Synchronized long-read genome, methylome, epigenome and transcriptome profiling resolve a Mendelian condition.

Resolving the molecular basis of a Mendelian condition remains challenging owing to the diverse mechanisms by which genetic variants cause disease. To address this, we developed a synchronized long-read genome, methylome, epigenome and transcriptome sequencing approach, which enables accurate single-nucleotide, insertion-deletion and structural variant calling and diploid de novo genome assembly. This permits the simultaneous elucidation of haplotype-resolved CpG methylation, chromatin accessibility and full-length transcript information in a single long-read sequencing run.

Synchronized long-read genome, methylome, epigenome, and transcriptome for resolving a Mendelian condition.

Resolving the molecular basis of a Mendelian condition (MC) remains challenging owing to the diverse mechanisms by which genetic variants cause disease. To address this, we developed a synchronized long-read genome, methylome, epigenome, and transcriptome sequencing approach, which enables accurate single-nucleotide, insertion-deletion, and structural variant calling and diploid genome assembly, and permits the simultaneous elucidation of haplotype-resolved CpG methylation, chromatin accessibility, and full-length transcript information in a single long-read sequencing run. Application of this approach to an Undiagnosed Diseases Network (UDN) participant with a chromosome X;13 balanced translocation of uncertain significance revealed that this translocation disrupted the functioning of four separate genes (, , , and ) previously associated with single-gene MCs.

The Control of Methicillin-Resistant Staphylococcus aureus Blood Stream Infections in England.

Methicillin-resistant Staphylococcus aureus (MRSA) blood stream infection (BSI) is a major healthcare burden in some but not all healthcare settings, and it is associated with 10%-20% mortality. The introduction of mandatory reporting in England of MRSA BSI in 2001 was followed in 2004 by the setting of target reductions for all National Health Service hospitals. The original national target of a 50% reduction in MRSA BSI was considered by many experts to be unattainable, and yet this goal has been far exceeded (∼80% reduction with rates still declining).

Changes to clinician attire have done more harm than good.

The introduction of 'bare below the elbows' policies to facilitate handwashing led to the disappearance of the white coat from medical and surgical wards. While rates of key healthcare acquired infections in hospitals, e.g.

Does ambulance response time influence patient condition among patients with specific medical and trauma emergencies?

Objectives: The relation between patient outcome and ambulance response times is unknown. We sought to measure the influence of shorter response times on patient outcomes. The objective of the study was to determine whether ambulance response time makes a difference in the outcomes of emergency medical services (EMS) patients with specific traumatic and medical emergencies.

MRSA: why have we got it and can we do anything about it?

MRSA, first identified in 1960, became a major cause of healthcare-associated infection with the emergence of epidemic strains EMRSA 15 and 16 in the 1990 s. MRSA bacteraemia surveillance in England showed a peak of 7700 in 2003-2004. A target was set to halve MRSA bacteraemias by 2008 backed by a central improvement programme for infection prevention and control.

Contribution of a government target to controlling Clostridium difficile in the NHS in England.

The introduction of mandatory surveillance of Clostridium difficile infection (CDI) in 2004 showed the scale of the challenge: cases in patients >64 years old reached 55,681 in 2006. The first type 027 outbreaks had been in 2005 and CDI was a headline issue. The prevention and control of CDI requires a tripartite partnership between clinicians, health service managers, and the government/Department of Health which needs to set standards, ensure that CDI is a priority, set targets and monitor outcome.

Analysis of prevalence, risk factors and molecular epidemiology of Clostridium difficile infection in Kuwait over a 3-year period.

We conducted a prospective study to evaluate the prevalence and epidemiology of CDI in Kuwait government hospitals over a 3-year period, January 2003 to December 2005, to determine the ribotypes responsible for CDI and to estimate the prevalence of ribotype 027. We also conducted a case-control study to identify the risk factors in our patient population. A total of 697 stool samples from patients with suspected CDI were obtained and sent to Anaerobe Reference Laboratory, Faculty of Medicine, Kuwait University for Clostridium difficile toxin detection, culture and PCR ribotyping.

Correlation of multidrug resistance, toxinotypes and PCR ribotypes in Clostridium difficile isolates from Kuwait.

Clostridium difficile is a common cause of nosocomial diarrhea. its role in community-acquired diarrhea is also becoming an important public health concern. Hardly any studies have correlated strain ribotypes, toxinotypes and multidrug resistant (MDR) profiles.

Responsibility for managing healthcare-associated infections: where does the buck stop?

The prevention and control of healthcare-associated infections (HCAIs) requires a tripartite partnership between clinicians and carers, managers and government/Department of Health (DoH) across the whole health and social care community. Mandatory surveillance of meticillin-resistant Staphylococcus aureus bacteraemia and Clostridium difficile infection has shown a significant fall from peak numbers in 2003/04 and 2006, respectively, and there is now a zero tolerance approach to preventable infections and poor practice. Success so far has been based on senior management commitment, enhanced real-time surveillance, implementation of clinical protocols (high impact interventions, prudent prescribing), improved hand hygiene and environmental cleaning, and training and audit, backed up by a heightened performance management focus through targets and legislation (Code of Practice).

Is the war being won?--An expert's view.

With new DH guidance on dealing effectively with Clostridium difficile in healthcare settings recently published (see p40-43), Health Estate Journal spoke to Department of Health inspector of microbiology and infection control Professor Brian Duerden, asking him how much progress has been made in preventing outbreaks and containing spread, what lessons have been learned from some of the more high profile outbreaks, and what innovations, if any, are on the horizon to help eradicate the bacterium in future.

Distribution and antimicrobial susceptibility patterns of Clostridium difficile PCR ribotypes in English hospitals, 2007-08.

A surveillance study designed to provide a representative sample of the strains of Clostridium difficile causing infections in hospitals in England was in operation from April 2007 to the end of March 2008. Six hundred and seventy-seven isolates were obtained from 186 hospitals in the nine geographical regions of England as recognised by the Health Protection Agency's Regional Microbiology Network. Typing studies revealed that PCR ribotype 027 is now the most common strain isolated from symptomatic patients, accounting for over 41.

Infection control measures to limit the spread of Clostridium difficile.

Clostridium difficile-associated diarrhoea (CDAD) presents mainly as a nosocomial infection, usually after antimicrobial therapy. Many outbreaks have been attributed to C. difficile, some due to a new hyper-virulent strain that may cause more severe disease and a worse patient outcome.

Controlling healthcare-associated infections in the NHS.

The prevention and control of healthcare-associated infection (HCAI) is a priority for the NHS. The delivery of national targets for reducing methicillin resistant Staphylococcus aureus bacteraemias and Clostridium difficile infection are supported by enhanced mandatory surveillance through the Health Protection Agency and a Department of Health improvement programme that promotes policies and protocols for enhancing clinical procedures and places infection prevention and control at the centre of clinical and corporate governance. The Health Act 2006 Code of Practice makes such policies and protocols a legal requirement and compliance will be assessed by the Healthcare Commission.

Heat and acid tolerance of Clostridium novyi Type A spores and their survival prior to preparation of heroin for injection.

Clostridium novyi Type A was implicated as a cause of an outbreak of serious illness and deaths among drug users in the United Kingdom who injected heroin intramuscularly. A contaminated batch of heroin was believed to be the source of infection. To test the ability of the outbreak strain to survive certain processes associated with heroin use, it was tested for its ability to survive a range of temperature and pH and the process used in preparation of "street" heroin for injection.

Molecular investigation of genetic elements contributing to metronidazole resistance in Bacteroides strains.

Objectives: The aim of this study was to investigate the constitution of nim gene types, their activating insertion sequence (IS) element, their localization (plasmid or chromosome) and cfiA gene status in metronidazole-resistant Bacteroides strains (n=26) in order to examine their interchangeability.

Methods: Southern hybridization and conjugative plasmid transfer were used to localize the nimA-E genes and plasmid functions. PCR was used to detect the IS elements and the cfiA genes.

Twenty-first-century medical microbiology services in the UK.

With infection once again a high priority for the UK National Health Service (NHS), the medical microbiology and infection-control services require increased technology resources and more multidisciplinary staff. Clinical care and health protection need a coordinated network of microbiology services working to consistent standards, provided locally by NHS Trusts and supported by the regional expertise and national reference laboratories of the new Health Protection Agency. Here, I outline my thoughts on the need for these new resources and the ways in which clinical microbiology services in the UK can best meet the demands of the twenty-first century.

Soft tissue infections caused by spore-forming bacteria in injecting drug users in the United Kingdom.

From 2000 to May 2004 there has been a marked increase in illness resulting from spore-forming bacteria in injecting heroin users in the United Kingdom. Clostridium novyi caused 63 cases of severe illness in 2000 and seven further cases from 2001. Wound botulism first occurred in 2000 (six cases) with 51 further cases to March 2004.

Extensive DNA inversions in the B. fragilis genome control variable gene expression.

The obligately anaerobic bacterium Bacteroides fragilis, an opportunistic pathogen and inhabitant of the normal human colonic microbiota, exhibits considerable within-strain phase and antigenic variation of surface components. The complete genome sequence has revealed an unusual breadth (in number and in effect) of DNA inversion events that potentially control expression of many different components, including surface and secreted components, regulatory molecules, and restriction-modification proteins. Invertible promoters of two different types (12 group 1 and 11 group 2) were identified.

Actinomyces dentalis sp. nov., from a human dental abscess.

A previously undescribed filamentous, beaded, Gram-positive, rod-shaped bacterium was isolated from pus of a human dental abscess. Based on its cellular morphology and the results of biochemical testing the organism was tentatively identified as a member of the genus Actinomyces, but it did not correspond to any currently recognized species of this genus. Comparative 16S rRNA gene sequencing studies showed the bacterium represents a distinct subline within the genus Actinomyces, clustering within a group of species that includes Actinomyces bovis, the type species of the genus.