A Web-Based Peer Support Network to Help Care Partners of People With Serious Illness: Co-Design Study.

BACKGROUND: Care partners of people with serious illness experience significant challenges and unmet needs during the patient's treatment period and after their death. Learning from others with shared experiences can be valuable, but opportunities are not consistently available. OBJECTIVE: This study aims to design and prototype a regional, facilitated, and web-based peer support network to help active and bereaved care partners of persons with serious illness be better prepared to cope with the surprises that arise during serious illness and in bereavement. METHODS: An 18-member co-design team included active care partners and those in bereavement, people who had experienced serious illness, regional health care and support partners, and clinicians. It was guided by facilitators and peer network subject-matter experts. We conducted design exercises to identify the functions and specifications of a peer support network. Co-design members independently prioritized network specifications, which were incorporated into an early iteration of the web-based network. RESULTS: The team prioritized two functions: (1) connecting care partners to information and (2) facilitating emotional support. The design process generated 24 potential network specifications to support these functions. The highest priorities included providing a supportive and respectful community; connecting people to trusted resources; reducing barriers to asking for help; and providing frequently asked questions and responses. The network platform had to be simple and intuitive, provide technical support for users, protect member privacy, provide publicly available information and a private discussion forum, and be easily accessible. It was feasible to enroll members in the ConnectShareCare web-based network over a 3-month period. CONCLUSIONS: A co-design process supported the identification of critical features of a peer support network for care partners of people with serious illnesses in a rural setting, as well as initial testing and use. Further testing is underway to assess the long-term viability and impact of the network.

Symptomatic and Functional Outcomes Among Individuals at High Risk for Psychosis Participating in Step-Based Care.

Validated, multicomponent treatments designed to address symptoms and functioning of individuals at clinical high risk for psychosis are currently lacking. The authors report findings of a study with such individuals participating in step-based care-a program designed to provide low-intensity, non-psychosis-specific interventions and advancement to higher-intensity, psychosis-specific interventions only if an individual is not meeting criteria for a clinical response. Among individuals with symptomatic or functional concerns at enrollment, 67% met criteria for a symptomatic response (median time to response=11.1 weeks), and 64% met criteria for a functional response (median time to response=8.9 weeks).

Phylogenetic relationship and virulence composition of Escherichia coli O26:H11 cattle and human strain collections in Scotland; 2002-2020.

O26 is the commonest non-O157 Shiga toxin (stx)-producing Escherichia coli serogroup reported in human infections worldwide. Ruminants, particularly cattle, are the primary reservoir source for human infection. In this study, we compared the whole genomes and virulence profiles of O26:H11 strains (n = 99) isolated from Scottish cattle with strains from human infections (n = 96) held by the Scottish Escherichia coli O157/STEC Reference Laboratory, isolated between 2002 and 2020. Bovine strains were from two national cross-sectional cattle surveys conducted between 2002-2004 and 2014-2015. A maximum likelihood phylogeny was constructed from a core-genome alignment with the O26:H11 strain 11368 reference genome. Genomes were screened against a panel of 2,710 virulence genes using the Virulence Finder Database. All stx-positive bovine O26:H11 strains belonged to the ST21 lineage and were grouped into three main clades. Bovine and human source strains were interspersed, and the stx subtype was relatively clade-specific. Highly pathogenic stx2a-only ST21 strains were identified in two herds sampled in the second cattle survey and in human clinical infections from 2010 onwards. The closest pairwise distance was 9 single-nucleotide polymorphisms (SNPs) between Scottish bovine and human strains and 69 SNPs between the two cattle surveys. Bovine O26:H11 was compared to public EnteroBase ST29 complex genomes and found to have the greatest commonality with O26:H11 strains from the rest of the UK, followed by France, Italy, and Belgium. Virulence profiles of stx-positive bovine and human strains were similar but more conserved for the stx2a subtype. O26:H11 stx-negative ST29 (n = 17) and ST396 strains (n = 5) were isolated from 19 cattle herds; all were eae-positive, and 10 of these herds yielded strains positive for ehxA, espK, and Z2098, gene markers suggestive of enterohaemorrhagic potential. There was a significant association (p < 0.001) between nucleotide sequence percent identity and stx status for the bacteriophage insertion site genes yecE for stx2 and yehV for stx1. Acquired antimicrobial resistance genes were identified in silico in 12.1% of bovine and 17.7% of human O26:H11 strains, with sul2, tet, aph(3″), and aph(6″) being most common. This study describes the diversity among Scottish bovine O26:H11 strains and investigates their relationship to human STEC infections.

Analysis of Escherichia coli O157 strains in cattle and humans between Scotland and England & Wales: implications for human health.

For the last two decades, the human infection frequency of Escherichia coli O157 (O157) in Scotland has been 2.5-fold higher than in England and Wales. Results from national cattle surveys conducted in Scotland and England and Wales in 2014/2015 were combined with data on reported human clinical cases from the same time frame to determine if strain differences in national populations of O157 in cattle could be associated with higher human infection rates in Scotland. Shiga toxin subtype (Stx) and phage type (PT) were examined within and between host (cattle vs human) and nation (Scotland vs England and Wales). For a subset of the strains, whole genome sequencing (WGS) provided further insights into geographical and host association. All three major O157 lineages (I, II, I/II) and most sub-lineages (Ia, Ib, Ic, IIa, IIb, IIc) were represented in cattle and humans in both nations. While the relative contribution of different reservoir hosts to human infection is unknown, WGS analysis indicated that the majority of O157 diversity in human cases was captured by isolates from cattle. Despite comparable cattle O157 prevalence between nations, strain types were localized. PT21/28 (sub-lineage Ic, Stx2a+) was significantly more prevalent in Scottish cattle [odds ratio (OR) 8.7 (2.3-33.7; P<0.001] and humans [OR 2.2 (1.5-3.2); P<0.001]. In England and Wales, cattle had a significantly higher association with sub-lineage IIa strains [PT54, Stx2c; OR 5.6 (1.27-33.3); P=0.011] while humans were significantly more closely associated with sub-lineage IIb [PT8, Stx1 and Stx2c; OR 29 (4.9-1161); P<0.001]. Therefore, cattle farms in Scotland were more likely to harbour Stx2a+O157 strains compared to farms in E and W (P<0.001). There was evidence of limited cattle strain migration between nations and clinical isolates from one nation were more similar to cattle isolates from the same nation, with sub-lineage Ic (mainly PT21/28) exhibiting clear national association and evidence of local transmission in Scotland. While we propose the higher rate of O157 clinical cases in Scotland, compared to England and Wales, is a consequence of the nationally higher level of Stx2a+O157 strains in Scottish cattle, we discuss the multiple additional factors that may also contribute to the different infection rates between these nations.

Prevalence of Shiga Toxin-Producing Escherichia coli O157 in Wild Scottish Deer with High Human Pathogenic Potential.

Shiga toxin-producing E. coli (STEC) infections associated with wildlife are increasing globally, highlighting many 'spillover' species as important reservoirs for these zoonotic pathogens. A human outbreak of STEC serogroup O157 in 2015 in Scotland, associated with the consumption of venison meat products, highlighted several knowledge gaps, including the prevalence of STEC O157 in Scottish wild deer and the potential risk to humans from wild deer isolates. In this study, we undertook a nationwide survey of wild deer in Scotland and determined that the prevalence of STEC O157 in wild deer is low 0.28% (95% confidence interval = 0.06-0.80). Despite the low prevalence of STEC O157 in Scottish wild deer, identified isolates were present in deer faeces at high levels (>104 colony forming units/g faeces) and had high human pathogenic potential based on whole genome sequencing and virulence gene profiling. A retrospective epidemiological investigation also identified one wild deer isolate from this study as a possible source of a Scottish human outbreak in 2017. These results emphasise the importance of food hygiene practices during the processing of wild deer carcasses for human consumption.

Harnessing the Collective Expertise of Patients, Care Partners, Clinical Teams, and Researchers Through a Coproduction Learning Health System: A Case Study of the Dartmouth Health Promise Partnership.

The coproduction learning health system (CLHS) model extends the definition of a learning health system to explicitly bring together patients and care partners, health care teams, administrators, and scientists to share the work of optimizing health outcomes, improving care value, and generating new knowledge. The CLHS model highlights a partnership for coproduction that is supported by data that can be used to support individual patient care, quality improvement, and research. We provide a case study that describes the application of this model to transform care within an oncology program at an academic medical center.

Shiga toxin-producing Escherichia coli clonal complex 32, including serotype O145:H28, in the UK and Ireland.

Introduction. Shiga toxin-producing Escherichia coli (STEC) O157:H7 has been the most clinically significant STEC serotype in the UK for over four decades. Over the last 10 years we have observed a decrease in STEC O157:H7 and an increase in non-O157 STEC serotypes, such as O145:H28.Gap Statement. Little is known about the microbiology and epidemiology of STEC belonging to CC32 (including O145:H28) in the UK. The aim of this study was to integrate genomic data with patient information to gain a better understanding of the virulence, disease severity, epidemic risk assessment and population structure of this clinically significant clonal complex.Methodology. Isolates of E. coli belonging to CC32 (n=309) in the archives of public health agencies in the UK and Ireland were whole-genome-sequenced, virulence-profiled and integrated with enhanced surveillance questionnaire (ESQ) data, including exposures and disease severity.Results. Overall, diagnoses of STEC belonging to CC32 (290/309, 94 %) in the UK have increased every year since 2014. Most cases were female (61 %), and the highest proportion of cases belonged to the 0-4 age group (53/211,25 %). The frequency of symptoms of diarrhoea (92 %), abdominal pain (84 %), blood in stool (71 %) and nausea (51 %) was similar to that reported in cases of STEC O157:H7, although cases of STEC CC32 were more frequently admitted to hospital (STEC CC32 48 % vs O157:H7  34 %) and/or developed haemolytic uraemic syndrome (HUS) (STEC CC32 9 % vs O157:H7 4 %).The majority of STEC isolates (268/290, 92 %) had the stx2a/eae virulence gene combination, most commonly associated with progression to STEC HUS. There was evidence of person-to-person transmission and small, temporally related, geographically dispersed outbreaks, characteristic of foodborne outbreaks linked to nationally distributed products.Conclusion. We recommend more widespread use of polymerase chain reaction (PCR) for the detection of all STEC serogroups, the development of consistent strategies for the follow-up testing of PCR-positive faecal specimens, the implementation of more comprehensive and standardized collection of epidemiological data, and routine sharing of sequencing data between public health agencies worldwide.

Prioritizing Measures That Matter Within a Person-Centered Oncology Learning Health System.

BACKGROUND: Despite progress in developing learning health systems (LHS) and associated metrics of success, a gap remains in identifying measures to guide the implementation and assessment of the impact of an oncology LHS. Our aim was to identify a balanced set of measures to guide a person-centered oncology LHS. METHODS: A modified Delphi process and clinical value compass framework were used to prioritize measures for tracking LHS performance. A multidisciplinary group of 77 stakeholders, including people with cancer and family members, participated in 3 rounds of online voting followed by 50-minute discussions. Participants rated metrics on perceived importance to the LHS and discussed priorities. RESULTS: Voting was completed by 94% of participants and prioritized 22 measures within 8 domains. Patient and caregiver factors included clinical health (Eastern Cooperative Oncology Group Performance Status, survival by cancer type and stage), functional health and quality of life (Patient Reported Outcomes Measurement Information System [PROMIS] Global-10, Distress Thermometer, Modified Caregiver Strain Index), experience of care (advance care planning, collaboRATE, PROMIS Self-Efficacy Scale, access to care, experience of care, end-of-life quality measures), and cost and resource use (avoidance and delay in accessing care and medications, financial hardship, total cost of care). Contextual factors included team well-being (Well-being Index; voluntary staff turnover); learning culture (Improvement Readiness, compliance with Commission on Cancer quality of care measures); scholarly engagement and productivity (institutional commitment and support for research, academic productivity index); and diversity, equity, inclusion, and belonging (screening and follow-up for social determinants of health, inclusivity of staff and patients). CONCLUSIONS: The person-centered LHS value compass provides a balanced set of measures that oncology practices can use to monitor and evaluate improvement across multiple domains.

Epidemiology and genomic analysis of Shiga toxin-producing Escherichia coli clonal complex 165 in the UK.

Introduction. Shiga toxin-producing Escherichia coli (STEC) is a zoonotic, foodborne gastrointestinal pathogen that has the potential to cause severe clinical outcomes, including haemolytic uraemic syndrome (HUS). STEC-HUS is the leading cause of renal failure in children and can be fatal. Over the last decade, STEC clonal complex 165 (CC165) has emerged as a cause of STEC-HUS.Gap Statement. There is a need to understand the pathogenicity and prevalence of this emerging STEC clonal complex in the UK, to facilitate early diagnosis, improve clinical management, and prevent and control outbreaks.Aim. The aim of this study was to characterize CC165 through identification of virulence factors (VFs) and antimicrobial resistance (AMR) determinants in the genome and to integrate the genome data with the available epidemiological data to better understand the incidence and pathogenicity of this clonal complex in the UK.Methodology. All isolates belonging to CC165 in the archives at the UK public health agencies were sequenced and serotyped, and the virulence gene and AMR profiles were derived from the genome using PHE bioinformatics pipelines and the Centre for Genomic Epidemiology virulence database.Results. There were 48 CC165 isolates, of which 43 were STEC, four were enteropathogenic E. coli (EPEC) and one E. coli. STEC serotypes were predominately O80:H2 (n=28), and other serotypes included O45:H2 (n=9), O55:H9 (n=4), O132:H2 (n=1) and O180:H2 (n=1). All but one STEC isolate had Shiga toxin (stx) subtype stx2a or stx2d and 47/48 isolates had the eae gene encoding intimin involved in the intimate attachment of the bacteria to the human gut mucosa. We detected extra-intestinal virulence genes including those associated with iron acquisition (iro) and serum resistance (iss), indicating that this pathogen has the potential to translocate to extra-intestinal sites. Unlike other STEC clonal complexes, a high proportion of isolates (93%, 40/43) were multidrug-resistant, including resistance to aminoglycosides, beta-lactams, chloramphenicol, sulphonamides, tetracyclines and trimethoprim.Conclusion. The clinical significance of this clonal complex should not be underestimated. Exhibiting high levels of AMR and a combination of STEC and extra-intestinal pathogenic E. coli (ExPEC) virulence profiles, this clonal complex is an emerging threat to public health.

Prevalence and Epidemiology of Non-O157 Escherichia coli Serogroups O26, O103, O111, and O145 and Shiga Toxin Gene Carriage in Scottish Cattle, 2014-2015.

Cattle are a reservoir for Shiga toxin-producing Escherichia coli (STEC), zoonotic pathogens that cause serious clinical disease. Scotland has a higher incidence of STEC infection in the human population than the European average. The aim of this study was to investigate the prevalence and epidemiology of non-O157 serogroups O26, O103, O111, and O145 and Shiga toxin gene carriage in Scottish cattle. Fecal samples (n = 2783) were collected from 110 herds in 2014 and 2015 and screened by real-time PCR. Herd-level prevalence (95% confidence interval [CI]) for O103, O26, and O145 was estimated as 0.71 (0.62, 0.79), 0.43 (0.34, 0.52), and 0.23 (0.16, 0.32), respectively. Only two herds were positive for O111. Shiga toxin prevalence was high in both herds and pats, particularly for stx2 (herd level: 0.99; 95% CI: 0.94, 1.0). O26 bacterial strains were isolated from 36 herds on culture. Fifteen herds yielded O26 stx-positive isolates that additionally harbored the intimin gene; six of these herds shed highly pathogenic stx2-positive strains. Multiple serogroups were detected in herds and pats, with only 25 herds negative for all serogroups. Despite overlap in detection, regional and seasonal effects were observed. Higher herd prevalence for O26, O103, and stx1 occurred in the South West, and this region was significant for stx2 at the pat level (P = 0.015). Significant seasonal variation was observed for O145 prevalence, with the highest prevalence in autumn (P = 0.032). Negative herds were associated with Central Scotland and winter. Herds positive for all serogroups were associated with autumn and larger herd size and were not housed at sampling.IMPORTANCE Cattle are reservoirs for Shiga toxin-producing Escherichia coli (STEC), bacteria shed in animal feces. Humans are infected through consumption of contaminated food or water and by direct contact, resulting in serious disease and kidney failure in the most vulnerable. The contribution of non-O157 serogroups to STEC illness was underestimated for many years due to the lack of specific tests. Recently, non-O157 human cases have increased, with O26 STEC of particular note. It is therefore vital to investigate the level and composition of non-O157 in the cattle reservoir and to compare them historically and by the clinical situation. In this study, we found cattle prevalence high for toxin, as well as for O103 and O26 serogroups. Pathogenic O26 STEC were isolated from 14% of study herds, with toxin subtypes similar to those seen in Scottish clinical cases. This study highlights the current risk to public health from non-O157 STEC in Scottish cattle.

ITGB2 (Integrin ß2) Immunomodulatory Gene Variants in Premature Infants With Necrotizing Enterocolitis.

ABSTRACT: Aberrant toll-like receptor (TLR) activation is central to necrotizing enterocolitis (NEC) pathogenesis. ß2 integrins regulate TLR signaling, and integrin ß2 (ITGB2) deficiency causes TLR hyperresponsiveness. To test the hypothesis that ITGB2 genetic variants modulate NEC susceptibility, we sequenced the exonic ITGB2 locus to compare the prevalence of deleterious variants among 221 preterm infants with and without NEC. ITGB2 variants were not associated with NEC in our entire cohort (NEC [9/56] versus controls [16/165], P = 0.19) or in extremely low birthweight infants (ELBW, controls [7.9%] versus NEC [18.2%]; P = 0.11) but were increased compared to the populace (4.5%, gnomad.broadinstitute.org). Combined annotation-dependent depletion -predicted deleterious ITGB2 variants increased proportionately with increasing NEC severity in ELBW infants (controls [6.7%] versus medical NEC [16.7%] versus surgical NEC [19%] (P = 0.03). Although ITGB2 variants were not associated with NEC in our preterm cohort, subgroup analysis showed a trend towards enrichment with NEC severity in ELBW infants.

Childhood Pulmonary Function, Exercise Capacity, and Exhaled Nitric Oxide Levels: Outcomes following Neonatal Treatment with Inhaled Nitric Oxide to Prevent Bronchopulmonary Dysplasia.

OBJECTIVE: The goal was to determine if inhaled nitric oxide (iNO) for 3 weeks during neonatal care of high-risk preterm infants was associated with improved pulmonary function and exercise capacity or altered exhaled nitric oxide (FeNO) levels in later childhood. STUDY DESIGN: Thirty-four very preterm children previously enrolled in a randomized, neonatal trial of iNO to prevent chronic lung disease, were assessed in follow-up at 7 to 9 years of age, including pulmonary function testing (PFT), exercise testing, and measurement of FeNO. RESULTS: There were no differences in PFTs or exercise capacity between iNO treated and controls. FeNO levels showed large interpatient variability but tended to be lower in the iNO treated. CONCLUSION: Findings indicate no overall differences in pulmonary function or exercise capacity for children who had neonatal iNO treatment compared with placebo.

The NSIGHT1-randomized controlled trial: rapid whole-genome sequencing for accelerated etiologic diagnosis in critically ill infants.

Genetic disorders are a leading cause of morbidity and mortality in infants in neonatal and pediatric intensive care units (NICU/PICU). While genomic sequencing is useful for genetic disease diagnosis, results are usually reported too late to guide inpatient management. We performed an investigator-initiated, partially blinded, pragmatic, randomized, controlled trial to test the hypothesis that rapid whole-genome sequencing (rWGS) increased the proportion of NICU/PICU infants receiving a genetic diagnosis within 28 days. The participants were families with infants aged <4 months in a regional NICU and PICU, with illnesses of unknown etiology. The intervention was trio rWGS. Enrollment from October 2014 to June 2016, and follow-up until November 2016. Of all, 26 female infants, 37 male infants, and 2 infants of undetermined sex were randomized to receive rWGS plus standard genetic tests (n = 32, cases) or standard genetic tests alone (n = 33, controls). The study was terminated early due to loss of equipoise: 73% (24) controls received genomic sequencing as standard tests, and 15% (five) controls underwent compassionate cross-over to receive rWGS. Nevertheless, intention to treat analysis showed the rate of genetic diagnosis within 28 days of enrollment (the primary end-point) to be higher in cases (31%, 10 of 32) than controls (3%, 1 of 33; difference, 28% [95% CI, 10-46%]; p = 0.003). Among infants enrolled in the first 25 days of life, the rate of neonatal diagnosis was higher in cases (32%, 7 of 22) than controls (0%, 0 of 23; difference, 32% [95% CI, 11-53%];p = 0.004). Median age at diagnosis (25 days [range 14-90] in cases vs. 130 days [range 37-451] in controls) and median time to diagnosis (13 days [range 1-84] in cases, vs. 107 days [range 21-429] in controls) were significantly less in cases than controls (p = 0.04). In conclusion, rWGS increased the proportion of NICU/PICU infants who received timely diagnoses of genetic diseases.

Validation of Whole-Genome Sequencing for Identification and Characterization of Shiga Toxin-Producing Escherichia coli To Produce Standardized Data To Enable Data Sharing.

Whole-genome sequencing (WGS) is rapidly becoming the method of choice for outbreak investigations and public health surveillance of microbial pathogens. The combination of improved cluster resolution and prediction of resistance and virulence phenotypes provided by a single tool is extremely advantageous. However, the data produced are complex, and standard bioinformatics pipelines are required to translate the output into easily interpreted epidemiologically relevant information for public health action. The main aim of this study was to validate the implementation of WGS at the Scottish Escherichia coli O157/STEC Reference Laboratory (SERL) using the Public Health England (PHE) bioinformatics pipeline to produce standardized data to enable interlaboratory comparison of results generated at two national reference laboratories. In addition, we evaluated the BioNumerics whole-genome multilocus sequence typing (wgMLST) and E. coli genotyping plug-in tools using the same data set. A panel of 150 well-characterized isolates of Shiga toxin-producing E. coli (STEC) that had been sequenced and analyzed at PHE using the PHE pipeline and database (SnapperDB) was assembled to provide identification and typing data, including serotype (O:H type), sequence type (ST), virulence genes (eae and Shiga toxin [stx] subtype), and a single-nucleotide polymorphism (SNP) address. To validate the implementation of sequencing at the SERL, DNA was reextracted from the isolates and sequenced and analyzed using the PHE pipeline, which had been installed at the SERL; the output was then compared with the PHE data. The results showed a very high correlation between the data, ranging from 93% to 100%, suggesting that the standardization of WGS between our reference laboratories is possible. We also found excellent correlation between the results obtained using the PHE pipeline and BioNumerics, except for the detection of stx2a and stx2c when these subtypes are both carried by strains.

Utility of Whole-Genome Sequencing of Escherichia coli O157 for Outbreak Detection and Epidemiological Surveillance.

Detailed laboratory characterization of Escherichia coli O157 is essential to inform epidemiological investigations. This study assessed the utility of whole-genome sequencing (WGS) for outbreak detection and epidemiological surveillance of E. coli O157, and the data were used to identify discernible associations between genotypes and clinical outcomes. One hundred five E. coli O157 strains isolated over a 5-year period from human fecal samples in Lothian, Scotland, were sequenced with the Ion Torrent Personal Genome Machine. A total of 8,721 variable sites in the core genome were identified among the 105 isolates; 47% of the single nucleotide polymorphisms (SNPs) were attributable to six "atypical" E. coli O157 strains and included recombinant regions. Phylogenetic analyses showed that WGS correlated well with the epidemiological data. Epidemiological links existed between cases whose isolates differed by three or fewer SNPs. WGS also correlated well with multilocus variable-number tandem repeat analysis (MLVA) typing data, with only three discordant results observed, all among isolates from cases not known to be epidemiologically related. WGS produced a better-supported, higher-resolution phylogeny than MLVA, confirming that the method is more suitable for epidemiological surveillance of E. coli O157. A combination of in silico analyses (VirulenceFinder, ResFinder, and local BLAST searches) were used to determine stx subtypes, multilocus sequence types (15 loci), and the presence of virulence and acquired antimicrobial resistance genes. There was a high level of correlation between the WGS data and our routine typing methods, although some discordant results were observed, mostly related to the limitation of short sequence read assembly. The data were used to identify sublineages and clades of E. coli O157, and when they were correlated with the clinical outcome data, they showed that one clade, Ic3, was significantly associated with severe disease. Together, the results show that WGS data can provide higher resolution of the relationships between E. coli O157 isolates than that provided by MLVA. The method has the potential to streamline the laboratory workflow and provide detailed information for the clinical management of patients and public health interventions.

Applying phylogenomics to understand the emergence of Shiga-toxin-producing Escherichia coli O157:H7 strains causing severe human disease in the UK.

Shiga-toxin-producing Escherichia coli (STEC) O157:H7 is a recently emerged zoonotic pathogen with considerable morbidity. Since the emergence of this serotype in the 1980s, research has focussed on unravelling the evolutionary events from the E. coli O55:H7 ancestor to the contemporaneous globally dispersed strains observed today. In this study, the genomes of over 1000 isolates from both human clinical cases and cattle, spanning the history of STEC O157:H7 in the UK, were sequenced. Phylogenetic analysis revealed the ancestry, key acquisition events and global context of the strains. Dated phylogenies estimated the time to evolution of the most recent common ancestor of the current circulating global clone to be 175 years ago. This event was followed by rapid diversification. We show the acquisition of specific virulence determinates has occurred relatively recently and coincides with its recent detection in the human population. We used clinical outcome data from 493 cases of STEC O157:H7 to assess the relative risk of severe disease including haemolytic uraemic syndrome from each of the defined clades in the population and show the dramatic effect Shiga toxin repertoire has on virulence. We describe two strain replacement events that have occurred in the cattle population in the UK over the last 30 years, one resulting in a highly virulent strain that has accounted for the majority of clinical cases in the UK over the last decade. There is a need to understand the selection pressures maintaining Shiga-toxin-encoding bacteriophages in the ruminant reservoir and the study affirms the requirement for close surveillance of this pathogen in both ruminant and human populations.

Multilocus sequence typing (MLST) of Staphylococcus aureus.

MLST is a widely accepted method of sequence-based typing that relies on analysis of relatively conserved genes that encode essential proteins. For Staphylococcus aureus the level of discrimination provided by MLST is sufficient to provide a relatively detailed picture of the global dissemination of the pathogen. The method is not restrictive in the precise methodology used to acquire the sequences, but the method of assigning types requires that the data be of high quality. Excellent web-based tools have been developed and are curated by the groups that launched MLST. These tools have allowed the scheme to be maintained as a coherent global asset and assist users in the analysis of their data.

Validation of an improved pediatric weight estimation strategy.

OBJECTIVES: To validate the recently described Mercy method for weight estimation in an independent cohort of children living in the United States. METHODS: Anthropometric data including weight, height, humeral length, and mid upper arm circumference were collected from 976 otherwise healthy children (2 months to 14 years old). The data were used to examine the predictive performances of the Mercy method and four other weight estimation strategies (the Advanced Pediatric Life Support [APLS] method, the Broselow tape, and the Luscombe and Owens and the Nelson methods). RESULTS: THE MERCY METHOD DEMONSTRATED ACCURACY COMPARABLE TO THAT OBSERVED IN THE ORIGINAL STUDY (MEAN ERROR: -0.3 kg; mean percentage error: -0.3%; root mean square error: 2.62 kg; 95% limits of agreement: 0.83-1.19). This method estimated weight within 20% of actual for 95% of children compared with 58.7% for APLS, 78% for Broselow, 54.4% for Luscombe and Owens, and 70.4% for Nelson. Furthermore, the Mercy method was the only weight estimation strategy which enabled prediction of weight in all of the children enrolled. CONCLUSIONS: The Mercy method proved to be highly accurate and more robust than existing weight estimation strategies across a wider range of age and body mass index values, thereby making it superior to other existing approaches.

Molecular longitudinal tracking of Mycobacterium abscessus spp. during chronic infection of the human lung.

The Mycobacterium abscessus complex is an emerging cause of chronic pulmonary infection in patients with underlying lung disease. The M. abscessus complex is regarded as an environmental pathogen but its molecular adaptation to the human lung during long-term infection is poorly understood. Here we carried out a longitudinal molecular epidemiological analysis of 178 M. abscessus spp. isolates obtained from 10 cystic fibrosis (CF) and 2 non CF patients over a 13 year period. Multi-locus sequence and molecular typing analysis revealed that 11 of 12 patients were persistently colonized with the same genotype during the course of the infection while replacement of a M. abscessus sensu stricto strain with a Mycobacterium massiliense strain was observed for a single patient. Of note, several patients including a pair of siblings were colonized with closely-related strains consistent with intra-familial transmission or a common infection reservoir. In general, a switch from smooth to rough colony morphology was observed during the course of long-term infection, which in some cases correlated with an increasing severity of clinical symptoms. To examine evolution during long-term infection of the CF lung we compared the genome sequences of 6 sequential isolates of Mycobacterium bolletii obtained from a single patient over an 11 year period, revealing a heterogeneous clonal infecting population with mutations in regulators controlling the expression of virulence factors and complex lipids. Taken together, these data provide new insights into the epidemiology of M. abscessus spp. during long-term infection of the CF lung, and the molecular transition from saprophytic organism to human pathogen.

Multiplex real-time PCR assay for the detection of meticillin-resistant Staphylococcus aureus and Panton-Valentine leukocidin from clinical samples.

The control and prevention of meticillin-resistant Staphylococcus aureus (MRSA) is a major challenge for healthcare establishments, especially as this pathogen continues to evolve. The emergence and spread of community associated MRSA producing Panton-Valentine leukocidin (PVL) causing severe, sometimes fatal, infections in otherwise healthy people is a significant cause of concern. Patient screening to detect MRSA is now widely used as part of an effective control program to limit the spread of this pathogen. Real-time PCR targeting specific MRSA markers offers a rapid alternative to conventional methods enabling earlier intervention, such as patient isolation and decolonization treatment. Herein we describe a multiplex real-time assay that combines primers and probes to detect MRSA and the genes for PVL to provide a rapid and informative assay.