Phylodynamic reconstruction reveals norovirus GII.4 epidemic expansions and their molecular determinants.

Noroviruses are the most common cause of viral gastroenteritis. An increase in the number of globally reported norovirus outbreaks was seen the past decade, especially for outbreaks caused by successive genogroup II genotype 4 (GII.4) variants. Whether this observed increase was due to an upswing in the number of infections, or to a surveillance artifact caused by heightened awareness and concomitant improved reporting, remained unclear. Therefore, we set out to study the population structure and changes thereof of GII.4 strains detected through systematic outbreak surveillance since the early 1990s. We collected 1383 partial polymerase and 194 full capsid GII.4 sequences. A Bayesian MCMC coalescent analysis revealed an increase in the number of GII.4 infections during the last decade. The GII.4 strains included in our analyses evolved at a rate of 4.3-9.0x10(-3) mutations per site per year, and share a most recent common ancestor in the early 1980s. Determinants of adaptation in the capsid protein were studied using different maximum likelihood approaches to identify sites subject to diversifying or directional selection and sites that co-evolved. While a number of the computationally determined adaptively evolving sites were on the surface of the capsid and possible subject to immune selection, we also detected sites that were subject to constrained or compensatory evolution due to secondary RNA structures, relevant in virus-replication. We highlight codons that may prove useful in identifying emerging novel variants, and, using these, indicate that the novel 2008 variant is more likely to cause a future epidemic than the 2007 variant. While norovirus infections are generally mild and self-limiting, more severe outcomes of infection frequently occur in elderly and immunocompromized people, and no treatment is available. The observed pattern of continually emerging novel variants of GII.4, causing elevated numbers of infections, is therefore a cause for concern.

Norovirus disease associated with excess mortality and use of statins: a retrospective cohort study of an outbreak following a pilgrimage to Lourdes.

Although norovirus infection is generally known to be a mild disease, there is some evidence for severe outcome. An outbreak in a Dutch psychiatric institution, originating from pilgrims returning from Lourdes (France), provided an opportunity for performing a retrospective cohort study in order to identify risk factors for norovirus disease and excess mortality. Relative risks (RR) including 95% confidence intervals (CI) showed that attending the pilgrimage (RR 2·0, 95% CI 1·4-3·0) and age >70 (RR 1·7, 95% CI 1·2-2·2) were risk factors for symptomatic infection. In a subset of patients, for whom more detailed information was available, the use of statins was associated with norovirus disease when adjusted for underlying condition (adjusted odds ratio 3·9, 95% CI 1·2-13·0). Mortality was higher in cases infected during the pilgrimage compared to other residents (RR 20·9, 95% CI 4·7-93·8). Norovirus disease can lead to severe outcome. The newly identified risk of statins for contracting norovirus disease may have considerable consequences for the Western world and needs prospective confirmation.

Chronic shedders as reservoir for nosocomial transmission of norovirus.

Norovirus (NoV) infection in immunocompromised patients may lead to prolonged norovirus shedding. Here, we demonstrate the involvement of three chronic shedders in hospital outbreaks. Combined epidemiological and molecular evidence suggests that in one case, NoV transmission occurred at least 17 days after the first diagnosis.

Genetic analysis of the capsid gene of genotype GII.2 noroviruses.

Noroviruses (NoVs) are considered to be a major cause of acute nonbacterial gastroenteritis in humans. The NoV genus is genetically diverse, and genotype GII.4 has been most commonly identified worldwide in recent years. In this study we analyzed the complete capsid gene of NoV strains belonging to the less prevalent genotype GII.2. We compared a total of 36 complete capsid sequences of GII.2 sequences obtained from the GenBank (n = 5) and from outbreaks or sporadic cases that occurred in The Netherlands (n = 10) and in Osaka City, Japan (n = 21), between 1976 and 2005. Alignment of all capsid sequences did not show fixation of amino acid substitutions over time as an indication for genetic drift. In contrast, when strains previously recognized as recombinants were excluded from the alignment, genetic drift was observed. Substitutions were found at five informative sites (two in the P1 subdomain and three in the P2 subdomain), segregating strains into five genetic groups (1994 to 1997, 1999 to 2000, 2001 to 2003, 2004, and 2005). Only one amino acid position changed consistently between each group (position 345). Homology modeling of the GII.2 capsid protein showed that the five amino acids were located on the surface of the capsid and close to each other at the interface of two monomers. The data suggest that these changes were induced by selective pressure, driving virus evolution. Remarkably, this was observed only for nonrecombinant genomes, suggesting differences in behavior with recombinant strains.

A prospective cohort study comparing the VAS spine score and Roland-Morris disability questionnaire in patients with a type A traumatic thoracolumbar spinal fracture.

The Roland Morris Disability Questionnaire (RMDQ-24) and the VAS spine score have been regularly used to measure functional outcome in patients with back pain. The RMDQ-24 is primarily used in degenerative disease of the spine and the VAS Spine is used in trauma patients. The aim of this study is to compare these scores and to see if there is a correlation in patients with a traumatic thoracolumbar spinal fracture. Prospective cohort study comparing the RMDQ-24 and the VAS spine score in patients with a traumatic type A fracture thoracolumbar spine fracture. Fifteen non-operatively patients (group one) completed 118 questionnaires and 17 operatively treated patients (group two) completed 140 questionnaires. Group one scored an average of 6.6 and 65.9 for the RMDQ-24 and VAS Spine, in group two this was 5.1 and 82.9. Spearman's correlation test showed a significant correlation, in group one 0.83 and for the second group 0.87. RMDQ-24 and VAS Spine have a strong positive correlation in measuring disability in a group of patients with back pain because of a spinal fracture. In both non-operatively and operatively treated groups this correlation is significant.

Import of norovirus infections in the Netherlands and Ireland following pilgrimages to Lourdes, 2008--preliminary report.

Between mid-September and 19 October 2008, nine clusters of norovirus infection involving around 90 primary cases and over a hundred secondary cases were identified in patients from the Netherlands, Ireland, Italy and France, linked to pilgrimage to Lourdes, France.

[Surgical treatment of lung cancer with complete VATS lobectomy, new in the Netherlands]. / De chirurgische behandeling van longkanker via complete VATS-lobectomie, nieuw in Nederland.

* In view of the favourable results of lobectomy with complete 'video-assisted thoracoscopic surgery' (c-VATS) as described in literature, we started to perform these totally endoscopic lung resections in the Netherlands in March 2006. * Essential aspects of the procedure are that the ribs are not spread and that the procedure is performed using the image on the monitor only. * In the 23 c-VATS lobectomies performed, the advantages described in literature were confirmed. The operation was safe and entirely similar to the 'open' thoracotomy in an oncological sense. The effects for the patient were favourable. There was less pain after the operation, the function of the shoulder remained intact and the patients recovered rapidly. * It takes a considerable time before the learning curve reaches the plateau phase. The use of proper instruments is essential. * It is expected that the c-VATS lobectomy will be performed more often in the Netherlands in the coming years.

[Cardiac symptoms due to pectus excavatum in a man over the age of 55]. / Cardiale klachten door een pectus excavatum bij een 55-plusser.

A healthy 59-year-old man, a retired general practitioner, suffered from increasing palpitations, fatigue and postural dyspnoea: bending over led to a significant increase in his shortness of breath. Cardiological and pulmonological examination, performed at regular intervals, showed occasional supraventricular arrhythmia and nodal tachycardia but did not yield a satisfactory explanation for the symptoms. In the years that followed, the physical impairment became a considerable handicap. Finally, the patient himself suggested a possible explanation on the basis of an Internet search: his pectus excavatum. A literature search confirmed this hypothesis. A lateral chest X-ray in bending position and a CT-scan of the chest revealed compression of the heart by the sternum. Ten years after the onset of symptoms, a modified Ravitch operation finally brought nearly complete recovery.

[Trauma during pregnancy]. / Trauma tijdens de zwangerschap.

Mortality due to trauma in pregnancy is not very common in the Netherlands. More often a pregnant woman presents herself for examination after trauma. Blunt trauma is more common in the third trimester. Minor trauma also needs good care, with special attention for solutio placentae. Maternal mortality after penetrating trauma is low because of the protection of vital organs by the uterus. With good treatment the mortality in pregnant trauma patients will not be higher than in nonpregnant patients. A rapid and effective resuscitation of the mother will give the foetus the best chance of survival.

[The body-packer syndrome]. / Het 'body-packer'-syndroom.

In four patients, a woman of 35 and men of 27, 38, and 22 years old, body-packer syndrome was diagnosed. Body-packer syndrome is seen in people concealing drugs in special containers within the body; this may lead to rupture with acute intoxication or to ileus. The clinical presentation can be very deceptive. An abdominal X-ray often reveals the packages. If there are no symptoms treatment is with mild laxatives, acute intoxication requires immediate laparotomy. One of the four patients died, the others recovered. Legally (Dutch law), the physician best delivers the drug to the police as a lost object, without revealing the patient's identity.

Radionuclide-guided biopsy of bone lesions in cancer patients; a reliable, well-tolerated technique.

OBJECTIVES: In follow up or primary analysis of malignant disease, use of nuclear medicine imaging techniques is increasing. The lesions, detected by these modalities, do not always present with a matching radiological abnormality suitable for radiological biopsy. A method to confirm, or rule out, metastatic disease in these cases is radionuclide-guided surgical biopsy. PATIENTS AND METHODS: The technique consists of injecting a Technetium-99m labelled diphosphonate, performing a bone scan and marking the lesion; under general anaesthetic a surgical biopsy is taken through a small incision guided by a gamma-probe. We evaluated the combined experience of two teaching hospitals, by conducting retrospective chart review, over a period of 5.5 years. Sixty-nine procedures with 79 biopsies were performed. The patients previously had breast cancer in 42%, lung cancer in 32%, both gastrointestinal and urological malignancies in 13%, miscellaneous or unknown origin were reported in 9%. In 68% a rib biopsy was taken, in 15% a pelvis biopsy and in 11% a sternum or clavicle biopsy. RESULTS: In 46.8% metastatic disease was confirmed and in 2.6% a new malignancy (M Kahler) was identified. During a median clinical follow up period of 31 months (1-70), 1 initially negative biopsy site was found to contain a metastasis. Apart from local pain in 3% of cases, no complications were mentioned. CONCLUSION: In conclusion we can state that, in this retrospective series, radionuclide-guided bone biopsy is a reliable, well-tolerated technique to determine the diagnosis of these lesions with a sensitivity of 97% and specificity of 100%.

Symptomatic pectus excavatum in seniors (SPES): a cardiovascular problem? : A prospective cardiological study of 42 senior patients with a symptomatic pectus excavatum.

OBJECTIVES: There is no consensus among physicians as to whether or not pectus excavatum can produce symptoms sometimes even severe enough to justify a surgical procedure. The aim of this study was to assess the prevalence and severity of complaints and symptoms of senior patients with a pectus excavatum and to evaluate the results of surgical correction of the chest deformation. DESIGN: This is a prospective clinical study, case series. PARTICIPANTS: The participants are 42 senior patients with a pectus excavatum and somatic complaints. METHODS: Cardiological screening included medical history taking, physical examination, electrocardiography, transthoracic echocardiography and treadmill cardiography. Complaints, symptoms and test results were arranged in a clinical score. Indication for a therapeutic surgical correction by a modified Ravitch operation was a high clinical score in combination with radiological evidence of cardiac compression on CT or MRI. RESULTS: The clinical picture of the 42 senior patients with a pectus excavatum showed complaints of fatigue and low exercise tolerance, shortness of breath, palpitations, inspiratory obstruction and sometimes chest discomfort or pain. The serious and sometimes invalidating complaints of 19 patients (45%) had started only in their fourth or fifth decade of life and were labelled in 12 patients (63%) as 'Unexplained cardiovascular complaints'. To date, 11 patients have undergone surgical procedures. Symptoms were reduced substantially or had disappeared completely. All patients indicated that their health-related quality of life was significantly improved. CONCLUSION: Recognising the clinical picture of SPES is relevant because surgical reconstruction of the chest can provide substantial relief of symptoms.

Norovirus illness is a global problem: emergence and spread of norovirus GII.4 variants, 2001-2007.

BACKGROUND: Noroviruses (NoVs) are the most common cause of viral gastroenteritis. Their high incidence and importance in health care facilities result in a great impact on public health. Studies from around the world describing increasing prevalence have been difficult to compare because of differing nomenclatures for variants of the dominant genotype, GII.4. We studied the global patterns of GII.4 epidemiology in relation to its genetic diversity. METHODS: Data from NoV outbreaks with dates of onset from January 2001 through March 2007 were collected from 15 institutions on 5 continents. Partial genome sequences (n=775) were collected, allowing phylogenetic comparison of data from different countries. RESULTS: The 15 institutions reported 3098 GII.4 outbreaks, 62% of all reported NoV outbreaks. Eight GII.4 variants were identified. Four had a global distribution--the 1996, 2002, 2004, and 2006b variants. The 2003Asia and 2006a variants caused epidemics, but they were geographically limited. Finally, the 2001 Japan and 2001 Henry variants were found across the world but at low frequencies. CONCLUSIONS: NoV epidemics resulted from the global spread of GII.4 strains that evolved under the influence of population immunity. Lineages show notable (and currently unexplained) differences in geographic prevalence. Establishing a global NoV network by which data on strains with the potential to cause pandemics can be rapidly exchanged may lead to improved prevention and intervention strategies.

High prevalence of prolonged norovirus shedding and illness among hospitalized patients: a model for in vivo molecular evolution.

During a 2-year survey in an academic hospital, 8 (8.4%) of all norovirus (NoV)-positive patients showed prolonged norovirus illness and shedding (duration, 21-182 days). All patients had underlying illnesses, resulting in some level of immunodeficiency in 5. Four patients were admitted to the hospital with gastroenteritis, 2 acquired norovirus while hospitalized, and 2 were outpatients. Genotypes GII.4 and GIIb-GII.3 were found. Reinfection occurred in 3 patients. Full capsid sequences were determined from strains detected in sequentially collected stool specimens to study evolution. The greatest number of amino acid mutations in a given patient was 11; they were detected in NoV isolates recovered over a 119-day period and were mapped to positions at or near putative antigenic sites. In the patient with most severe immune dysfunction, only 5 amino acids mutated over 182 days, suggesting immune-driven selection. The severe impact on patients and hospitals and the potential role of prolonged shedders as a reservoir for viral antigenic variants lead us to stress the importance of confinement of outbreaks of NoV infection that occur in hospitals.

Gastroenteritis caused by norovirus GGII.4, The Netherlands, 1994-2005.

From 1994 through 2005, gastroenteritis outbreaks caused by norovirus generally increased in the Netherlands, with 3 epidemic seasons associated with new GGII.4 strains. Increased percentages of GGII.4 strains during these epidemics, followed by a sharp decrease in their absolute and relative numbers, suggest development of immunity.

Epochal evolution of GGII.4 norovirus capsid proteins from 1995 to 2006.

Noroviruses are the causative agents of the majority of viral gastroenteritis outbreaks in humans. During the past 15 years, noroviruses of genotype GGII.4 have caused four epidemic seasons of viral gastroenteritis, during which four novel variants (termed epidemic variants) emerged and displaced the resident viruses. In order to understand the mechanisms and biological advantages of these epidemic variants, we studied the genetic changes in the capsid proteins of GGII.4 strains over this period. A representative sample was drawn from 574 GGII.4 outbreak strains collected over 15 years of systematic surveillance in The Netherlands, and capsid genes were sequenced for a total of 26 strains. The three-dimensional structure was predicted by homology modeling, using the Norwalk virus (Hu/NoV/GGI.1/Norwalk/1968/US) capsid as a reference. The highly significant preferential accumulation and fixation of mutations (nucleotide and amino acid) in the protruding part of the capsid protein provided strong evidence for the occurrence of genetic drift and selection. Although subsequent new epidemic variants differed by up to 25 amino acid mutations, consistent changes were observed in only five positions. Phylogenetic analyses showed that each variant descended from its chronologic predecessor, with the exception of the 2006b variant, which is more closely related to the 2002 variant than to the 2004 variant. The consistent association between the observed genetic findings and changes in epidemiology leads to the conclusion that population immunity plays a role in the epochal evolution of GGII.4 norovirus strains.

Molecular epidemiology of white spot syndrome virus within Vietnam.

White spot syndrome virus (WSSV), the sole member of the virus family Nimaviridae, is a large double-stranded DNA virus that infects shrimp and other crustaceans. By alignment of three completely sequenced isolates originating from Taiwan (WSSV-TW), China (WSSV-CN) and Thailand (WSSV-TH), the variable loci in the genome were mapped. The variation suggests the spread of WSSV from a common ancestor originating from either side of the Taiwan Strait to Thailand, but support for this hypothesis through analysis of geographical intermediates is sought. RFLP analysis of eight Vietnamese WSSV isolates, of which six were collected along the central coast (VN-central) and two along the south coast (VN-south), showed apparent sequence variation in the variable loci identified previously. These loci were characterized in detail by PCR amplification, cloning and sequencing. Relative to WSSV-TW, all VN-central isolates showed a approximately 8.5 kb deletion in the major variable region ORF23/24, whereas the VN-south isolates contain a deletion of approximately 11.5 or approximately 12.2 kb, compared to a approximately 1.2 or approximately 13.2 kb deletion in WSSV-CN and WSSV-TH, respectively. The minor variable region ORF14/15 showed deletions of various sizes compared with WSSV-TH for all eight VN isolates. The data suggest that the VN isolates and WSSV-TH have a common lineage, which branched off from WSSV-TW and WSSV-CN early on, and that WSSV entered Vietnam by multiple introductions. A model is presented for the spread of WSSV from either side of the Taiwan Strait into Vietnam based on the gradually increasing deletions of both 'variable regions'. The number and order of repeat units within ORF75 and ORF125 appeared to be suitable markers to study regional spread of WSSV.