Evaluating next-generation sequencing for direct clinical diagnostics in diarrhoeal disease.

The accurate microbiological diagnosis of diarrhoea involves numerous laboratory tests and, often, the pathogen is not identified in time to guide clinical management. With next-generation sequencing (NGS) becoming cheaper, it has huge potential in routine diagnostics. The aim of this study was to evaluate the potential of NGS-based diagnostics through direct sequencing of faecal samples. Fifty-eight clinical faecal samples were obtained from patients with diarrhoea as part of the routine diagnostics at Hvidovre University Hospital, Denmark. Ten samples from healthy individuals were also included. DNA was extracted from faecal samples and sequenced on the Illumina MiSeq system. Species distribution was determined with MGmapper and NGS-based diagnostic prediction was performed based on the relative abundance of pathogenic bacteria and Giardia and detection of pathogen-specific virulence genes. NGS-based diagnostic results were compared to conventional findings for 55 of the diarrhoeal samples; 38 conventionally positive for bacterial pathogens, two positive for Giardia, four positive for virus and 11 conventionally negative. The NGS-based approach enabled detection of the same bacterial pathogens as the classical approach in 34 of the 38 conventionally positive bacterial samples and predicted the responsible pathogens in five of the 11 conventionally negative samples. Overall, the NGS-based approach enabled pathogen detection comparable to conventional diagnostics and the approach has potential to be extended for the detection of all pathogens. At present, however, this approach is too expensive and time-consuming for routine diagnostics.

The epidemiology of irritable bowel syndrome: Symptom development over a 3-year period in Denmark. A prospective, population-based cohort study.

BACKGROUND: We aimed to explore the natural history of irritable bowel syndrome (IBS) in Denmark over 3 years by studying development of IBS symptoms and associated factors. METHODS: A cohort study was carried out using a web panel representative of the Danish general population 18-50 years. The survey, including a questionnaire based on the Rome III criteria for IBS, was conducted in January 2010, January 2011, and March 2013. KEY RESULTS: The prevalence of IBS was 15.4% (920/5986). The incidence was 10.3%, and was three times higher for persons with unspecific gastrointestinal (GI) symptoms compared to asymptomatic persons. Of respondents with IBS symptoms in both 2010 and 2011, 69% (131/191) also reported symptoms of IBS in 2013, which was significantly more compared to respondents with IBS symptoms in 2010 reporting to be asymptomatic or having unspecific GI symptoms in 2011 (20% and 39%, respectively, P<.001). Being diagnosed with IBS predicted fulfilling the criteria for IBS 3 years later (OR: 2.59, 95% CI: 1.11-6.10). Fulfilling criteria for IBS after 1 year also led to a high risk of IBS symptoms 3 years later in asymptomatic persons and persons with unspecific symptoms at baseline. CONCLUSIONS & INFERENCES: The vast majority of persons fulfilling criteria for IBS report GI symptoms after one and 3 years. Fulfilling IBS criteria after 1 year led to a high risk of reporting IBS symptoms after 3 years. In the general population having an IBS diagnosis predicts persistently fulfilling the Rome III criteria for IBS 3 years later.

The role of Coxsackievirus A16 in a case of sudden unexplained death in an infant - A SUDI case.

The Coxsackievirus A16 (CV-A16) is one of the main pathogens causing hand-foot-and-mouth disease in young children. It is a low-virulence virus rarely involved in serious illness. It is seen sporadically or in outbreaks all over the world. We report a case of sudden unexplained death in infancy, SUDI, in a 3 and 1/2 months old infant, in which a thorough post mortem investigation pointed at a fatal infection with CV-A16 as the most likely cause of death. Only five cases of fatal CV-A16 infection have been published and none of these presented as sudden death. The fatal cases involved two infants, two young children and an elderly man. Post mortem, pre-autopsy CT-scan and C-reactive protein analysis allowed for an autopsy procedure targeted at a microbiological cause of death. The case illustrates the usefulness of supplementary testing during autopsy.

The Rome II and Rome III criteria identify the same subtype-populations in irritable bowel syndrome: agreement depends on the method used for symptom report.

BACKGROUND: For comparing trials using different classifications for irritable bowel syndrome (IBS) subtypes, it is important to know whether these identify the same sub-populations. Our aim was to determine the agreement between Rome II and Rome III subtypes, and to explore whether agreement depends on the symptom reporting method. METHODS: Rome II IBS patients from two identical, randomized placebo-controlled trials of probiotics were included. Retrospective subtypes were based on the Rome II questionnaire. Prospective subtypes were based on diary cards for 2 weeks of run-in. Agreement was determined between: (i) retrospective Rome II and Rome III, (ii) prospective Rome II and Rome III, (iii) retrospective Rome II and prospectively Rome III, (iv) retrospective and prospective Rome II, and (v) retrospective and prospective Rome III. KEY RESULTS: A total of 126 patients, 72% women, mean age 46 ± 15 years, were included. The agreement between subtypes using the same symptom reporting method was: (i) 90.3% (κ = 0.85) for retrospective subtypes, and (ii) 84% (κ = 0.76) for prospective subtypes. The agreement between subtypes using different symptom reporting methods was, (iii) 49% (κ = 0.23) for retrospective Rome II and prospective Rome III, (iv) 51% (κ = 0.26) for Rome II subtypes, and (v) 41% (κ = 0.25) for Rome III subtypes. CONCLUSIONS & INFERENCES: Agreement between Rome II and Rome III subtypes is good to very good when using the same symptom reporting method. When mixing methods, agreement is only fair even within the same classification. This has implications for comparison of trials using different symptom reporting methods for subtyping.

Short-term stability of subtypes in the irritable bowel syndrome: prospective evaluation using the Rome III classification.

BACKGROUND: In irritable bowel syndrome (IBS) subtyping is used in research and clinical practice. Knowledge of subtype stability is needed for proper design of trials and treatment strategies. AIMS: To evaluate the stability of Rome III IBS subtypes over time and to determine the optimal time period for prospective, diary-based subtyping. METHODS: Rome III IBS patients aged 18-70 years enrolled in two identical, randomised, placebo-controlled trials of probiotics, were included. No difference was found on stool pattern, thus patients were analysed as one group. Patients scored defaecations according to Bristol Stool Form Scale for 10 weeks. IBS subtypes were determined for all 1- and 2-week periods. Subtype distribution and stool pattern over time were determined. The proportions of patients having the same subtype all weeks (stable patients) or having a predominant subtype (same subtype ≥60% of time) were determined. RESULTS: A total of 126 patients, mean age 46 ± 15 years, 72% women were included. Subtype distribution was similar over time with IBS with constipation, IBS with diarrhoea and IBS unsubtyped constituting one-third of the population each. Even though only 18-35% had the same subtype all weeks, the majority of patients had the same subtype for ≥60% of time (82-98%). Sixty-nine per cent had the same predominant and baseline subtypes. Two-week data increased the proportion of stable patients, of patients with a predominant subtype, and of patients who had similar baseline and predominant subtype. CONCLUSIONS: Most IBS patients change subtype over time. However, an underlying stool pattern stability was demonstrated in the majority of patients. To increase stability, we recommend 2-week data for IBS subtyping.