Detection of Cryptosporidium parvum and Giardia lamblia in Water Supply by Quantitative Real-Time PCR

2012 ◽  
Vol 518-523 ◽  
pp. 3707-3711
Author(s):  
Peng Peng ◽  
Rui Bao Jia ◽  
Yu Mei Liu ◽  
Li Li
Keyword(s):  
Water Supply ◽  
Real Time ◽  
Cryptosporidium Parvum ◽  
Real Time Pcr ◽  
Giardia Lamblia ◽  
Pcr Primers ◽  
Quantitative Real Time Pcr ◽  
Rt Pcr ◽  
High Recovery ◽  
Pcr Method

Cryptosporidium parvum and Giardia lamblia are common pathogenic protozoa in water, which pose high risk to drinking water supply. In the present study, detection of C. parvum and G. lamblia was performed by quantitative real-time PCR (RT-PCR). Pairs of PCR primers were evaluated for the detection specificity to pathogenic C. parvum and G. lamblia. The recovery of the RT-PCR detection procedure was examined and high recovery rates (i.e., more than 45％ for C. parvum and more than 50％ for G. lamblia ) were achieved. The RT-PCR method was used to detect C. parvum and G. lamblia in a secondary water supply. The results indicated the potential application of the quantitative RT- PCR method in detection of C. parvum and G. lamblia in water supply.

scholarly journals Real-Time PCR Assay for Detection and Enumeration of Dekkera bruxellensis in Wine

2003 ◽  
Vol 69 (12) ◽  
pp. 7430-7434 ◽  
Author(s):  
Trevor G. Phister ◽  
David A. Mills
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Pcr Primers ◽  
Rrna Gene ◽  
Dekkera Bruxellensis ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Spoilage Yeast ◽  
Pcr Method ◽  
26S Rrna

ABSTRACT Traditional methods to detect the spoilage yeast Dekkera bruxellensis from wine involve lengthy enrichments. To overcome this difficulty, we developed a quantitative real-time PCR method to directly detect and enumerate D. bruxellensis in wine. Specific PCR primers to D. bruxellensis were designed to the 26S rRNA gene, and nontarget yeast and bacteria common to the winery environment were not amplified. The assay was linear over a range of cell concentrations (6 log units) and could detect as little as 1 cell per ml in wine. The addition of large amounts of nontarget yeasts did not impact the efficiency of the assay. This method will be helpful to identify possible routes of D. bruxellensis infection in winery environments. Moreover, the time involved in performing the assay (3 h) should enable winemakers to more quickly make wine processing decisions in order to reduce the threat of spoilage by D. bruxellensis.

A Quantitative Real-Time PCR Method Using an X-Linked Gene for Sex Typing in Pigs

2012 ◽  
Vol 54 (2) ◽  
pp. 493-496 ◽  
Author(s):  
Maria Ballester ◽  
Anna Castelló ◽  
Yuliaxis Ramayo-Caldas ◽  
Josep M. Folch
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Quantitative Real Time Pcr ◽  
Sex Typing ◽  
Linked Gene ◽  
Pcr Method

Quantitative Detection of Campylobacter jejuni on Fresh Chicken Carcasses by Real-Time PCR

2007 ◽  
Vol 70 (6) ◽  
pp. 1373-1378 ◽  
Author(s):  
ANNA-CLARA RÖNNER ◽  
HANS LINDMARK
Keyword(s):  
Detection Limit ◽  
Real Time ◽  
Dna Extraction ◽  
Campylobacter Jejuni ◽  
Real Time Pcr ◽  
Close Correlation ◽  
Quantitative Detection ◽  
Quantitative Real Time Pcr ◽  
Direct Plating ◽  
Pcr Method

Campylobacter jejuni infection is a significant cause of foodborne gastroenteritis worldwide. Consumption and handling of poultry products is believed to be the primary risk factor for campylobacteriosis. Risk assessments require quantitative data, and C. jejuni is enumerated usually by direct plating, which sometimes allows growth of non-Campylobacter bacteria. The objective of the present study was to develop a quantitative real-time PCR method (q-PCR) for enumerating C. jejuni in chicken rinse without a culturing step. The procedure to obtain the template for the PCR assay involved (i) filtration of 10 ml of chicken rinse, (ii) centrifugation of the sample, and (iii) total DNA extraction from the pellet obtained using a commercial DNA extraction kit. The detection limit of the method was comparable to that for plating 100 μl of chicken rinse on modified charcoal cefoperazone deoxycholate agar, and the detection limit could be further improved 10-fold by concentrating the DNA eluate by ethanol precipitation. A close correlation for spiked chicken rinse was obtained for the results of the quantitative real-time PCR method and direct plating (r = 0.99). The coefficient of correlation for the methods was 0.87 when samples from chicken carcasses on the slaughter line were analyzed, whereas a lower correlation (r = 0.76) was obtained when samples from retail carcasses were analyzed. Greater variation in the proportion of dead and/or viable but not culturable Campylobacter types in the retail samples may explain the decreased correlation between the methods. Overall, the new method is simple and fast and the results obtained are closely correlated with those for direct plating for samples containing a low proportion of dead Campylobacter cells.

Improved specificity for Giardia lamblia cyst quantification in wastewater by development of a real-time PCR method

2004 ◽  
Vol 57 (1) ◽  
pp. 41-53 ◽  
Author(s):  
Isabelle Bertrand ◽  
Christophe Gantzer ◽  
Thierry Chesnot ◽  
Janine Schwartzbrod
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Giardia Lamblia ◽  
Pcr Method

Quantification of Cryptosporidium parvum in anaerobic digesters treating manure by (reverse-transcription) quantitative real-time PCR, infectivity and excystation tests

2006 ◽  
Vol 53 (8) ◽  
pp. 195-202 ◽  
Author(s):  
G. Garcés ◽  
M. Effenberger ◽  
M. Najdrowski ◽  
C. Wackwitz ◽  
A. Gronauer ◽  
...  
Keyword(s):  
Anaerobic Digestion ◽  
Real Time ◽  
Cryptosporidium Parvum ◽  
Reverse Transcription ◽  
Real Time Pcr ◽  
Combined Treatment ◽  
Quantitative Real Time Pcr ◽  
Anaerobic Digesters ◽  
Hsp70 Mrna ◽  
Cryptosporidium Parvum Oocysts

The survival of Cryptosporidium parvum oocysts in anaerobic digesters treating manure was investigated for mesophilic, thermophilic, and a combined treatment (mesophilic–thermophilic–mesophilic) under different retention times of oocysts in the reactors. C. parvum DNA was extracted with an optimised protocol, and its amount determined by quantitative real-time PCR (qPCR). Results indicated noteworthy differences in DNA content after the different treatments. DNA was not degraded during the process. However, excystation and infectivity tests showed a reduction of viable oocyst numbers of ≥2 and ≥5 log units after the thermophilic treatment in two different experiments. Thus qPCR-targeting DNA can overestimate the number of oocysts that survive and remain viable after anaerobic digestion. However, targeting DNA is suitable to indicate the presence or absence of oocysts. Reverse transcription qPCR (RT-qPCR) targeting C. parvum hsp70 mRNA successfully indicated the presence of viable fraction of oocysts.

Detecting Cryptosporidium parvum and Giardia lamblia by coagulation concentration and real-time PCR quantification

2012 ◽  
Vol 7 (1) ◽  
pp. 49-54 ◽  
Author(s):  
Huining Zhang ◽  
Xiaohu Zhang ◽  
Shuting Zhang ◽  
Bo Wei ◽  
Qipei Jiang ◽  
...  
Keyword(s):  
Real Time ◽  
Cryptosporidium Parvum ◽  
Real Time Pcr ◽  
Giardia Lamblia

scholarly journals A quantitative real-time PCR method for absolute telomere length

BioTechniques ◽  
2008 ◽  
Vol 44 (6) ◽  
pp. 807-809 ◽  
Author(s):  
Nathan J. O'Callaghan ◽  
Varinderpal S. Dhillon ◽  
Philip Thomas ◽  
Michael Fenech
Keyword(s):  
Telomere Length ◽  
Real Time ◽  
Real Time Pcr ◽  
Quantitative Real Time Pcr ◽  
Pcr Method
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