Nested PCR with the PGMY09/11 and GP5+/6+ primer sets improves detection of HPV DNA in cervical samples

2004 ◽  
Vol 122 (1) ◽  
pp. 87-93 ◽  
Author(s):  
Andrea L. Fuessel Haws ◽  
Qin He ◽  
Peter L. Rady ◽  
Lifang Zhang ◽  
James Grady ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Hpv Dna ◽  
Primer Sets

Bedeutung von p16(INK4A) hinsichtlich des Nachweises von HPV-DNA und in der Stratifizierung der tumorspezifischen Mortalität bei 34 Patienten mit invasivem Peniskarzinom in Langzeit-Nachbeobachtung

2018 ◽  
Vol 50 (05) ◽  
pp. 491-501
Author(s):  
Johannes Bründl ◽  
Christina Neppl ◽  
Thomas Hommer ◽  
Wolfgang Otto ◽  
Johannes Breyer ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Prädiktiver Wert ◽  
Hpv Dna ◽  
P16ink4a Expression ◽  
Hpv Status ◽  
P16 Ink4a

Zusammenfassung Hintergrund Bei etwa jedem dritten Patienten mit einem penilen Plattenepithelkarzinom (PSCC) gilt die Infektion mit Humanen Papillomaviren (HPV) als entscheidender Faktor der Karzinogenese. Erste Daten zur optimierten Risikostratifizierung und Therapie anhand des HPV-Status existieren; somit hat der Nachweis einer HPV-Genese möglicherweise mehr als lediglich eine epidemiologische Dimension. Referenz zur Detektion von HPV ist die aufwendige Polymerase-Kettenreaktion (PCR). Unklar ist, ob Alternativen wie die In-situ-Hybridisierung (ISH) oder verschiedene Surrogatmarker (immunhistochemischer Nachweis einer p16INK4a-Expression, histologischer Subtyp, Tumorinvasionsfront, Koilozytose), zur Bestimmung des HPV-Status ausreichend sind. Methoden In dieser unizentrischen Studie an 34 Patienten mit PSCC wurden eine Multiplex nested-PCR zur Festlegung des HPV-Status und Genotypisierung der HPV-DNA sowie eine ISH durchgeführt. Diverse histologische Kriterien und p16INK4a wurden durch zentralen Review bestimmt. Der Einfluss verschiedener Kriterien auf die karzinomspezifische Mortalität (CSM) wurde mit Cox-Regressionsmodellen geprüft (FU: 92 Mon.). Ferner erfolgte der Vergleich der diskriminativen Qualitäten verschiedener Tumorinvasionsmuster (u. a. TNM-Klassifikation 7. vs. 8. Auflage) hinsichtlich der CSM-Prädiktion. Ergebnisse Gemäß PCR-Analyse wiesen 26 % der Patienten HR-HPV auf (n = 9). ISH und die untersuchten histologischen Kriterien zeigten eine unzureichende Vorhersagegenauigkeit des HPV-Status (p > 0,3). p16INK4a erreichte in den Testkriterien Sensitivität, Spezifität, positiv-prädiktiver und negativ-prädiktiver Wert sowie Gesamtübereinstimmung 66,7 %, 84 %, 60 %, 87,5 % bzw. 79,4 % (AUC: 0,753; p = 0,026). Keines der Untersuchungskriterien (inkl. PCR) korrelierte signifikant mit der CSM. Im Vergleich zur 7. wies die 8. Auflage der pT-Klassifikation eine bessere CSM-Prädiktion auf (C-Indizes 70,2 % vs. 72,9 %). In Addition zur Schwellkörper-Invasion besaß die Urethra-Infiltration keinen eigenständigen prädiktiven Wert. Eine durch uns vorgeschlagene Neu-Gruppierung der Schwellkörper-Invasionsmuster erzielte eine Diskriminationssteigerung bzgl. der CSM (C-Index 77,9 %). Schlussfolgerung p16INK4a erlaubt – im Gegensatz zur ISH und den untersuchten histologischen Kriterien – eine akzeptable Vorhersage des HPV-Status. Der Nachweis von HPV-DNA inkl. seiner Surrogatmarker hatte keinen Einfluss auf die CSM. Die 8. Auflage der pT-Klassifikation scheint prognostisch sinnvoll, da die urethrale Tumorinvasion keinen eigenständigen Einfluss auf die CSM aufweist.

scholarly journals Detection of human papillomavirus in cervical cell specimens by hybrid capture and PCR with different primers.

2006 ◽  
Vol 53 (3) ◽  
pp. 603-607 ◽  
Author(s):  
Slawa Szostek ◽  
Malgorzata Klimek ◽  
Barbara Zawilinska ◽  
Janusz Rys ◽  
Jolanta Kope ◽  
...  
Keyword(s):  
Human Papillomavirus ◽  
High Risk ◽  
Consensus Primer ◽  
Hybrid Capture ◽  
Hpv Dna ◽  
Cervical Smears ◽  
Capture Assay ◽  
Cervical Cell ◽  
Target Dna ◽  
Primer Sets

The purpose of this study was to compare hybrid capture assay with PCRs using different primers for the L1, E6-E7 regions for the detection of human papillomavirus (HPV) genome. One hundred twenty-five cervical smears with normal (n=42) and abnormal (n=83) cytology were investigated. Those at high-risk for HPV were studied by hybridization antibody capture assay and PCR with the pU-1M/pU-2R primers. Target DNA from the HPV L1 region was amplified by SPF10 primer set and home-PCR with MY09/MY11 primers. The presence of HPV DNA in cervical smears was detected by SPF10 (in 72% of cases), MY09/MY11 (58%), hybrid capture (55%) and pU-1M/pU-2R (39%). Results obtained with the SPF10 and MY09/MY11 consensus primer sets as well as hybrid capture and pU-1M/pU-2R specific for high-risk types differed significantly (chi2, P

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 Comparison of PCR versus Culture for Detection of Mycobacterium bovis after Experimental Inoculation of Various Matrices Held under Environmental Conditions for Extended Periods

2013 ◽  
Vol 79 (20) ◽  
pp. 6501-6506 ◽  
Author(s):  
Angela P. Adams ◽  
Steven R. Bolin ◽  
Amanda E. Fine ◽  
Carole A. Bolin ◽  
John B. Kaneene
Keyword(s):  
Mycobacterium Bovis ◽  
Nested Pcr ◽  
Weather Conditions ◽  
Content Type ◽  
Mycobacterial Culture ◽  
Monthly Sample ◽  
Sample Testing ◽  
Weekly Sample ◽  
Primer Sets ◽  
Initial Contamination

ABSTRACTThe purpose of this study was to compare the performance of a molecular detection technique (nested PCR) with that of mycobacterial culture in the detection ofMycobacterium bovisDNA in a set of 687 samples of experimentally inoculated environmental substrates (hay, soil, corn, water) exposed to natural weather conditions in Michigan. Four replicates of each substrate were used; half were autoclaved for sterilization, all were inoculated with 50,000 CFU ofM. bovisisolated from Michigan livestock, and all were placed in outdoor enclosures, with half under shade and the other half exposed to direct sunlight. Samples were tested for the presence ofM. bovisduring one 12-month period, with monthly sample testing and during three 12-week periods (winter, spring, summer) with weekly sample testing. Samples were subjected to mycobacterial culture for isolation ofM. bovisand a nested PCR with two primer sets targeting IS6110to detectM. bovisDNA. In 128 samples tested during the 12-month period,M. boviswas not detectable by culture after 2 months butM. bovisDNA was detectable by PCR for at least 7 months. Of the 559 samples tested during the 12-week periods, PCR detectedM. bovisDNA for up to 88 days in all of the sample types. There were no significant differences in the detection ofM. bovisbetween shade and sun samples or between sterile and unsterilized samples, regardless of the detection method (PCR or culture). For use in epidemiologic investigations, the PCR assay was more rapid than mycobacterial culture, was not hindered by contaminating organisms, and detectedM. bovisDNA in environment samples much longer after initial contamination than mycobacterial culture did.

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

Human Papillomavirus Genome based Detection and Typing: A Holistic Molecular Approach

2019 ◽  
Vol 19 (4) ◽  
pp. 237-246 ◽  
Author(s):  
Abhilasha Gautam ◽  
Mallikarjuna R. Gedda ◽  
Madhukar Rai ◽  
Shyam Sundar ◽  
Jaya Chakravarty
Keyword(s):  
Human Papillomavirus ◽  
High Risk ◽  
Genome Structure ◽  
Cost Effective ◽  
Detection Sensitivity ◽  
Screening Methods ◽  
Consensus Primer ◽  
Hpv Dna ◽  
Primer Sets ◽  
Consensus Primers

Human Papillomavirus (HPV) is a species specific double-stranded DNA virus infecting human cutaneous or mucosal tissues. The genome structure of HPV is extremely polymorphic hence making it difficult to discriminate between them. HPV exhibits numerous dissimilar types that can be subdivided into high-risk (HR), probably high-risk and low-risk (LR), causing numerous types of cancers and warts around the genital organs in humans. Several screening methods are performed in order to detect cytological abnormalities and presence or absence of HPV genome. Currently available commercial kits and methods are designed to detect only a few HR/LR-HPV types, which are expensive adding to the economic burden of the affected individual and are not freely available. These gaps could be minimized through Polymerase Chain reaction (PCR) method, which is a gold standard and a cost-effective technique for the detection of most HPV (both known and unknown) types by using specific consensus primers in minimal lab setup. In this context, numerous studies have validated the effectiveness of different sets of consensus primers in the screening of HPVs. Numerous consensus primers, such as E6, E6/E7, GP-E6/E7, MY09/11, GP5+/GP6+, SPF10, and PGMY09/11 have been developed to detect the presence of HPV DNA. In addition, HPV detection sensitivity could be achieved through consensus primer sets targeting specific ORF regions like L1 and E6, which may finally assist in better diagnosis of several unknown HR-HPVs. The present review, provides a summary of the available methods, kits and consensus primer sets for HPV genome based detection, their advantages and limitations along with future goals to be set for HPV detection.

scholarly journals RICERCA DI HPV-DNA MEDIANTE NESTED-PCR. NEI BRUSH LINGUALI DI SOGGETTI SANI

2004 ◽  
Vol 19 (2) ◽  
Author(s):  
R. Caldarelli-Stefano ◽  
S. Avezzu ◽  
V. Molina
Keyword(s):  
Nested Pcr ◽  
Hpv Dna

Verucciform Xanthoma of the Penis Not Associated With Human Papillomavirus Infection

2005 ◽  
Vol 129 (3) ◽  
pp. e62-e64 ◽  
Author(s):  
Çağatay Erşahin ◽  
Anna M. Szpaderska ◽  
Kimberly Foreman ◽  
Sherri Yong
Keyword(s):  
Human Papillomavirus ◽  
Hpv Infection ◽  
Nested Polymerase Chain Reaction ◽  
Hpv Dna ◽  
Negative Results ◽  
Papillomavirus Infection ◽  
Rare Lesion ◽  
Hpv Types ◽  
Polymerase Chain ◽  
Primer Sets

Abstract Verruciform xanthoma (VX) is a rare lesion with a predilection for oral mucosa. Only 16 cases of VX of the penis have been reported. Histologically, VX lesions in different locations are identical; however, the etiology is controversial. Previous studies have reported the presence of human papillomavirus (HPV) in VX of the skin. The purpose of this study was to determine whether HPV is a causative agent in this rare case of VX of the penis. Microscopically, the lesion demonstrated prominent verrucoid squamous hyperplasia with hyperkeratosis, parakeratosis, and acanthosis. Histiocytes, a hallmark of VX, were identified in the elongated dermal papillae. Nested polymerase chain reaction was performed on the DNA with the commonly used primer sets MY9/MY11 and GP5+/GP6+, which identify more than 40 HPV types. The results failed to identify HPV DNA in the sample, although HPV could be readily detected in genomic DNA extracted from paraffin-embedded condyloma acuminatum, a known HPV-associated lesion. Additionally, we tested a VX lesion of the palate for HPV DNA and obtained negative results. Our results indicate that VX can arise without HPV infection and suggest other possible origins may be involved.

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