

2020 ◽  
Author(s):  
Reza Ranjbar ◽  
Shahin Zayeri ◽  
Amir Mirzaie
Keyword(s):  
Acinetobacter Baumannii ◽  
Multiplex Pcr ◽  
Rapid Detection ◽  
Simultaneous Detection ◽  
Cross Reactivity ◽  
Clinical Isolates ◽  
Clinical Samples ◽  
Pcr Assay ◽  
Multiplex Pcr Assay

Background and Objectives: Acinetobacter baumannii has been known as a major pathogen causing nosocomial infec- tions. The aim of this study was to develop multiplex PCR for rapid and simultaneous detection of metallo-β-lactamase (MBL) genes in clinical isolates of A. baumannii. Materials and Methods: In this study, we used three sets of primers to amplify the MBL genes including bla        ,     bla   and bla   OXA-48 . The multiplex PCR assay was optimized for rapid and simultaneous detection of MBL genes in A. bau-   OXA-23   NDM   mannii strains recovered from clinical samples. Results: A. baumannii strains recovered from clinical samples were subjected to the study. The multiplex PCR produced 3   OXA-48   OXA-23   bands of 501 bp for bla        , 744 bp for bla observed in multiplex PCR.   OXA-48   and 623 bp for bla   NDM   genes. In addition to, no any cross-reactivity was   Conclusion: Based on obtained data, the multiplex PCR had a good specificity without any cross reactivity and it appears that the multiplex PCR is reliable assay for simultaneous detection of MBL genes in A. baumannii strains.  

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