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Article ; Online: Efficiency-corrected PCR quantification for identification of prevalence and load of respiratory disease-causing agents in feedlot cattle.

Barnewall, R J / Marsh, I B / Williams, T M / Cusack, Pmv / Sales, N / Galea, F / Szentirmay, A N / Quinn, J C

2022 Volume 100, Issue 11, Page(s) 539–549

Abstract: ... for BRD than non-BRD ailments. In addition, M. bovis was rarely detected at feedlot induction but was ...

Abstract	Bovine respiratory disease (BRD) is the most prevalent disease in feedlot cattle worldwide with Bovine alphaherpesvirus 1 (BoAHV1), Histophilus somni, Mannheimia haemolytica, Mycoplasma bovis, Pasteurella multocida and Trueperella pyogenes accepted to be common etiological agents associated with BRD. Although these agents are common in the upper and lower airways in clinical BRD cases, some also exist as normal flora suggesting their presence in the upper airways alone is not necessarily informative with respect to disease status or risk. To determine the relationship between presence, load and disease status, we investigated the relationship between load in the upper airways at induction and active BRD cases in feedlot cattle using efficiency-corrected PCR quantification. By this approach, we were able to accurately determine the prevalence and load of the key BRD agents in the upper respiratory tract showing that cattle in the hospital pen had a higher prevalence, and load, of these agents both singly and in combination compared to cattle sampled at feedlot induction. A combination of agents was the most accurate indicator of BRD risk with cattle with four or more agents detected in the upper airway more likely to be undergoing treatment for BRD than non-BRD ailments. In addition, M. bovis was rarely detected at feedlot induction but was identified at high prevalence in cattle in the hospital pen. These findings present a potential new technological approach for the investigation, analysis and identification of BRD-associated viral and bacterial agents for Australian feedlot systems as well as for BRD disease management and treatment.
MeSH term(s)	Cattle ; Animals ; Prevalence ; Australia/epidemiology ; Mannheimia haemolytica ; Cattle Diseases/epidemiology ; Cattle Diseases/microbiology ; Polymerase Chain Reaction/veterinary ; Bovine Respiratory Disease Complex/epidemiology ; Bovine Respiratory Disease Complex/microbiology
Language	English
Publishing date	2022-08-18
Publishing country	England
Document type	Journal Article
ZDB-ID	41542-x
ISSN	1751-0813 ; 0005-0423
ISSN (online)	1751-0813
ISSN	0005-0423
DOI	10.1111/avj.13200
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Article ; Online: Efficiency Correction Is Required for Accurate Quantitative PCR Analysis and Reporting.

Ruijter, Jan M / Barnewall, Rebecca J / Marsh, Ian B / Szentirmay, Andrew N / Quinn, Jane C / van Houdt, Robin / Gunst, Quinn D / van den Hoff, Maurice J B

Clinical chemistry

2021 Volume 67, Issue 6, Page(s) 829–842

Abstract: Background: Quantitative PCR (qPCR) aims to measure the DNA or RNA concentration in diagnostic and biological samples based on the quantification cycle (Cq) value observed in the amplification curves. Results of qPCR experiments are regularly calculated ...

Abstract	Background: Quantitative PCR (qPCR) aims to measure the DNA or RNA concentration in diagnostic and biological samples based on the quantification cycle (Cq) value observed in the amplification curves. Results of qPCR experiments are regularly calculated as if all assays are 100% efficient or reported as just Cq, ΔCq, or ΔΔCq values. Contents: When the reaction shows specific amplification, it should be deemed to be positive, regardless of the observed Cq. Because the Cq is highly dependent on amplification efficiency that can vary among targets and samples, accurate calculation of the target quantity and relative gene expression requires that the actual amplification efficiency be taken into account in the analysis and reports. PCR efficiency is frequently derived from standard curves, but this approach is affected by dilution errors and hampered by properties of the standard and the diluent. These factors affect accurate quantification of clinical and biological samples used in diagnostic applications and collected in challenging conditions. PCR efficiencies determined from individual amplification curves avoid these confounders. To obtain unbiased efficiency-corrected results, we recommend absolute quantification with a single undiluted calibrator with a known target concentration and efficiency values derived from the amplification curves of the calibrator and the unknown samples. Summary: For meaningful diagnostics or biological interpretation, the reported results of qPCR experiments should be efficiency corrected. To avoid ambiguity, the Minimal Information for Publications on Quantitative Real-Time PCR Experiments (MIQE) guidelines checklist should be extended to require the methods that were used (1) to determine the PCR efficiency and (2) to calculate the reported target quantity and relative gene expression value.
MeSH term(s)	Calibration ; Genetic Techniques ; Humans ; RNA ; Real-Time Polymerase Chain Reaction
Chemical Substances	RNA (63231-63-0)
Language	English
Publishing date	2021-04-23
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	80102-1
ISSN	1530-8561 ; 0009-9147
ISSN (online)	1530-8561
ISSN	0009-9147
DOI	10.1093/clinchem/hvab052
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Article ; Online: Efficiency‐corrected PCR quantification for identification of prevalence and load of respiratory disease‐causing agents in feedlot cattle

Barnewall, RJ / Marsh, IB / Williams, TM / Cusack, PMV / Sales, N / Galea, F / Szentirmay, AN / Quinn, JC

Australian Veterinary Journal. 2022 Nov., v. 100, no. 11 p.539-549

2022

Abstract: ... for BRD than non‐BRD ailments. In addition, M. bovis was rarely detected at feedlot induction but was ...

Abstract	Bovine respiratory disease (BRD) is the most prevalent disease in feedlot cattle worldwide with Bovine alphaherpesvirus 1 (BoAHV1), Histophilus somni, Mannheimia haemolytica, Mycoplasma bovis, Pasteurella multocida and Trueperella pyogenes accepted to be common etiological agents associated with BRD. Although these agents are common in the upper and lower airways in clinical BRD cases, some also exist as normal flora suggesting their presence in the upper airways alone is not necessarily informative with respect to disease status or risk. To determine the relationship between presence, load and disease status, we investigated the relationship between load in the upper airways at induction and active BRD cases in feedlot cattle using efficiency‐corrected PCR quantification. By this approach, we were able to accurately determine the prevalence and load of the key BRD agents in the upper respiratory tract showing that cattle in the hospital pen had a higher prevalence, and load, of these agents both singly and in combination compared to cattle sampled at feedlot induction. A combination of agents was the most accurate indicator of BRD risk with cattle with four or more agents detected in the upper airway more likely to be undergoing treatment for BRD than non‐BRD ailments. In addition, M. bovis was rarely detected at feedlot induction but was identified at high prevalence in cattle in the hospital pen. These findings present a potential new technological approach for the investigation, analysis and identification of BRD‐associated viral and bacterial agents for Australian feedlot systems as well as for BRD disease management and treatment.
Keywords	Arcanobacterium pyogenes ; Bovine alphaherpesvirus 1 ; Haemophilus somni ; Mannheimia haemolytica ; Mycoplasma bovis ; Pasteurella multocida ; bovine respiratory disease ; cattle ; disease control ; etiology ; feedlots ; flora ; hospitals ; respiratory system ; risk
Language	English
Dates of publication	2022-11
Size	p. 539-549.
Publishing place	Wiley Publishing Asia Pty Ltd
Document type	Article ; Online
Note	JOURNAL ARTICLE
ZDB-ID	41542-x
ISSN	1751-0813 ; 0005-0423
ISSN (online)	1751-0813
ISSN	0005-0423
DOI	10.1111/avj.13200
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Article: Luminescence probe studies of ionomers-II Steady-state measurements from sulphonated polyethylene and teflon membranes.

Szentirmay, M N / Prieto, N E / Martin, C R

Talanta

2006 Volume 32, Issue 8 Pt 2, Page(s) 745–749

Abstract: Luminescence probe studies of sulphonated Teflon and sulphonated polyethylene ionomer membranes are described. These studies have shown that the micropolarities of the ionic clusters within these ionomers are quite dynamic, varying with the nature of the ...

Abstract	Luminescence probe studies of sulphonated Teflon and sulphonated polyethylene ionomer membranes are described. These studies have shown that the micropolarities of the ionic clusters within these ionomers are quite dynamic, varying with the nature of the chain material and the membrane water content. These studies also suggest that polymer chain material intrudes into the ionic cluster phase.
Language	English
Publishing date	2006-01-23
Publishing country	Netherlands
Document type	Journal Article
ZDB-ID	1500969-5
ISSN	1873-3573 ; 0039-9140
ISSN (online)	1873-3573
ISSN	0039-9140
DOI	10.1016/0039-9140(85)80177-6
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Article ; Online: Spatial organization of RNA polymerase II transcription in the nucleus.

Szentirmay, M N / Sawadogo, M

Nucleic acids research

2000 Volume 28, Issue 10, Page(s) 2019–2025

Abstract: In eukaryotic cells, mRNA synthesis is carried out by large, multifunctional complexes that are also involved in coordinating transcription with other nuclear processes. This survey focuses on the distribution and structural arrangement of these ... ...

Abstract	In eukaryotic cells, mRNA synthesis is carried out by large, multifunctional complexes that are also involved in coordinating transcription with other nuclear processes. This survey focuses on the distribution and structural arrangement of these complexes within the nucleus, in relationship with the discrete positioning of particular chromosomal loci. To better understand the link between the spatial organization of the nucleus and the regulation of gene expression, it is necessary to combine information from biochemical studies with results from microscopic observations of preserved nuclear structures. Recent experimental approaches have made this possible. The subnuclear locations of specific chromosome loci, RNA transcripts, RNA polymerases, and transcription and pre-mRNA-processing factors can now be observed with computer-assisted microscopy and specific molecular probes. The results indicate that RNA polymerase II (RNAPII) transcription takes place at discrete sites scattered throughout the nucleoplasm, and that these sites are also the locations of pre-mRNA processing. Transcribing polymerases appear to be grouped into clusters at each transcription site. Cell cycle-dependent zones of transcription and processing factors have been identified, and certain subnuclear domains appear specialized for expression or silencing of particular genes. The arrangement of transcription in the nucleus is dynamic and depends on its transcriptional activity, with the RNAPII itself playing a central role in marshalling the large complexes involved in gene expression.
MeSH term(s)	Animals ; Cell Cycle ; Cell Nucleus/metabolism ; Cell Nucleus/ultrastructure ; Chromosomes/genetics ; Chromosomes/ultrastructure ; Mammals ; RNA Polymerase II/metabolism ; RNA, Messenger/genetics ; RNA, Messenger/metabolism ; Transcription, Genetic
Chemical Substances	RNA, Messenger ; RNA Polymerase II (EC 2.7.7.-)
Language	English
Publishing date	2000-05-15
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/28.10.2019
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Article ; Online: Correction: Determination of the phylogenetic origins of the Árpád Dynasty based on Y chromosome sequencing of Béla the Third.

European journal of human genetics : EJHG

2020 Volume 29, Issue 8, Page(s) 1317

Language	English
Publishing date	2020-12-29
Publishing country	England
Document type	Published Erratum
ZDB-ID	1141470-4
ISSN	1476-5438 ; 1018-4813
ISSN (online)	1476-5438
ISSN	1018-4813
DOI	10.1038/s41431-020-00795-5
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Article ; Online: Determination of the phylogenetic origins of the Árpád Dynasty based on Y chromosome sequencing of Béla the Third.

European journal of human genetics : EJHG

2020 Volume 29, Issue 1, Page(s) 164–172

Abstract: We set out to identify the origins of the Árpád Dynasty based on genome sequencing of DNA derived from the skeletal remains of Hungarian King Béla III (1172-1196) and eight additional individuals (six males, two females) originally interred at the Royal ... ...

Abstract	We set out to identify the origins of the Árpád Dynasty based on genome sequencing of DNA derived from the skeletal remains of Hungarian King Béla III (1172-1196) and eight additional individuals (six males, two females) originally interred at the Royal Basilica of Székesfehérvár. Y-chromosome analysis established that two individuals, Béla III and HU52 assign to haplogroups R-Z2125 whose distribution centres near South Central Asia with subsidiary expansions in the regions of modern Iran, the Volga Ural region and the Caucasus. Out of a cohort of 4340 individuals from these geographic areas, we acquired whole-genome data from 208 individuals derived for the R-Z2123 haplogroup. From these data we have established that the closest living kin of the Árpád Dynasty are R-SUR51 derived modern day Bashkirs predominantly from the Burzyansky and Abzelilovsky districts of Bashkortostan in the Russian Federation. Our analysis also reveals the existence of SNPs defining a novel Árpád Dynasty specific haplogroup R-ARP. Framed within the context of a high resolution R-Z2123 phylogeny, the ancestry of the first Hungarian royal dynasty traces to the region centering near Northern Afghanistan about 4500 years ago and identifies the Bashkirs as their closest kin, with a separation date between the two populations at the beginning of the first millennium CE.
MeSH term(s)	Chromosomes, Human, Y/genetics ; Famous Persons ; Female ; Human Migration ; Humans ; Hungary ; Male ; Pedigree ; Phylogeny ; Polymorphism, Single Nucleotide ; Sequence Analysis, DNA/methods
Language	English
Publishing date	2020-07-07
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1141470-4
ISSN	1476-5438 ; 1018-4813
ISSN (online)	1476-5438
ISSN	1018-4813
DOI	10.1038/s41431-020-0683-z
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Article: Sarkosyl block of transcription reinitiation by RNA polymerase II as visualized by the colliding polymerases reinitiation assay.

Szentirmay, M N / Sawadogo, M

Nucleic acids research

1994 Volume 22, Issue 24, Page(s) 5341–5346

Abstract: There are indications that different concentrations of Sarkosyl can block transcription initiation by RNA polymerase II in vitro at different functional steps [Hawley and Roeder (1985) J. Biol. Chem. 260, 8163-8172]. Consequently, this reagent could be a ...

Abstract	There are indications that different concentrations of Sarkosyl can block transcription initiation by RNA polymerase II in vitro at different functional steps [Hawley and Roeder (1985) J. Biol. Chem. 260, 8163-8172]. Consequently, this reagent could be a very useful tool for mechanistic studies. So far, however, evidence for the selectivity of Sarkosyl effects on RNA polymerase II transcription has been only indirect. To directly investigate the effect of Sarkosyl on transcription initiation and reinitiation by RNA polymerase II, we employed the reinitiation assay based on utilization of templates containing G-free cassettes (colliding polymerases reinitiation assay, or CoPRA). These experiments showed unambiguously that, under the appropriate conditions, Sarkosyl can be used to block transcription reinitiation by RNA polymerase II while allowing a first round of initiations from preassembled initiation complexes. This inhibition is not due to a disruption of the SII-dependent elongation of the reinitiated transcripts, and the levels of Sarkosyl that prevent transcription reinitiation coincide with the levels that block preinitiation complex assembly. However, Sarkosyl addition to transcription reactions reconstituted with partially purified transcription factors was found to have several undesirable side effects. The usefulness and limitations of the Sarkosyl-based and CoPRA assays for measurements of transcription reinitiation are discussed.
MeSH term(s)	RNA Polymerase II/antagonists & inhibitors ; RNA Polymerase II/metabolism ; RNA, Messenger/biosynthesis ; Sarcosine/analogs & derivatives ; Sarcosine/pharmacology ; Transcription Factors/isolation & purification ; Transcription Factors/metabolism ; Transcription, Genetic/drug effects
Chemical Substances	RNA, Messenger ; Transcription Factors ; sarkosyl (632GS99618) ; RNA Polymerase II (EC 2.7.7.-) ; Sarcosine (Z711V88R5F)
Language	English
Publishing date	1994-12-11
Publishing country	England
Document type	Comparative Study ; Journal Article ; Research Support, U.S. Gov't, P.H.S.
ZDB-ID	186809-3
ISSN	0305-1048 ; 0301-5610
ISSN	0305-1048 ; 0301-5610
DOI	10.1093/nar/22.24.5341
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Article: Synthesis of reinitiated transcripts by mammalian RNA polymerase II is controlled by elongation factor SII.

Szentirmay, M N / Sawadogo, M

The EMBO journal

1993 Volume 12, Issue 12, Page(s) 4677–4684

Abstract: Previous studies have revealed that the in vitro synthesis of reinitiated transcripts by RNA polymerase II requires an additional activity, designated reinitiation transcription factor (RTF), which is distinct from all of the general class II initiation ... ...

Abstract	Previous studies have revealed that the in vitro synthesis of reinitiated transcripts by RNA polymerase II requires an additional activity, designated reinitiation transcription factor (RTF), which is distinct from all of the general class II initiation factors. While further characterizing this activity, it was found that RTF displays properties indistinguishable from those of the RNA polymerase II elongation factor SII. In addition, Western blot analysis using SII-specific antibodies revealed that human SII is a major component in purified RTF preparations. The functional equivalence of the two proteins was established using recombinant SII, which proved fully capable of substituting for RTF in the reinitiation assay. In these reconstituted reactions, transcription complexes resulting from reinitiation events required SII to proceed through a 400 bp G-free cassette, while complexes resulting from the first round of initiations were SII-independent. Reinitiations can take place in the absence of SII; however, addition of the elongation factor is essential for full extension of the reinitiated transcripts. These results suggest that events taking place at the promoter (e.g. first-round initiations versus reinitiations) can create marked differences in the properties of RNA polymerase II elongation complexes.
MeSH term(s)	Animals ; Blotting, Western ; HeLa Cells ; Humans ; Mice ; RNA Polymerase II/metabolism ; Transcription Factors/metabolism ; Transcription Factors, General ; Transcription, Genetic ; Transcriptional Elongation Factors
Chemical Substances	Transcription Factors ; Transcription Factors, General ; Transcriptional Elongation Factors ; transcription factor S-II ; RNA Polymerase II (EC 2.7.7.-)
Language	English
Publishing date	1993-12
Publishing country	England
Document type	Journal Article ; Research Support, U.S. Gov't, P.H.S.
ZDB-ID	586044-1
ISSN	1460-2075 ; 0261-4189
ISSN (online)	1460-2075
ISSN	0261-4189
DOI	10.1002/j.1460-2075.1993.tb06156.x
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Article: Transcription factor requirement for multiple rounds of initiation by human RNA polymerase II.

Szentirmay, M N / Sawadogo, M

Proceedings of the National Academy of Sciences of the United States of America

1991 Volume 88, Issue 23, Page(s) 10691–10695

Abstract: We have investigated conditions that allow multiple rounds of transcription initiation from the adenovirus major late promoter in an in vitro system derived from HeLa cell nuclear extracts. Templates containing guanine-free cassettes provided a direct ... ...

Abstract	We have investigated conditions that allow multiple rounds of transcription initiation from the adenovirus major late promoter in an in vitro system derived from HeLa cell nuclear extracts. Templates containing guanine-free cassettes provided a direct assay for discriminating between reinitiated transcripts and transcripts generated by a first-round of transcription initiations. When reactions were reconstituted with the previously characterized class II transcription factors (TFIIA, TFIIB, TFIID, TFIIE/F), transcription by human RNA polymerase II from the adenovirus major late promoter was essentially restricted to a single round of initiations. Reinitiations at previously transcribed major late templates required an additional activity, designated reinitiation transcription factor (RTF). The RTF activity could be separated from the required transcription initiation factors. Semipurified human RTF also promoted transcription reinitiations at minimal promoters derived from the human c-myc, histone H4, and heat shock 70-kDa protein genes, indicating that the same reinitiation factor may be utilized by many, if not all, genes. The possible role of RTF in regulating the transcription rate of various class II genes is discussed.
MeSH term(s)	Cloning, Molecular ; Escherichia coli/genetics ; HeLa Cells ; Humans ; Kinetics ; Promoter Regions, Genetic ; RNA Polymerase II/metabolism ; Recombinant Proteins/isolation & purification ; Recombinant Proteins/metabolism ; Templates, Genetic ; Transcription Factor TFIID ; Transcription Factors/genetics ; Transcription Factors/isolation & purification ; Transcription Factors/metabolism ; Transcription, Genetic
Chemical Substances	Recombinant Proteins ; Transcription Factor TFIID ; Transcription Factors ; RNA Polymerase II (EC 2.7.7.-)
Language	English
Publishing date	1991-12-01
Publishing country	United States
Document type	Journal Article ; Research Support, U.S. Gov't, P.H.S.
ZDB-ID	209104-5
ISSN	1091-6490 ; 0027-8424
ISSN (online)	1091-6490
ISSN	0027-8424
DOI	10.1073/pnas.88.23.10691
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