Identification of equine mares as reservoir hosts for pathogenic species of Leptospira.

Equine leptospirosis can result in abortion, stillbirth, neonatal death, placentitis, and uveitis. Horses can also act as subclinical reservoir hosts of infection, which are characterized as asymptomatic carriers that persistently excrete leptospires and transmit disease. In this study, PCR and culture were used to assess urinary shedding of pathogenic Leptospira from 37 asymptomatic mares. Three asymptomatic mares, designated as H2, H8, and H9, were PCR-positive for lipL32, a gene specific for pathogenic species of Leptospira. One asymptomatic mare, H9, was culture-positive, and the recovered isolate was classified as L. kirschneri serogroup Australis serovar Rushan. DNA capture and enrichment of Leptospira genomic DNA from PCR-positive, culture-negative samples determined that asymptomatic mare H8 was also shedding L. kirschneri serogroup Australis, whereas asymptomatic mare H2 was shedding L. interrogans serogroup Icterohaemorrhagiae. Sera from all asymptomatic mares were tested by the microscopic agglutination test (MAT) and 35 of 37 (94.6%) were seropositive with titers ranging from 1:100 to 1:3200. In contrast to asymptomatic mares, mare H44 presented with acute spontaneous abortion and a serum MAT titer of 1:102,400 to L. interrogans serogroup Pomona serovar Pomona. Comparison of L. kirschneri serogroup Australis strain H9 with that of L. interrogans serogroup Pomona strain H44 in the hamster model of leptospirosis corroborated differences in virulence of strains. Since lipopolysaccharide (LPS) is a protective antigen in bacterin vaccines, the LPS of strain H9 (associated with subclinical carriage) was compared with strain H44 (associated with spontaneous abortion). This revealed different LPS profiles and immunoreactivity with reference antisera. It is essential to know what species and serovars of Leptospira are circulating in equine populations to design efficacious vaccines and diagnostic tests. Our results demonstrate that horses in the US can act as reservoir hosts of leptospirosis and shed diverse pathogenic Leptospira species via urine. This report also details the detection of L. kirschneri serogroup Australis serovar Rushan, a species and serotype of Leptospira, not previously reported in the US.

Development of a nano-size off-grid energy system using renewables and IoT technologies at the Meteoria visitor center: A Finnish case study.

The increased utilization of non-renewable energy during the last century has influenced the climate, with increased carbon dioxide emissions and elevated temperature as a result. Thus, the need to develop and demonstrate new sustainable solutions regarding both energy supply and consumption, but also energy system optimization, is obvious. This case study presents the nano-size off-grid energy system at the Meteoria visitor center in Ostrobothnia, Finland, and the real-time measuring techniques that have been installed to follow up the energy production and consumption. The Meteoria consists of several buildings, which are open to the public from April to October. The case site is operated by energy derived from wind power, solar power, and a diesel generator (as a backup), with batteries for energy storage. The Internet of Things (IoT) has been retrofitted to the existing energy system to enable energy measurements and follow various electrical parameters in real-time. In addition, a graphical visualization platform open to the public has been developed. In this study, the completeness of data sampling and the IoT system was checked, and the results show high availability of data. Furthermore, various errors/limitations regarding the IoT system were identified. The energy supply/demand at the Meteoria in 2021 was monitored and the challenges regarding the existing energy system in a cold climate zone are discussed as well as the potential role of the Meteoria to function as a living lab.

Concurrent colonization of rodent kidneys with multiple species and serogroups of pathogenic Leptospira.

Rodents are important reservoir hosts of pathogenic leptospires in the US Virgin Islands. Our previous work determined that trapped rodents were colonized with Leptospira borgpetersenii serogroup Ballum (n = 48) and/or Leptospira kirschneri serogroup Icterohaemorrhagiae (n = 3). In addition, nine rodents appeared to be colonized with a mixed population comprising more than one species/serogroup. The aim of this study was to validate this finding by characterizing clonal isolates derived from cultures of mixed species. Cultures of presumptive mixed species (designated LR1, LR5, LR37, LR57, LR60, LR61, LR68, LR70, and LR72) were propagated in different media including Hornsby-Alt-Nally (HAN) media, incubated at both 29℃ and 37℃, and T80/40/LH incubated at 29℃. Polyclonal reference antisera specific for serogroup Ballum and Icterohaemorrhagiae were used to enrich for different serogroups followed by subculture on agar plates. Individual colonies were then selected for genotyping and serotyping. Of the nine cultures of mixed species/serogroups, a single clonal isolate was separated in five of them: L. borgpetersenii serogroup Ballum in LR1, LR5, and LR37, and L. kirschneri serogroup Icterohaemorrhagiae in LR60 and LR72. In four of the cultures with mixed species (LR57, LR61, LR68, and LR70), clonal isolates of both L. borgpetersenii serogroup Ballum and L. kirschneri serogroup Icterohaemorrhagiae were recovered. Our results definitively establish that rodents can be colonized with more than one species/serogroup of Leptospira concurrently. The identification and characterization of multiple species/serogroups of Leptospira from individual reservoir hosts of infection are essential to understand the epidemiology and transmission of disease to both human and domestic animal populations.IMPORTANCEPathogenic Leptospira, the causative agent of human and animal leptospirosis, comprise a diverse genus of species/serogroups which are inherently difficult to isolate from mammalian hosts due to fastidious growth requirements. Molecular evidence has indicated that reservoir hosts of Leptospira may shed multiple species concurrently. However, evidence of this phenomena by culture has been lacking. Culture is definitive and is essential for comprehensive characterization of recovered isolates by high-resolution genome sequencing and serotyping. In this work, a protocol using recently developed novel media formulations, in conjunction with reference antisera, was developed and validated to demonstrate the recovery of multiple species/serogroups of pathogenic Leptospira from the same host. The identification and characterization of multiple species/serogroups of Leptospira from individual reservoir hosts of infection are essential to understand the epidemiology and transmission of disease to both human and domestic animal populations.

Reliability of whole mount radical prostatectomy histopathology as the ground truth for artificial intelligence assisted prostate imaging.

The development of artificial intelligence-based imaging techniques for prostate cancer (PCa) detection and diagnosis requires a reliable ground truth, which is generally based on histopathology from radical prostatectomy specimens. This study proposes a comprehensive protocol for the annotation of prostatectomy pathology slides. To evaluate the reliability of the protocol, interobserver variability was assessed between five pathologists, who annotated ten radical prostatectomy specimens consisting of 74 whole mount pathology slides. Interobserver variability was assessed for both the localization and grading of PCa. The results indicate excellent overall agreement on the localization of PCa (Gleason pattern ≥ 3) and clinically significant PCa (Gleason pattern ≥ 4), with Dice similarity coefficients (DSC) of 0.91 and 0.88, respectively. On a per-slide level, agreement for primary and secondary Gleason pattern was almost perfect and substantial, with Fleiss Kappa of .819 (95% CI .659-.980) and .726 (95% CI .573-.878), respectively. Agreement on International Society of Urological Pathology Grade Group was evaluated for the index lesions and showed agreement in 70% of cases, with a mean DSC of 0.92 for all index lesions. These findings show that a standardized protocol for prostatectomy pathology annotation provides reliable data on PCa localization and grading, with relatively high levels of interobserver agreement. More complicated tissue characterization, such as the presence of cribriform growth and intraductal carcinoma, remains a source of interobserver variability and should be treated with care when used in ground truth datasets.

Leptospira borgpetersenii serovar Hardjo and Leptospira santarosai serogroup Pyrogenes isolated from bovine dairy herds in Puerto Rico.

Leptospirosis is one of the most common zoonotic diseases in the world and endemic in the Caribbean Islands. Bovine leptospirosis is an important reproductive disease. Globally, cattle are recognized as a reservoir host for L. borgpetersenii serovar Hardjo, which is transmitted via urine, semen, and uterine discharges, and can result in abortion and poor reproductive performance. The dairy industry in Puerto Rico comprises up to 25% of agriculture-related income and is historically the most financially important agricultural commodity on the island. In this study, we report the isolation of two different pathogenic Leptospira species, from two different serogroups, from urine samples collected from dairy cows in Puerto Rico: L. borgpetersenii serogroup Sejroe serovar Hardjo and L. santarosai serogroup Pyrogenes. Recovered isolates were classified using whole-genome sequencing, serotyping with reference antisera and monoclonal antibodies, and immunoblotting. These results demonstrate that dairy herds in Puerto Rico can be concurrently infected with more than one species and serovar of Leptospira, and that bacterin vaccines and serologic diagnostics should account for this when applying intervention and diagnostic strategies.

Bovine Leptospirosis Due to Persistent Renal Carriage of Leptospira borgpetersenii Serovar Tarassovi.

Leptospirosis is a global zoonotic disease that causes significant morbidity and mortality in human and animal populations. Leptospira interrogans is a leading cause of human disease, and L. borgpetersenii is a leading cause of animal disease. Cattle are reservoir hosts of L. borgpetersenii serovar Hardjo, which is transmitted via urine, semen, and uterine discharges resulting in abortion and poor reproductive performance. Bovine bacterin vaccines can only protect against those serovars included in vaccine formulations and typically include serovar Hardjo among others. Genotyping and serotyping represent two different and unique methods for classifying leptospires that do not always correlate well; comprehensive characterization using either method requires recovery of isolates from infected animals. In this study, we report for the first time, isolation of L. borgpetersenii serovar Tarassovi from the urine of a dairy cow in the U.S. The classification of the isolate, designated strain MN900, was confirmed by whole-genome sequencing, serotyping with reference antisera and monoclonal antibodies, Matrix Assisted Laser Desorption/Ionization (MALDI), and immunoblotting with reference antisera. Strain MN900 was excreted in urine samples for 18 weeks even as the cow was seronegative for serovar Tarassovi. Strain MN900 has an unusual morphology since it is not as motile as other leptospires and lacks hooked ends. Serovar Tarassovi is not included in U.S. bacterin vaccines. These results demonstrate the importance of culture and concomitant genotyping and serotyping to accurately classify leptospires, and as required to design efficacious vaccine and diagnostic strategies to not only limit animal disease but reduce zoonotic risk.

Leptospira sanjuanensis sp. nov., a pathogenic species of the genus Leptospira isolated from soil in Puerto Rico

Two spirochetes (designated strains LGVF01 and LGVF02T) were isolated from soil samples in San Juan, Puerto Rico in HAN media after selection using a combination of ELISA, agar plating, and colony screening by Fluorescent Antibody Testing (FAT) and PCR for lipL32 and secY. Isolates were helix-shaped, aerobic, fast-growing, and highly motile. Genome sequence analysis indicated that both strains should be classified as members of a novel species within the pathogenic (P1) clade of the genus Leptospira. The average nucleotide identity between the two strains was 99.2 %, but below 93.2 % when compared to any previously described leptospiral species. Serotyping of strain LGVF02T indicates that it does not belong within any serogroup of Leptospira suggesting it also represents a new serovar. Collectively, strains LGVF01 and LGVF02T represent a new species of pathogenic leptospires for which the name Leptospira sanjuanensis sp. nov. is proposed. The type strain is LGVF02T (=NVSL-LGVF02T=KIT0302T).

Leptospira sanjuanensis sp. nov., a pathogenic species of the genus Leptospira isolated from soil in Puerto Rico

Two spirochetes (designated strains LGVF01 and LGVF02T) were isolated from soil samples in San Juan, Puerto Rico in HAN media after selection using a combination of ELISA, agar plating, and colony screening by Fluorescent Antibody Testing (FAT) and PCR for lipL32 and secY. Isolates were helix-shaped, aerobic, fast-growing, and highly motile. Genome sequence analysis indicated that both strains should be classified as members of a novel species within the pathogenic (P1) clade of the genus Leptospira. The average nucleotide identity between the two strains was 99.2 %, but below 93.2 % when compared to any previously described leptospiral species. Serotyping of strain LGVF02T indicates that it does not belong within any serogroup of Leptospira suggesting it also represents a new serovar. Collectively, strains LGVF01 and LGVF02T represent a new species of pathogenic leptospires for which the name Leptospira sanjuanensis sp. nov. is proposed. The type strain is LGVF02T (=NVSL-LGVF02T=KIT0302T).

Safe Upper Limits of Serum Cobalt and Chromium Levels for a Metal-on-Metal Total Hip Bearing: A 10-Year Follow-Up Study.

BACKGROUND: Long-term survival of metal-on-metal (MoM) prostheses and the development of adverse reaction to metal debris (ARMD) around these bearings are still unclear. Serum levels of cobalt (Co) and chromium (Cr) are used as a screening tool to anticipate failure in MoM bearings and detect ARMD. METHODS: One hundred sixty primary large head MoM prostheses were followed up for 10 years. To estimate the revision risk, the cumulative incidence function (CIF) was used. Subdistribution hazard modeling was used to investigate the associations between cumulative incidence of revision for ARMD and Co levels, Cr levels, gender, age, head size, and cup inclination. Furthermore, the safe upper limits (SULs) for Co and Cr were determined. RESULTS: Univariate analyses showed an increased risk in revision for ARMD in females (subdistribution hazard ratio [sdHR] 3.43, 95% confidence interval [CI] 1.01-11.7, P = .049) and cup inclination angles over 45° (sdHR 4.70, 95% CI 1.63-13.58, P = .004). In addition, a higher last measured Co level (sdHR 1.05, 95% CI 1.03-1.07, P < .001) and last measured Cr level (sdHR 1.21, 95% CI 1.14-1.29, P < .001) were associated with a higher probability of revision for ARMD. We determined our bearing-specific SULs at 4.1 parts per billion (ppb) and 4.2 ppb for Co and Cr, respectively. CONCLUSION: Guidelines regarding follow-up and surveillance should include a complete clinical assessment with bearing-specific SULs of serum metal ion levels. For the M2a-Magnum MoM bearing we advise an SUL for Co and Cr levels of 4.1 and 4.2 ppb, respectively.

Searching and Finding the Hidden Treasure: A Retrospective Analysis of Rickettsial Disease Among Dutch International Travelers.

BACKGROUND: Rickettsial disease (RD) is a prevalent and underestimated cause of febrile illness worldwide, especially in the absence of an inoculation eschar. We attempted to quantify this underestimation at our clinic, by investigating past cases of febrile illness in travelers who had tested negative for leptospirosis, a disease that can initially present similarly to non-eschar RD, and which we routinely consider when other important causes of unspecified febrile illness have tested negative. METHODS: We performed a retrospective analysis in febrile returned travelers from Asia, Africa, or the Americas between 2010 and 2017, who had tested negative for leptospirosis. Serologic immunofluorescence assays were performed for Orientia tsutsugamushi (scrub typhus), typhus group, and spotted fever group RD. We performed a medical records review of all patients who tested positive. In case of a fitting medical history, cases were deemed either confirmed (based on convalescent serology) or suspected (based on single serology). RESULTS: Among 97 patients, convalescent serology was available in 16 (16.5%) patients, and a single serology in 81 (83.5%) patients. RD was the likely diagnosis in 8 of 16 (50.0%) patients with convalescent serology, and in 8 of 81 (9.9%) with single serology. Of the 16 confirmed/suspected cases, 11 (69%) had been missed and 7 (44%) had not received adequate empiric antibiotic therapy. CONCLUSIONS: This study shows that non-eschar RD is an important and poorly recognized cause of illness in travelers, even in a specialized travel clinic. A lower threshold to test and treat for RD is warranted in returning travelers with febrile illness.

Genetic diversity of pathogenic leptospires from wild, domestic and captive host species in Portugal.

Leptospirosis is a neglected zoonotic disease of worldwide distribution with a significant veterinary and public health impact. It is caused by pathogenic bacteria of the genus Leptospira. The availability of effective tools to accurately identify and type leptospires is of utmost importance for the diagnosis of the disease and for assessing its epidemiology. Several multi-locus sequence typing (MLST) approaches were described for the typing of worldwide isolates of Leptospira but an extensive agreement towards the adoption of a unique consensus scheme for this agent is still lacking. Most genotyped strains originate from Asian and South American countries, with a minority originating from Europe (being most countries represented only by one or a few isolates). The knowledge of the diversity of circulating leptospires is the key to understanding the disease transmission and its zoonotic implications. In this study, we revisited the taxonomy of several isolates of pathogenic Leptospira obtained from domestic, wild and captive animals in Portugal, between 1990 and 2012. A selection of these isolates was genotyped using two previously published MLST schemes. A total of seven distinct sequence types (STs) were detected among the Portuguese isolates with two STs representing L. borgpetersenii (ST149 and ST152), two STs representing L. kirschneri (ST117 and ST100) and three STs representing L. interrogans (ST17, ST24 and ST140). Global widespread (and maybe more virulent) Leptospira genotypes seem to circulate in Portugal, particularly the L. interrogans ST17 isolates which are associated with several outbreaks of leptospirosis among humans and animals in different regions of the world. This study contributes to the enrichment of the global MLST databases with a new set of allele and sequence type information also providing novel data on circulating Leptospira serovars in Portugal.

Introduction of primary screening using high-risk HPV DNA detection in the Dutch cervical cancer screening programme: a population-based cohort study.

BACKGROUND: In January 2017, the Dutch cervical cancer screening programme transitioned from cytomorphological to primary high-risk HPV (hrHPV) DNA screening, including the introduction of self-sampling, for women aged between 30 and 60 years. The Netherlands was the first country to switch to hrHPV screening at the national level. We investigated the health impact of this transition by comparing performance indicators from the new hrHPV-based programme with the previous cytology-based programme. METHODS: We obtained data from the Dutch nationwide network and registry of histo- and cytopathology (PALGA) for 454,573 women eligible for screening in 2017 who participated in the hrHPV-based programme between 1 January 2017 and 30 June 2018 (maximum follow-up of almost 21 months) and for 483,146 women eligible for screening in 2015 who participated in the cytology-based programme between 1 January 2015 and 31 March 2016 (maximum follow-up of 40 months). We compared indicators of participation (participation rate), referral (screen positivity; referral rate) and detection (cervical intraepithelial neoplasia (CIN) detection; number of referrals per detected CIN lesion). RESULTS: Participation in the hrHPV-based programme was significantly lower than that in the cytology-based programme (61% vs 64%). Screen positivity and direct referral rates were significantly higher in the hrHPV-based programme (positivity rate: 5% vs 9%; referral rate: 1% vs 3%). CIN2+ detection increased from 11 to 14 per 1000 women screened. Overall, approximately 2.2 times more clinical irrelevant findings (i.e. ≤CIN1) were found in the hrHPV-based programme, compared with approximately 1·3 times more clinically relevant findings (i.e. CIN2+); this difference was mostly due to a national policy change recommending colposcopy, rather than observation, of hrHPV-positive, ASC-US/LSIL results in the hrHPV-based programme. CONCLUSIONS: This is the first time that comprehensive results of nationwide implementation of hrHPV-based screening have been reported using high-quality data with a long follow-up. We have shown that both benefits and potential harms are higher in one screening round of a well-implemented hrHPV-based screening programme than in an established cytology-based programme. Lower participation in the new hrHPV programme may be due to factors such as invitation policy changes and the phased roll-out of the new programme. Our findings add further to evidence from trials and modelling studies on the effectiveness of hrHPV-based screening.

Colorectal metastasis to the gallbladder mimicking a primary gallbladder malignancy: histopathological and molecular characteristics.

AIMS: Outcomes of colorectal cancer (CRC) treatment and survival have steadily improved during the past decades, accompanied by an increased risk of developing second primary tumours and metastatic tumours at unusual sites. Metastatic CRC can show mucosal colonisation, thereby mimicking a second primary tumour. This potential confusion could lead to incorrect diagnosis and consequently inadequate treatment of the patient. The aim of this study was to differentiate between metastatic CRC and a second primary (gallbladder cancer, GBC) using a combination of standard histopathology and molecular techniques. METHODS AND RESULTS: Ten consecutive patients with both CRC and GBC were identified in our region using the Dutch National Pathology Archive (PALGA). Two patients served as negative controls. Histology of GBC was reviewed by nine pathologists. A combination of immunohistochemistry, microsatellite analysis, genomewide DNA copy number analysis and targeted somatic mutation analysis was used to aid in differential diagnosis. In two patients, CRC and GBC were clonally related, as confirmed by somatic mutation analysis. For one case, this was confirmed by genomewide DNA copy number analysis. However, in both cases, pathologists initially considered the GBC as a second primary tumour. CONCLUSIONS: Metastatic CRC displaying mucosal colonisation is often misinterpreted as a second primary tumour. A combination of traditional histopathology and molecular techniques improves this interpretation, and lowers the risk of inadequate treatment.

Contrast-enhanced ultrasound tractography for 3D vascular imaging of the prostate.

Diffusion tensor tractography (DTT) enables visualization of fiber trajectories in soft tissue using magnetic resonance imaging. DTT exploits the anisotropic nature of water diffusion in fibrous structures to identify diffusion pathways by generating streamlines based on the principal diffusion vector. Anomalies in these pathways can be linked to neural deficits. In a different field, contrast-enhanced ultrasound is used to assess anomalies in blood flow with the aim of locating cancer-induced angiogenesis. Like water diffusion, blood flow and transport of contrast agents also shows a principal direction; however, this is now determined by the local vasculature. Here we show how the tractographic techniques developed for magnetic resonance imaging DTT can be translated to contrast-enhanced ultrasound, by first estimating contrast flow velocity fields from contrast-enhanced ultrasound acquisitions, and then applying tractography. We performed 4D in-vivo contrast-enhanced ultrasound of three human prostates, proving the feasibility of the proposed approach with clinically acquired datasets. By comparing the results to histopathology after prostate resection, we observed qualitative agreement between the contrast flow tracts and typical markers of cancer angiogenic microvasculature: higher densities and tortuous geometries in tumor areas. The method can be used in-vivo using a standard contrast-enhanced ultrasound protocol, opening up new possibilities in the area of vascular characterization for cancer diagnostics.

Concordance of Gleason grading with three-dimensional ultrasound systematic biopsy and biopsy core pre-embedding.

PURPOSE: To determine the value of a three-dimensional (3D) greyscale transrectal ultrasound (TRUS)-guided prostate biopsy system and biopsy core pre-embedding method on concordance between Gleason scores of needle biopsies and radical prostatectomy (RP) specimens. METHODS: Retrospective analysis of prostate biopsies and subsequent RP for PCa in the Jeroen Bosch Hospital, the Netherlands, from 2007 to 2016. Two cohorts were analysed: conventional 2D TRUS-guided biopsies and RP (2007-2013, n = 266) versus 3D TRUS-guided biopsies with pre-embedding (2013-2016, n = 129). The impact of 3D TRUS-guidance with pre-embedding on Gleason score (GS) concordance between biopsy and RP was evaluated using the κ-coefficient. Predictors of biopsy GS 6 upgrading were assessed using logistic regression models. RESULTS: Gleason concordance was comparable between the two cohorts with a κ = 0.44 for the 3D cohort, compared to κ = 0.42 for the 2D cohort. 3D TRUS-guidance with pre-embedding, did not significantly affect the risk of biopsy GS 6 upgrading in univariate and multivariate analysis. CONCLUSIONS: 3D TRUS-guidance with biopsy core pre-embedding did not improve Gleason concordance. Improved detection techniques are needed for recognition of low-grade disease upgrading.

Prostate Cancer Risk Assessment in Biopsy-naïve Patients: The Rotterdam Prostate Cancer Risk Calculator in Multiparametric Magnetic Resonance Imaging-Transrectal Ultrasound (TRUS) Fusion Biopsy and Systematic TRUS Biopsy.

BACKGROUND: The value of multiparametric magnetic resonance imaging (mpMRI) and targeted biopsy (TBx) remains controversial for biopsy-naïve men when compared to transrectal ultrasound (TRUS)-guided systematic biopsy (SBx). Risk-based patient selection could help to selectively identify men with significant prostate cancer (PCa) and thus reduce unnecessary mpMRI and biopsies. OBJECTIVES: To compare PCa detection rates for mpMRI TBx with SBx and to determine the rate of potentially avoided mpMRI and biopsies through risk-based selection using the Rotterdam Prostate Cancer Risk Calculator (RPCRC). DESIGN, SETTING, AND PARTICIPANTS: Two-hundred consecutive biopsy-naïve men in two centres underwent mpMRI scanning, 12-core SBx, and subsequent MRI-TRUS TBx in the case of suspicious lesion(s) (Prostate Imaging-Reporting and Data System v.2 score ≥3). OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: We measured the detection rate for high-grade (Gleason score ≥ 3+4) PCa for TBx and SBx. We carried out a retrospective stratification according to RPCRC biopsy advice to determine the rate of mpMRI and biopsies that could potentially be avoided by RPCRC-based patient selection in relation to the rate of high-grade PCa missed. RESULTS AND LIMITATIONS: TBx yielded high-grade PCa in 51 men (26%) and low-grade PCa in 14 men (7%), while SBx yielded high-grade PCa in 63 men (32%) and low-grade PCa in 41 men (21%). Four out of 73 men (5%) with negative RPCRC advice and 63 out of 127 men (50%) with positive advice had high-grade PCa. Upfront RPCRC-based patient selection for mpMRI and TBx would have avoided 73 out of 200 (37%) mpMRI scans, missing two out of 51 (4%) high-grade PCas. Limitations include the RPCRC definition of high- and low-grade PCa and different mpMRI techniques. CONCLUSIONS: mpMRI with TBx detected PCa with high Gleason score and avoided biopsy in low-grade PCa, but failed to detect all high-grade PCa when compared to SBx among biopsy-naïve men. Risk-based patient selection using the RPCRC can avoid one-third of mpMRI scans and SBx in biopsy-naïve men. PATIENT SUMMARY: Men with a suspicion of prostate cancer are increasingly undergoing a magnetic resonance imaging (MRI) scan. Although promising, MRI-targeted biopsy is not accurate enough to safely replace systematic prostate biopsy for now. Individualised assessment of prostate cancer risk using the Rotterdam Prostate Cancer Risk Calculator could avoid one-third of MRI scans and systematic prostate biopsies.

Strategic framework for education and training in Quality by Design (QbD) and process analytical technology (PAT).

The regulatory and technical landscape of the pharmaceutical field is rapidly evolving from one focused predominantly on development of small molecules, using well established manufacturing technologies towards an environment in which biologicals and complex modalities are being developed using advanced science and technology coupled with the application of modern Quality by Design (QbD) principles. In order that Europe keeps pace with these changes and sustains its position as major player in the development and commercialization of medicines, it is essential that measures are put in place to maintain a highly skilled workforce. A number of challenges however exist to equipping academic, industrial and health agency staff with the requisite knowledge, skills and experience to develop the next generation of medicines. In this regard, the EUFEPS QbD and PAT Sciences Network has proposed a structured framework for education, training and continued professional development, which comprises a number of pillars covering the fundamental principles of modern pharmaceutical development including the underpinning aspects of science, engineering and technology innovation. The framework is not prescriptive and is not aimed at describing specific course content in detail. It should however be used as a point of reference for those institutions delivering pharmaceutical based educational courses, to ensure that the necessary skills, knowledge and experience for successful pharmaceutical development are maintained. A positive start has been made and a number of examples of formal higher education courses and short training programs containing elements of this framework have been described. The ultimate vision for this framework however, is to see widespread adoption and proliferation of this curriculum with it forming the backbone of QbD and PAT science based skills development.

The Prognostic Value of Microsatellite Instability, KRAS, BRAF and PIK3CA Mutations in Stage II Colon Cancer Patients.

In the era of personalized cancer medicine, identifying mutations within patient tumors plays an important role in defining high-risk stage II colon cancer patients. The prognostic role of BRAF V600E mutation, microsatellite instability (MSI) status, KRAS mutation and PIK3CA mutation in stage II colon cancer patients is not settled. We retrospectively analyzed 186 patients with stage II colon cancer who underwent an oncological resection but were not treated with adjuvant chemotherapy. KRAS mutations, PIK3CA mutation, V600E BRAF mutation and MSI status were determined. Survival analyses were performed. Mutations were found in the patients with each mutation in the following percentages: 23% (MSI), 35% (KRAS), 19% (BRAF) and 11% (PIK3CA). A trend toward worse overall survival (OS) was seen in patients with an MSI (5-year OS 74% versus 82%, adjusted hazard ratio [HR] 1.8, 95% confidence interval [CI] 0.6-4.9) and a KRAS-mutated tumor (5-year OS 77% versus 82%, adjusted HR 1.7, 95% CI 0.8-3.5). MSI and BRAF-mutated tumors tended to correlate with poorer disease-free survival (DFS) (5-year DFS 60% versus 78%, adjusted HR 1.6, 95% CI 0.5-2.1 and 5-year DFS 57% versus 77%, adjusted HR 1.1, 95% CI 0.4-2.6 respectively). In stage II colon cancer patients not treated with adjuvant chemotherapy, BRAF mutation and MSI status both tended to have a negative prognostic effect on disease-free survival. KRAS and MSI status also tended to be correlated with worse overall survival.