scholarly journals Mornitoring and Identification of Human Astrovirus from Groundwater in Korea Based on Highly Sensitive RT-nested PCR Primer Sets

2021 ◽  
Vol 27 (4) ◽  
pp. 255-263
Author(s):  
Siwon Lee ◽  
Kyung Seon Bae ◽  
Jihyun Park ◽  
Jin-Ho Kim ◽  
Jin-Young Lee ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Highly Sensitive ◽  
Human Astrovirus ◽  
Primer Sets ◽  
Pcr Primer

Multilocus Genetic Characterization of Two Ant Vectors (Group II “Dirty 22” Species) Known To Contaminate Food and Food Products and Spread Foodborne Pathogens†

2012 ◽  
Vol 75 (8) ◽  
pp. 1447-1452 ◽  
Author(s):  
IRSHAD M. SULAIMAN ◽  
MICKEY ANDERSON ◽  
DAVID H. OI ◽  
STEVEN SIMPSON ◽  
KHALIL KERDAHI
Keyword(s):  
Foodborne Pathogens ◽  
Nested Pcr ◽  
Genetic Characterization ◽  
Food Products ◽  
Species Group ◽  
Pest Species ◽  
Group I ◽  
Group Ii ◽  
Primer Sets ◽  
Pcr Primer

The U.S. Food and Drug Administration utilizes the presence of filth and extraneous materials as one of the criteria for implementing regulatory actions and assessing adulteration of food products of public health importance. Twenty-two prevalent pest species (also known as the “Dirty 22” species) have been considered by this agency as possible vehicles for the spread of foodborne diseases, and the presence of these species is considered an indicator of unsanitary conditions in food processing and storage facilities. In a previous study, we further categorized the Dirty 22 species into four groups: group I includes four cockroach species, group II includes two ant species, group III includes 12 fly species, and group IV includes four rodent species. Here, we describe the development of three nested PCR primer sets and multilocus genetic characterization by amplifying the small subunit rRNA, elongation factor 1-alpha, and wingless (WNT-1) genes of group II Dirty 22 ant species Monomorium pharaonis and Solenopsis molesta. These novel group II Dirty 22 species-specific nested PCR primer sets can be used when the specimens cannot be identified using conventional microscopic methods. These newly developed assays will provide correct identification of group II Dirty 22 ant species, and the information can be used in the control of foodborne pathogens.

scholarly journals Deep Sequencing of a Dimethylsulfoniopropionate-Degrading Gene (dmdA) by Using PCR Primer Pairs Designed on the Basis of Marine Metagenomic Data

2009 ◽  
Vol 76 (2) ◽  
pp. 609-617 ◽  
Author(s):  
Vanessa A. Varaljay ◽  
Erinn C. Howard ◽  
Shulei Sun ◽  
Mary Ann Moran
Keyword(s):  
Sequence Data ◽  
Gene Clusters ◽  
Amino Acid Identity ◽  
Metagenomic Data ◽  
Data Sets ◽  
Free Living ◽  
Metagenomic Sequence ◽  
Primer Sets ◽  
Design And Testing ◽  
Pcr Primer

ABSTRACT In silico design and testing of environmental primer pairs with metagenomic data are beneficial for capturing a greater proportion of the natural sequence heterogeneity in microbial functional genes, as well as for understanding limitations of existing primer sets that were designed from more restricted sequence data. PCR primer pairs targeting 10 environmental clades and subclades of the dimethylsulfoniopropionate (DMSP) demethylase protein, DmdA, were designed using an iterative bioinformatic approach that took advantage of thousands of dmdA sequences captured in marine metagenomic data sets. Using the bioinformatically optimized primers, dmdA genes were amplified from composite free-living coastal bacterioplankton DNA (from 38 samples over 5 years and two locations) and sequenced using 454 technology. An average of 6,400 amplicons per primer pair represented more than 700 clusters of environmental dmdA sequences across all primers, with clusters defined conservatively at >90% nucleotide sequence identity (∼95% amino acid identity). Degenerate and inosine-based primers did not perform better than specific primer pairs in determining dmdA richness and sometimes captured a lower degree of richness of sequences from the same DNA sample. A comparison of dmdA sequences in free-living versus particle-associated bacteria in southeastern U.S. coastal waters showed that sequence richness in some dmdA subgroups differed significantly between size fractions, though most gene clusters were shared (52 to 91%) and most sequences were affiliated with the shared clusters (∼90%). The availability of metagenomic sequence data has significantly enhanced the design of quantitative PCR primer pairs for this key functional gene, providing robust access to the capabilities and activities of DMSP demethylating bacteria in situ.

scholarly journals A highly sensitive modified nested PCR to enhance case detection in leishmaniasis

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Bhagya Deepachandi ◽  
Sudath Weerasinghe ◽  
Preethi Soysa ◽  
Nadira Karunaweera ◽  
Yamuna Siriwardana
Keyword(s):  
Nested Pcr ◽  
Case Detection ◽  
Highly Sensitive

scholarly journals Prevalence of the Helicobacter pylori babA2 Gene in Children Mainly Depends on the PCR Primer Set Used

2020 ◽  
Vol 2020 ◽  
pp. 1-9
Author(s):  
Anja Šterbenc ◽  
Maja M. Lunar ◽  
Matjaž Homan ◽  
Boštjan Luzar ◽  
Nina Zidar ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Helicobacter Pylori ◽  
Diagnostic Tool ◽  
Clinical Relevance ◽  
Detection Methods ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Primer Sets ◽  
H Pylori ◽  
Pcr Primer

Various polymerase chain reaction- (PCR-) based methods with varying positivity rates were designed to detect the Helicobacter pylori babA2 gene. To compare different primer sets, babA2 prevalence was determined in 279 H. pylori-positive pediatric samples using the 832 bp, 139 bp, and 271 bp PCR primer sets, resulting in 34.0%, 51.3%, and 79.6% prevalence of the babA2 gene, respectively. The babA2 status determined using the 832 bp and 139 bp PCR primer sets significantly correlated with bacterial density and activity of inflammation, whereas no such correlations were found using the 271 bp PCR primer set. The 139 and 832 bp PCR primer sets concordantly detected the babA2 gene in 93 cases; however, in comparison to the 832 bp PCR primer set, the 139 bp PCR primer set detected additional 50 babA2 cases, whereas only two 832 bp positive cases were missed. The 271 bp PCR primer set missed 32 babA2 cases that were 832 bp and/or 139 bp PCR positive, but tested solely positive in 109 cases. Interestingly, cloning of a subset of 271 bp PCR positive samples revealed amplification of the babA/B gene chimera. Hence, in our opinion, the 271 bp PCR protocol is not a reliable diagnostic tool for detecting the babA2 gene in children. Our results reaffirm previous observations that the use of certain babA2 PCR primer sets can significantly impact estimation of the prevalence and clinical relevance of the H. pylori babA2 gene in children, suggesting babA2 detection methods should be carefully selected.

Highly sensitive nested PCR and rapid immunochromatographic detection of Babesia bovis and Babesia bigemina infection in a cattle herd with acute clinical and fatal cases in Argentina

2019 ◽  
Vol 67 (S2) ◽  
pp. 159-164 ◽  
Author(s):  
Sabrina Ganzinelli ◽  
Daniel Benitez ◽  
Sambuu Gantuya ◽  
Azirwan Guswanto ◽  
Monica Florin‐Christensen ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Babesia Bovis ◽  
Cattle Herd ◽  
Babesia Bigemina ◽  
Highly Sensitive

Rapid identification and differentiation of fungal DNA in dermatological specimens by LightCycler PCR

2004 ◽  
Vol 53 (12) ◽  
pp. 1207-1214 ◽  
Author(s):  
Ralf Gutzmer ◽  
Susanne Mommert ◽  
Uta Küttler ◽  
Thomas Werfel ◽  
Alexander Kapp
Keyword(s):  
Skin Diseases ◽  
Rapid Identification ◽  
Diagnostic Information ◽  
Clinical Samples ◽  
Melting Points ◽  
Number Of Patients ◽  
Highly Sensitive ◽  
Fungal Dna ◽  
Pcr Method ◽  
Primer Sets

The aim was to develop a LightCycler PCR method for the rapid detection and differentiation of fungal DNA in dermatological specimens such as skin scales and skin swabs. LightCycler PCR assays were established for seven primer sets specific for fungal DNA. For each primer set LightCycler melting points were defined by amplification of DNA from 21 fungi and sensitivity was determined by amplification of serial dilutions of fungal DNA. A protocol was established that allows detection and differentiation of mould and yeast DNA with one highly sensitive PCR reaction by assessment of LightCycler melting points. Two subsequent LightCycler PCR reactions and one RFLP reaction allowed the differentiation of dermatophytes and non-dermatophyte moulds and the subclassification of yeasts. Analysis of clinical samples from 38 patients with fungal skin diseases provided conclusive new diagnostic information in 9/38 cases (23.7 %) by this PCR protocol that was not equally provided by direct microscopy and mycological culture. Thus the LightCycler PCR protocol established here represents a rapid diagnostic tool that aids in the diagnosis of fungal skin disease in a substantial number of patients.

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