<i>Pseudomonas aeruginosa</i> intensive care unit outbreak:winnowing of transmissions with molecular and genomic typing

Bioinformatics and computational biology analyses were supported by the University of St Andrews Bioinformatics Unit which is funded by a Wellcome Trust ISSF award [grant 097831/Z/11/Z]. BJP, KO, MP, MTGH, GP and SHG are funded by the Chief Scientist Office through the Scottish Infection Research Network, a part of the SHAIPI consortium grant reference number SIRN/10.Background: Pseudomonas aeruginosa healthcare outbreaks can be time consuming and difficult to investigate. Guidance does not specify which typing technique is most practical to base decisions on. Aim: We explore the usefulness of whole genome sequencing (WGS) in the investigation of a Pseudomonas aeruginosa outbreak describing how it compares with pulsed-field gel electrophoresis (PFGE) and variable number tandem repeat (VNTR) analysis. Methods: Six patient isolates and six environmental samples from an Intensive Care Unit (ICU) positive for P. aeruginosa over two years underwent VNTR, PFGE and WGS. Findings:  VNTR and PFGE were required to fully determine the potential source of infection and rule out others. WGS results unambiguously distinguished linked isolates giving greater assurance of the transmission route between wash hand basin (WHB) water and two patients supporting control measures employed. Conclusion:  WGS provided detailed information without need for further typing. When allied to epidemiological information it can be used to understand outbreak situations rapidly and with certainty. Implementation of WGS in real-time would be a major advance in day-to-day practice. It could become a standard of care as it becomes more widespread due to its reproducibility and reduction in costs.Publisher PDFPeer reviewe

Iodine Atoms: A New Molecular Feature for the Design of Potent Transthyretin Fibrillogenesis Inhibitors

The thyroid hormone and retinol transporter protein known as transthyretin (TTR) is in the origin of one of the 20 or so known amyloid diseases. TTR self assembles as a homotetramer leaving a central hydrophobic channel with two symmetrical binding sites. The aggregation pathway of TTR into amiloid fibrils is not yet well characterized but in vitro binding of thyroid hormones and other small organic molecules to TTR binding channel results in tetramer stabilization which prevents amyloid formation in an extent which is proportional to the binding constant. Up to now, TTR aggregation inhibitors have been designed looking at various structural features of this binding channel others than its ability to host iodine atoms. In the present work, greatly improved inhibitors have been designed and tested by taking into account that thyroid hormones are unique in human biochemistry owing to the presence of multiple iodine atoms in their molecules which are probed to interact with specific halogen binding domains sitting at the TTR binding channel. The new TTR fibrillogenesis inhibitors are based on the diflunisal core structure because diflunisal is a registered salicylate drug with NSAID activity now undergoing clinical trials for TTR amyloid diseases. Biochemical and biophysical evidence confirms that iodine atoms can be an important design feature in the search for candidate drugs for TTR related amyloidosis

Milkability assessment of Tsigai, Improved Valachian, Lacaune and F1Crossbred ewes (Tsigai × Lacaune, Improved Valachian × Lacaune) throughout lactation

This paper aimed to compare the milking ability of ewes from pure European breeds (Tsigai, Improved Valachian, Lacaune, and their crosses) on the basis of their milk flow kinetics during machine milking throughout lactation. Ewes of the Tsigai (TS, n = 13), Improved Valachian (IV, n = 12), Lacaune (LC, n = 12), TS x LC (50%, n = 12) and IV x LC (50%, n = 12) on their 1-3 lactation were used. The ewes were in their 74 +/- 12 d of lactation at the beginning of the experiment. Milk flow data of each ewe were recorded during one evening milking in the middle of four successive months (May, June, July, and August). Milk flow was recorded by means of an automatic collecting jar equipped with an electronic milk meter device working in every 1-s intervals. The milk flow curves were classified into three types: 1 peak (1P), 2 peaks (2P) and plateau (PL). The highest values of repeatability of parameters were found for stripped milk yield (SMY), maximum milk flow, latency time and milk yield at 30s (over 0.52). The lowest values were recorded for machine milking time (MMT), machine milk yield (MMY) and time to reach maximum milk flow (<0.33). Month of lactation significantly influenced most of the milkability data except latency time and time to maximum flow rate. The effect of genotype showed highly significant influence on most of the analyzed parameters. The lowest total milk yield (TMY) and MMY were measured in pure TS (0.348 +/- 0.021 and 0.266 +/- 0.0211, respectively) and IV breeds (0.320 +/- 0.0221 and 0.259 +/- 0.0211, respectively) as compared with Lacaune (0.524 +/- 0.0221 and 0.360 +/- 0.0211, respectively). The highest TMY and MMY were measured in ewes with PL milk flow types and the lowest in ewes with 1P. Ewes with 1P showed clearly shorter MMT and higher percentage of SMY as compared to PL and 2P though SMY did not differ among groups. Interaction of milk flow type and stage of lactation influenced TMY and MMY, maximum milk flow, milk yield in 30s of milking and MMT. A clear effect of milk flow type on TMY and MMY was observed each month

Target-enrichment sequencing yields valuable genomic data for challenging-to-culture bacteria of public health importance

Genomic data contribute invaluable information to the epidemiological investigation of pathogens of public health importance. However, whole-genome sequencing (WGS) of bacteria typically relies on culture, which represents a major hurdle for generating such data for a wide range of species for which culture is challenging. In this study, we assessed the use of culture-free target-enrichment sequencing as a method for generating genomic data for two bacterial species: (1) Bacillus anthracis, which causes anthrax in both people and animals and whose culture requires high-level containment facilities; and (2) Mycoplasma amphoriforme, a fastidious emerging human respiratory pathogen. We obtained high-quality genomic data for both species directly from clinical samples, with sufficient coverage (&gt;15×) for confident variant calling over at least 80% of the baited genomes for over two thirds of the samples tested. Higher qPCR cycle threshold (Ct) values (indicative of lower pathogen concentrations in the samples), pooling libraries prior to capture, and lower captured library concentration were all statistically associated with lower capture efficiency. The Ct value had the highest predictive value, explaining 52 % of the variation in capture efficiency. Samples with Ct values ≤30 were over six times more likely to achieve the threshold coverage than those with a Ct &gt; 30. We conclude that target-enrichment sequencing provides a valuable alternative to standard WGS following bacterial culture and creates opportunities for an improved understanding of the epidemiology and evolution of many clinically important pathogens for which culture is challenging

Staphylococcal bacteremia in cancer patients: Risk factors and outcome in 134 episodes prior to and after introduction of quinolones into infection prevention in neutropenia

A total of 134 episodes of staphylococcal bacteremia (SBE) appearing among 9987 admissions, and 979 episodes of bacteremia in cancer patients within 5 years, were analyzed for risk factors, clinical course and outcome; 64 were monomicrobial and 70 polymicrobial. The most frequent risk factors were acute leukemia, catheter insertion, long-lasting neutropenia, and prior prophylaxis with quinolones. There was no significant difference between polymicrobial and monomicrobial SBE in risk factors. The two groups differed only in the source of bacteremia (gastrointestinal and respiratory-tract infections were more common in monomicrobial SBE) and etiology - Staphylococcus aureus appeared more frequently in monomicrobial than in polymicrobial bacteremia (20.3% compared to 4.3%, P<0.05). More complications (14.3%) such as abscesses, endocarditis, etc. appeared in the group of polymicrobial SBE (P<0.05). No difference was observed in clinical course and outcome between monomicrobial and polymicrobial SBE. The incidence of SBE has increased since 1991, when quinolones were first used in prophylaxis in afebrile neutropenia at our center; however, the infection-associated mortality in monomicrobial SBE was low (4.3%)

Efficacy and Safety of Trastuzumab Emtansine plus Capecitabine vs Trastuzumab Emtansine Alone in Patients with Previously Treated ERBB2 (HER2)-Positive Metastatic Breast Cancer: A Phase 1 and Randomized Phase 2 Trial

Importance: ERBB2 (HER2)-targeted therapy provides benefits in metastatic breast cancer (mBC) and gastric cancer, but additional treatments are needed to maximize efficacy and quality of life. Objective: To determine maximum tolerated doses (MTDs) of trastuzumab emtansine (T-DM1) plus capecitabine in patients with previously treated ERBB2-positive mBC and locally advanced/metastatic gastric cancer (LA/mGC) (phase 1) and the efficacy and safety of this combination vs T-DM1 alone in patients with mBC (phase 2). Design, Setting, and Participants: The MTD in phase 1 was assessed using a 3 + 3 design with capecitabine dose modification. Phase 2 was an open-label, randomized, international multicenter study of patients with mBC treated with T-DM1 plus capecitabine or T-DM1 alone. Eligible patients had previously treated ERBB2-positive mBC or LA/mGC with no prior chemotherapy treatment for advanced disease. Interventions: Patients in the phase 1 mBC cohort received capecitabine (750 mg/m2, 700 mg/m2, or 650 mg/m2 twice daily, days 1-14 of a 3-week cycle) plus T-DM1 3.6 mg/kg every 3 weeks. Patients with LA/mGC received capecitabine at the mBC phase 1 MTD, de-escalating as needed, plus T-DM1 2.4 mg/kg weekly. In phase 2, patients with mBC were randomized (1:1) to receive capecitabine (at the phase 1 MTD) plus T-DM1 or T-DM1 alone. Main Outcomes and Measures: The phase 1 primary objective was to identify the MTD of capecitabine plus T-DM1. The phase 2 primary outcome was investigator-assessed overall response rate (ORR). Results: In phase 1, the median (range) age was 54.0 (37-71) and 57.5 (53-70) years for patients with mBC and patients with LA/mGC, respectively. The capecitabine MTD was identified as 700 mg/m2 in 11 patients with mBC and 6 patients with LA/mGC evaluable for dose-limiting toxic effects. In phase 2, between October 2014 and April 2016, patients with mBC (median [range] age, 52.0 [28-80] years) were randomized to receive combination therapy (n = 81) or T-DM1 (n = 80). The ORR was 44% (36 of 81 patients) and 36% (29 of 80 patients) in the combination and T-DM1 groups, respectively (difference, 8.2%; 90% CI, -4.5 to 20.9; P =.34; clinical cutoff, May 31, 2017). Adverse events (AEs) were reported in 78 of 82 patients (95%) in the combination group, with 36 (44%) experiencing grade 3-4 AEs, and 69 of 78 patients (88%) in the T-DM1 group, with 32 (41%) experiencing grade 3-4 AEs. No grade 5 AEs were reported. Conclusions and Relevance: Adding capecitabine to T-DM1 did not statistically increase ORR associated with T-DM1 in patients with previously treated ERBB2-positive mBC. The combination group reported more AEs, but with no unexpected toxic effects. Trial Registration: ClinicalTrials.gov Identifier: NCT01702558