Using recent findings by Tabor and Richardson, a novel thermostable DNA polymerase that generates sequence data comparable with T7 Sequenase DNA polymerase has been developed for improved base calling and longer reads.

Summary Introduction The 4Kscore Test is a prebiopsy blood test that incorporates four prostate protein biomarkers along with patient clinical information to determine a man's risk for high‐grade, aggressive (Gleason ≥7) prostate cancer. However, some men likely to benefit from the test may be seen in primary care settings where the digital rectal examination (DRE) information is not always obtained. In this study, we assessed the clinical validity of the 4Kscore Test when the DRE information was not included in the algorithm. Methods The Prospective 4Kscore Validation Study enrolled 1012 men scheduled for prostate biopsy across 26 urology practices in the United States. The 4Kscore was calculated for each patient with and without DRE information. The primary outcome was Gleason ≥7 prostate cancer on prostate biopsy. The contribution of DRE to the predictive accuracy of the test was evaluated by area under the receiver operating curve (AUC‐ROC), risk calibration and clinical consequences. Results High‐grade, aggressive prostate cancer was found in 231 (23%) of the 1012 patients. Both versions of the 4Kscore Test, with and without DRE, showed excellent discrimination (AUC=0.821 with DRE and AUC=0.818 without DRE input) and excellent calibration. No clinically significant difference was found between the two versions of the 4Kscore. Conclusions The 4Kscore Test algorithm, whether DRE findings are available or not, performs well in predicting a man's risk of high‐grade, aggressive prostate cancer. Patients who are suspected of having aggressive prostate cancer can safely have their risk better defined by 4Kscore even if a DRE has not been performed recently.

BackgroundWe have recently developed a simple, rapid, and relatively-cheap method for matrix-assisted laser-desorption and ionisation time-of-flight mass spectroscopy (MALDI-TOF MS) sample preparation that is applicable to plant material (in addition to microbial and insect material), and have used this to discriminate between closely-related Impatiens species and between regional biotypes of the invasive weed Impatiens glandulifera (commonly known as Himalayan balsam) using leaf samples. In the current paper, we have developed a complementary MALDI-TOF MS-based method for use with seeds. We have employed a combination of principal-component analysis and blind-tested comparison between reference-sample MALDI-TOF MS spectra and test-sample spectra to discriminate, on the basis of the acid-soluble seed-protein spectra generated by our method, between four regional biotypes of I. glandulifera from within the UK that differ in their susceptibility to the biological control agent Himalayan balsam rust (Puccinia komarovii var. glanduliferae).ResultsPeak-rich and highly-reproducible spectra were obtained and, in blind testing with test seeds collected in 2017 against reference seeds collected in 2017, we observed 100% identification accuracy in 12 blind tests. In blind testing with test seeds collected in 2016 against reference seeds collected in 2017, we observed 92% identification accuracy in 12 blind tests.ConclusionsMALDI-TOF MS analysis of seed material is able to discriminate between regional biotypes of I. glandulifera. MALDI-TOF MS therefore has the potential to improve the efficiency and efficacy of weed biological control using co-evolved natural enemies of invasive non-native plant species, through the matching of biological control agents with susceptible regional biotypes.Electronic supplementary materialThe online version of this article (10.1186/s13007-019-0412-1) contains supplementary material, which is available to authorized users.

Matrix-assisted laser-desorption and ionization time-of-flight mass spectrometry prepares proteins intact in the gas phase with predominantly a single positive charge. The times-of-flight of charged proteins along a tube held at high vacuum after acceleration in an electrical field are proportional to the square root of the mass-over-charge ratios for the proteins, thereby allowing a mass spectrum to be generated, which can then be used to characterize or identify a protein-containing sample. Several sample-preparation methods are currently available but not all of these are applicable to some forms of fungal biomass and few of these are well suited to the analysis of plant or insect material. We have therefore developed a simplified method that: lyses cells, selectively solubilizes basic proteins, dissolves matrix to a suitable concentration, generates spectra with good intensity and peak richness, costs no more (and generally less) than current methods, and is not constrained in terms of throughput by the availability of centrifuges. Using this method, and a reagent formulation comprising α-cyano-4-hydroxycinnamic acid matrix close to saturation in 60%–65% (v/v) acetonitrile in water containing 2.5% (v/v) trifluoroacetic acid, we have been able to differentiate between strains for a representative subset of aflatoxin-producing and aflatoxin-non-producing strains of Aspergillus fungi, to differentiate between Indian and Pakistani strains of Himalayan balsam rust, to differentiate between closely-related Crassula spp. and regional biotypes of Crassula helmsii , and to differentiate between rubbervine introduced into Australia and Brazil. We have also analyzed fall armyworm and stem-borer samples stored in 70% (v/v) ethanol and old dried insect specimens.

BackgroundMatrix-assisted laser-desorption and ionisation time-of-flight mass spectroscopy (MALDI-TOF MS) is a powerful tool for the characterisation and/or identification of protein-containing samples. Several MALDI-TOF MS sample-preparation methods are currently available but few of these are well suited to the analysis of plant material. We have recently developed a simple, rapid, and relatively-cheap method for MALDI-TOF MS that is applicable to plant material (in addition to microbial and insect material), and our aim in this study was to distinguish between closely-related plant species and/or between regional biotypes within an invasive weed species using this method with a view to optimising the selection of biological control agents that can be used for weed management.ResultsWe have employed a combination of principal-component analysis and closest-relatedness diagrams derived from MALDI-TOF MS spectral-comparison data to discriminate between the closely-related Impatiens spp. Impatiens noli-tangere, Impatiens parviflora, Impatiens scabrida, Impatiens balsamina, and two regional biotypes of the invasive weed Impatiens glandulifera. We have also developed a method for sample discrimination based upon comparison between blind-test MALDI-TOF MS spectra and reference-sample spectra. Using this latter method, we have been able to discriminate on the basis of the acid-soluble-protein mass spectra generated between four regional biotypes of I. glandulifera that differ in their susceptibility to the biological control agent Himalayan balsam rust (Puccinia komarovii var. glanduliferae) using mature leaf material. Using younger leaves, discrimination was not possible between these four regional biotypes.ConclusionsMALDI-TOF MS analysis is able to discriminate between closely-related plant species within the genus Impatiens and between regional biotypes of I. glandulifera. Because of this, MALDI-TOF MS holds great promise for improving weed biological control, a management technique which uses highly-specific co-evolved natural enemies for the control of an invasive non-native plant species, through the optimal matching of biological control agents with susceptible target species/regional biotypes.Electronic supplementary materialThe online version of this article (10.1186/s13007-018-0323-6) contains supplementary material, which is available to authorized users.