The Experts below are selected from a list of 57 Experts worldwide ranked by ideXlab platform
Peter Gill - One of the best experts on this subject based on the ideXlab platform.
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Genotyping and interpretation of STR-DNA: Low-template, mixtures and database matches—Twenty years of research and development
Forensic science international. Genetics, 2015Co-Authors: Peter Gill, Hinda Haned, Øyvind Bleka, Oskar Hansson, Guro Dørum, Thore EgelandAbstract:The introduction of Short Tandem Repeat (STR) DNA was a revolution within a revolution that transformed forensic DNA profiling into a tool that could be used, for the first time, to create National DNA databases. This transformation would not have been possible without the concurrent development of fluorescent automated sequencers, combined with the ability to Multiplex several loci together. Use of the polymerase chain reaction (PCR) increased the sensitivity of the method to enable the analysis of a handful of cells. The first Multiplexes were simple: 'the quad', introduced by the defunct UK Forensic Science Service (FSS) in 1994, rapidly followed by a more discriminating 'six-plex' (Second Generation Multiplex) in 1995 that was used to create the world's first national DNA database. The success of the database rapidly outgrew the functionality of the original system - by the year 2000 a new Multiplex of ten-loci was introduced to reduce the chance of adventitious matches. The technology was adopted world-wide, albeit with different loci. The political requirement to introduce pan-European databases encouraged standardisation - the development of European Standard Set (ESS) of markers comprising twelve-loci is the latest iteration. Although development has been impressive, the methods used to interpret evidence have lagged behind. For example, the theory to interpret complex DNA profiles (low-level mixtures), had been developed fifteen years ago, but only in the past year or so, are the concepts starting to be widely adopted. A plethora of different models (some commercial and others non-commercial) have appeared. This has led to a confusing 'debate' about the 'best' to use. The different models available are described along with their advantages and disadvantages. A section discusses the development of national DNA databases, along with details of an associated controversy to estimate the strength of evidence of matches. Current methodology is limited to searches of complete profiles - another example where the interpretation of matches has not kept pace with development of theory. STRs have also transformed the area of Disaster Victim Identification (DVI) which frequently requires kinship analysis. However, genotyping efficiency is complicated by complex, degraded DNA profiles. Finally, there is now a detailed understanding of the causes of stochastic effects that cause DNA profiles to exhibit the phenomena of drop-out and drop-in, along with artefacts such as stutters. The phenomena discussed include: heterozygote balance; stutter; degradation; the effect of decreasing quantities of DNA; the dilution effect.
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a comparison of adjustment methods to test the robustness of an str dna database comprised of 24 european populations
Forensic Science International, 2003Co-Authors: Peter Gill, John Buckleton, Lindsey A Foreman, Christopher M Triggs, Heather AllenAbstract:An aim of the European Network of Forensic Science Institutes (ENFSI) is to produce a DNA database of Second Generation Multiplex (SGM) STR profiles that is representative of the resident cosmopolitan populations. To achieve this, data were collected from 24 different populations. All of the data were combined to form one database of 5700 profiles from which allele proportions were calculated. The robustness of this combined European database was tested by estimating parameter d for every DNA profile, where d=log10(Pmc/PmE) Pmc is the match probability of the profile calculated from its cognate database and PmE is the match probability of the combined European database. Overall there was a small tendency for Pmc>PmE primarily because of sampling bias. This bias was removed by the simple expediency of applying an adjustment factor to the calculation of PmE. These were selected from the Balding size bias correction, the Balding and Nichols Fst correction, a minimum allele proportion (between 0.01 and 0.02), an upper bound of a 95% confidence interval (CI) and a lower bound on the genotype match probability. It was demonstrated that a single European database is a feasible proposition. A combination of different adjustment methods can be used to ensure that the result is conservative relative to the cognate database, and their effect measured by parameter d.
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Validation of the applied biosystems Prism™ 377 automated sequencer for forensic short tandem repeat analysis
Electrophoresis, 1996Co-Authors: Rachael R. E. Frazier, Emma S. Millican, S. Watson, N. Oldroyd, R Sparkes, Karen M. Taylor, Sasikala Panchal, Linda Bark, C. P. Kimpton, Peter GillAbstract:The Applied Biosystems (ABI) Prism 377 DNA sequencer has been evaluated in an attempt to increase the throughput of samples for short tandem repeat (STR) analysis, in both forensic casework and the UK National Criminal Intelligence DNA Database. The gel system assessed consisted of 0.2 mm, 4% acrylamide 6 M urea gels, with a well-to-read distance of 36 cm. Gels were run at a constant voltage of 3 kV and constant temperature of 51 degrees C. The run time of our Second Generation Multiplex (SGM) STR system was achieved in less than 2 h. Rigorous validation has been performed on the instrument hardware and software. Complete resolution of 1 base differences was obtained, up to and beyond 350 bases; sizing precision across gels was more than 2-fold higher than the 373A and the sensitivity was increased by one third.
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the validation of a 7 locus Multiplex str test for use in forensic casework ii artefacts casework studies and success rates
International Journal of Legal Medicine, 1996Co-Authors: R Sparkes, N. Oldroyd, C. P. Kimpton, S Gilbard, P Carne, J Andersen, D Thomas, A Urquhart, Peter GillAbstract:: PCR-based DNA typing of biological evidence is now widely used in forensic analyses due to the obvious advantages of enhanced sensitivity, the ability to distinguish discrete alleles and efficacy with degraded samples. A Multiplex short tandem repeat (STR) system has been previously developed which successfully co-amplifies six STR loci HUMTH01, D21S11, D18S51, D8S1179, HUMVWF31/A and HUMFIBRA (FGA) in conjunction with the X-Y homologous gene Amelogenin. This is known as the Second Generation Multiplex system (SGM). Detection of the PCR products is undertaken on ABD 373A or 377 automated sequencers using denaturing polyacrylamide gels coupled with fluorescent-based technology. We have evaluated this system for routine forensic use and demonstrated that the technique is robust and reproducible under conditions consistent with those encountered in a forensic environment. A total of 132 stains from simulated and actual casework were analysed, together with relevant control areas and reference samples. The success rate was high with 76% of stains giving full profiles; we were also able to successfully detect and interpret mixtures. No mistyping was observed. A detailed examination of each of these profiles has assisted in the development of guidelines for casework interpretation. Although artefacts, stutter peaks and undenatured DNA were occasionally observed, these did not interfere with the accuracy of interpretation. In addition 38 samples, previously examined using the quadruplex system, were analysed with the SGM to enable a direct comparison to be made between the systems. The performance of the system with poor quality samples demonstrated its use as a rapid and powerful technique for individual identification.
Peter M. Prodinger - One of the best experts on this subject based on the ideXlab platform.
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Improved pre-operative diagnostic accuracy for low-grade prosthetic joint infections using Second-Generation Multiplex Polymerase chain reaction on joint fluid aspirate
International Orthopaedics, 2020Co-Authors: Christian Suren, Susanne Feihl, Sabrina Cabric, Ingo J. Banke, Bernhard Haller, Andrej Trampuz, Rüdiger Eisenhart-rothe, Peter M. ProdingerAbstract:Background A major obstacle for the treatment of prosthetic joint infection (PJI) is the identification of the underlying causative organism. While the diagnostic criteria ruling PJI in or out have become ever more accurate, the detection of the causative pathogen(s) still relies mostly on conventional and time-consuming microbial culture. The aim of this study was to evaluate the diagnostic potential of a Second-Generation Multiplex PCR assay ( Unyvero ITI G2, Curetis AG, Holzgerlingen, Germany) used on synovial fluid specimens. Our hypothesis was that the method would yield a higher diagnostic accuracy in the pre-operative workup than synovial fluid culture. Thus, a more precise classification of septic and aseptic prosthesis failure could be achieved before revision surgery. Methods Prospectively collected frozen joint fluid specimens from 26 patients undergoing arthroplasty revision surgery of the hip or knee were tested as per the manufacturer’s protocol. Sensitivities, specificities, positive and negative predictive values as well as positive and negative likelihood ratios with corresponding confidence intervals were estimated using the statistical software R . A combination of the serum C-reactive protein (CRP) level, leukocyte count, erythrocyte sedimentation rate, joint fluid culture, tissue biopsy culture, and tissue biopsy histology served as the gold standard. Results Of the 26 patients included in the study, 15 were infected and 11 were aseptic. Conventional joint fluid culture showed a sensitivity of 0.67 and a specificity of 0.91. Joint fluid Multiplex PCR yielded a sensitivity of 0.8 and a specificity of 1.0. Conclusions Using the Second-Generation Unyvero ITI cartridge on joint fluid aspirate for the detection of prosthetic joint infection, we were able to achieve a higher diagnostic accuracy than with conventional culture. We conclude that to improve pathogen detection before revision surgery, this method represents a valuable and practicable tool.
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High Accuracy of Second Generation Unyvero ITI Automated Multiplex PCR Analysis of Joint Fluid Aspirate in Low-Grade Prosthetic Hip and Knee Joint Infections
Orthopaedic Journal of Sports Medicine, 2020Co-Authors: Christian Suren, Susanne Feihl, Sabrina Cabric, Ingo J. Banke, Bernhard Haller, Andrej Trampuz, Rüdiger Von Eisenhart-rothe, Peter M. ProdingerAbstract:Aims and Objectives: A major obstacle for the treatment of prosthetic joint infections (PJIs) is the identification of the organism causing it. While the diagnostic criteria ruling PJIs in or out have become ever more accurate, detecting the causative pathogen(s) still relies mostly on conventional culture. The aim of this study was to evaluate the diagnostic potential of a Second Generation Multiplex PCR assay (Unyvero ITI G2, Curetis AG, Holzgerlingen, Germany) used on synovial fluid specimens. Materials and Methods: Prospectively collected frozen joint fluid specimens from 26 patients undergoing arthroplasty revision surgery of the hip or knee for any reason were tested as per the manufacturer’s protocol. The sensitivity and specificity were calculated using the Chi-squared-test and a combination of the serum CRP level, leukocyte count, erythrocyte sedimentation rate, joint fluid culture, tissue biopsy culture, and tissue biopsy histology served as gold standard. Results: Of the 26 cases included in the study, 15 were infected and 11 aseptic. Conventional joint fluid culture showed a sensitivity 0.67 and a specificity of 0.91. Joint fluid Multiplex PCR yielded a sensitivity of 0.8 and a specificity of 1.0. Conclusion: This is the first study to assess the Second Generation Unyvero ITI cartridge on joint fluid aspirate for the detection of prosthetic joint infection. In our setting, the method achieved a higher diagnostic accuracy than conventional culture. We conclude that, to improve pathogen detection before revision surgery, this method represents a valuable tool.
Christian Suren - One of the best experts on this subject based on the ideXlab platform.
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Improved pre-operative diagnostic accuracy for low-grade prosthetic joint infections using Second-Generation Multiplex Polymerase chain reaction on joint fluid aspirate
International Orthopaedics, 2020Co-Authors: Christian Suren, Susanne Feihl, Sabrina Cabric, Ingo J. Banke, Bernhard Haller, Andrej Trampuz, Rüdiger Eisenhart-rothe, Peter M. ProdingerAbstract:Background A major obstacle for the treatment of prosthetic joint infection (PJI) is the identification of the underlying causative organism. While the diagnostic criteria ruling PJI in or out have become ever more accurate, the detection of the causative pathogen(s) still relies mostly on conventional and time-consuming microbial culture. The aim of this study was to evaluate the diagnostic potential of a Second-Generation Multiplex PCR assay ( Unyvero ITI G2, Curetis AG, Holzgerlingen, Germany) used on synovial fluid specimens. Our hypothesis was that the method would yield a higher diagnostic accuracy in the pre-operative workup than synovial fluid culture. Thus, a more precise classification of septic and aseptic prosthesis failure could be achieved before revision surgery. Methods Prospectively collected frozen joint fluid specimens from 26 patients undergoing arthroplasty revision surgery of the hip or knee were tested as per the manufacturer’s protocol. Sensitivities, specificities, positive and negative predictive values as well as positive and negative likelihood ratios with corresponding confidence intervals were estimated using the statistical software R . A combination of the serum C-reactive protein (CRP) level, leukocyte count, erythrocyte sedimentation rate, joint fluid culture, tissue biopsy culture, and tissue biopsy histology served as the gold standard. Results Of the 26 patients included in the study, 15 were infected and 11 were aseptic. Conventional joint fluid culture showed a sensitivity of 0.67 and a specificity of 0.91. Joint fluid Multiplex PCR yielded a sensitivity of 0.8 and a specificity of 1.0. Conclusions Using the Second-Generation Unyvero ITI cartridge on joint fluid aspirate for the detection of prosthetic joint infection, we were able to achieve a higher diagnostic accuracy than with conventional culture. We conclude that to improve pathogen detection before revision surgery, this method represents a valuable and practicable tool.
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High Accuracy of Second Generation Unyvero ITI Automated Multiplex PCR Analysis of Joint Fluid Aspirate in Low-Grade Prosthetic Hip and Knee Joint Infections
Orthopaedic Journal of Sports Medicine, 2020Co-Authors: Christian Suren, Susanne Feihl, Sabrina Cabric, Ingo J. Banke, Bernhard Haller, Andrej Trampuz, Rüdiger Von Eisenhart-rothe, Peter M. ProdingerAbstract:Aims and Objectives: A major obstacle for the treatment of prosthetic joint infections (PJIs) is the identification of the organism causing it. While the diagnostic criteria ruling PJIs in or out have become ever more accurate, detecting the causative pathogen(s) still relies mostly on conventional culture. The aim of this study was to evaluate the diagnostic potential of a Second Generation Multiplex PCR assay (Unyvero ITI G2, Curetis AG, Holzgerlingen, Germany) used on synovial fluid specimens. Materials and Methods: Prospectively collected frozen joint fluid specimens from 26 patients undergoing arthroplasty revision surgery of the hip or knee for any reason were tested as per the manufacturer’s protocol. The sensitivity and specificity were calculated using the Chi-squared-test and a combination of the serum CRP level, leukocyte count, erythrocyte sedimentation rate, joint fluid culture, tissue biopsy culture, and tissue biopsy histology served as gold standard. Results: Of the 26 cases included in the study, 15 were infected and 11 aseptic. Conventional joint fluid culture showed a sensitivity 0.67 and a specificity of 0.91. Joint fluid Multiplex PCR yielded a sensitivity of 0.8 and a specificity of 1.0. Conclusion: This is the first study to assess the Second Generation Unyvero ITI cartridge on joint fluid aspirate for the detection of prosthetic joint infection. In our setting, the method achieved a higher diagnostic accuracy than conventional culture. We conclude that, to improve pathogen detection before revision surgery, this method represents a valuable tool.
Paul Debenham - One of the best experts on this subject based on the ideXlab platform.
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Analysis of disputed single-parent/child and sibling relationships using 16 STR loci
International Journal of Legal Medicine, 2001Co-Authors: J. A. Thomson, V Pilotti, K. L. Ayres, M. N. Barrett, J. I. H. Walker, Paul DebenhamAbstract:This study describes the validation of short tandem repeat (STR) systems for the resolution of cases of disputed parentage where only a single parent is available for testing or where the claimed relationship of both parents is in doubt and also cases where sibship must be tested. Three separate Multiplex systems the Second Generation Multiplex, Powerplex 1.2 and FFFL have been employed, giving a total of 16 STR loci. Both empirical and theoretical approaches to the validation have been adopted. Appropriate equations have been derived to calculate likelihood ratios for different relationships, incorporating a correction for subpopulation effects. An F _ST point estimate of 1% has been applied throughout. Empirically, 101 cases of alleged father, alleged mother and child where analysed using six SLP systems and also using the three Multiplex STR systems. Of the 202 relationships tested, 197 were independently resolved by both systems, providing either clear evidence of non-parentage or strong support for the relationship.
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validation of short tandem repeat analysis for the investigation of cases of disputed paternity
Forensic Science International, 1999Co-Authors: James A. Thomson, V Pilotti, Karen L Ayres, P Stevens, Paul DebenhamAbstract:Abstract This study details validation of two separate Multiplex STR systems for use in paternity investigations. These are the Second Generation Multiplex (SGM) developed by the UK Forensic Science Service and the PowerPlex™1 Multiplex commercially available from Promega Inc. (Madison, WI, USA). These Multiplexes contain 12 different STR systems (two are duplicated in the two systems). Population databases from Caucasian, Asian and Afro-Caribbean populations have been compiled for all loci. In all but two of the 36 STR/ethnic group combinations, no evidence was obtained to indicate inconsistency with Hardy-Weinberg (HW) proportions. Empirical and theoretical approaches have been taken to validate these systems for paternity testing. Samples from 121 cases of disputed paternity were analysed using established Single Locus Probe (SLP) tests currently in use, and also using the two Multiplex STR systems. Results of all three test systems were compared and no non-conformities in the conclusions were observed, although four examples of apparent germ line mutations in the STR systems were identified. The data was analysed to give information on expected paternity indices and exclusion rates for these STR systems. The 12 systems combined comprise a highly discriminating test suitable for paternity testing. 99.96% of non-fathers are excluded from paternity on two or more STR systems. Where no exclusion is found, Paternity Index (PI) values of >10 000 are expected in >96% of cases.
R Sparkes - One of the best experts on this subject based on the ideXlab platform.
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The validation of a 7-locus Multiplex STIR test for use in forensic casework
International Journal of Legal Medicine, 1996Co-Authors: R Sparkes, N. Oldroyd, S Gilbard, P Carne, D Thomas, A Urquhart, C. Kimpton, J. Anderson, P. GillAbstract:PCR-based DNA typing of biological evidence is now widely used in forensic analyses due to the obvious advantages of enhanced sensitivity, the ability to distinguish discrete alleles and efficacy with degraded samples. A Multiplex short tandem repeat (STR) system has been previously developed which successfully co-amplifies six STR loci HUMTH01, D21S11, D18S51, D8S1179, HUMVWF31/A and HUMFIBRA (FGA) in conjunction with the X-Y homologous gene Amelogenin. This is known as the Second Generation Multiplex system (SGM). Detection of the PCR products is undertaken on ABD 373A or 377 automated sequencers using denaturing polyacrylamide gels coupled with fluorescent-based technology. We have evaluated this system for routine forensic use and demonstrated that the technique is robust and reproducible under conditions consistent with those encountered in a forensic environment. A total of 132 stains from simulated and actual casework were analysed, together with relevant control areas and reference samples. The success rate was high with 76% of stains giving full profiles; we were also able to successfully detect and interpret mixtures. No mistyping was observed. A detailed examination of each of these profiles has assisted in the development of guidelines for casework interpretation. Although artefacts, stutter peaks and undenatured DNA were occasionally observed, these did not interfere with the accuracy of interpretation. In addition 38 samples, previously examined using the quadruplex system, were analysed with the SGM to enable a direct comparison to be made between the systems. The performance of the system with poor quality samples demonstrated its use as a rapid and powerful technique for individual identification.
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Validation of the applied biosystems Prism™ 377 automated sequencer for forensic short tandem repeat analysis
Electrophoresis, 1996Co-Authors: Rachael R. E. Frazier, Emma S. Millican, S. Watson, N. Oldroyd, R Sparkes, Karen M. Taylor, Sasikala Panchal, Linda Bark, C. P. Kimpton, Peter GillAbstract:The Applied Biosystems (ABI) Prism 377 DNA sequencer has been evaluated in an attempt to increase the throughput of samples for short tandem repeat (STR) analysis, in both forensic casework and the UK National Criminal Intelligence DNA Database. The gel system assessed consisted of 0.2 mm, 4% acrylamide 6 M urea gels, with a well-to-read distance of 36 cm. Gels were run at a constant voltage of 3 kV and constant temperature of 51 degrees C. The run time of our Second Generation Multiplex (SGM) STR system was achieved in less than 2 h. Rigorous validation has been performed on the instrument hardware and software. Complete resolution of 1 base differences was obtained, up to and beyond 350 bases; sizing precision across gels was more than 2-fold higher than the 373A and the sensitivity was increased by one third.
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the validation of a 7 locus Multiplex str test for use in forensic casework ii artefacts casework studies and success rates
International Journal of Legal Medicine, 1996Co-Authors: R Sparkes, N. Oldroyd, C. P. Kimpton, S Gilbard, P Carne, J Andersen, D Thomas, A Urquhart, Peter GillAbstract:: PCR-based DNA typing of biological evidence is now widely used in forensic analyses due to the obvious advantages of enhanced sensitivity, the ability to distinguish discrete alleles and efficacy with degraded samples. A Multiplex short tandem repeat (STR) system has been previously developed which successfully co-amplifies six STR loci HUMTH01, D21S11, D18S51, D8S1179, HUMVWF31/A and HUMFIBRA (FGA) in conjunction with the X-Y homologous gene Amelogenin. This is known as the Second Generation Multiplex system (SGM). Detection of the PCR products is undertaken on ABD 373A or 377 automated sequencers using denaturing polyacrylamide gels coupled with fluorescent-based technology. We have evaluated this system for routine forensic use and demonstrated that the technique is robust and reproducible under conditions consistent with those encountered in a forensic environment. A total of 132 stains from simulated and actual casework were analysed, together with relevant control areas and reference samples. The success rate was high with 76% of stains giving full profiles; we were also able to successfully detect and interpret mixtures. No mistyping was observed. A detailed examination of each of these profiles has assisted in the development of guidelines for casework interpretation. Although artefacts, stutter peaks and undenatured DNA were occasionally observed, these did not interfere with the accuracy of interpretation. In addition 38 samples, previously examined using the quadruplex system, were analysed with the SGM to enable a direct comparison to be made between the systems. The performance of the system with poor quality samples demonstrated its use as a rapid and powerful technique for individual identification.
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the validation of a 7 locus Multiplex str test for use in forensic casework i mixtures ageing degradation and species studies
International Journal of Legal Medicine, 1996Co-Authors: R Sparkes, S. Watson, N. Oldroyd, C. P. Kimpton, Tim Clayton, L Barnett, J Arnold, C Thompson, R Hale, J ChapmanAbstract:PCR-based DNA typing of biological evidence is now widely used in forensic analyses due to the obvious advantages of enhanced sensitivity, the ability to distinguish discrete alleles and efficacy with degraded samples. A Multiplex short tandem repeat (STR) system has been previously developed which successfully co-amplifies six STR loci HUMTH01, D21S11, D18S51, D8S1179, HUMVWF31/A and HUMFIBRA (FGA) in conjunction with the X-Y homologous gene Amelogenin. This is known as the Second Generation Multiplex system (SGM). Detection of the PCR products is undertaken on ABD 373A or 377 automated sequencers using denaturing polyacrylamide gels coupled with fluorescent-based technology. We have evaluated this system for routine forensic use and demonstrated that the technique is robust and reproducible under conditions consistent with those encountered in a forensic environment. A total of 132 stains from simulated and actual casework were analysed, together with relevant control areas and reference samples. The success rate was high with 76% of stains giving full profiles; we were also able to successfully detect and interpret mixtures. No mistyping was observed. A detailed examination of each of these profiles has assisted in the development of guidelines for casework interpretation. Although artefacts, stutter peaks and undenatured DNA were occasionally observed, these did not interfere with the accuracy of interpretation. In addition 38 samples, previously examined using the quadruplex system, were analysed with the SGM to enable a direct comparison to be made between the systems. The performance of the system with poor quality samples demonstrated its use as a rapid and powerful technique for individual identification.