Coronaviruses are animal and human pathogens that can cause lethal zoonotic infections like SARS and MERS. They have polycistronic plus-stranded RNA genomes and belong to the order Nidovirales, a ...diverse group of viruses for which common ancestry was inferred from the common principles underlying their genome organization and expression, and from the conservation of an array of core replicase domains, including key RNA-synthesizing enzymes. Coronavirus genomes (~26-32 kilobases) are the largest RNA genomes known to date and their expansion was likely enabled by acquiring enzyme functions that counter the commonly high error frequency of viral RNA polymerases. The primary functions that direct coronavirus RNA synthesis and processing reside in nonstructural protein (nsp) 7 to nsp16, which are cleavage products of two large replicase polyproteins translated from the coronavirus genome. Significant progress has now been made regarding their structural and functional characterization, stimulated by technical advances like improved methods for bioinformatics and structural biology, in vitro enzyme characterization, and site-directed mutagenesis of coronavirus genomes. Coronavirus replicase functions include more or less universal activities of plus-stranded RNA viruses, like an RNA polymerase (nsp12) and helicase (nsp13), but also a number of rare or even unique domains involved in mRNA capping (nsp14, nsp16) and fidelity control (nsp14). Several smaller subunits (nsp7-nsp10) act as crucial cofactors of these enzymes and contribute to the emerging "nsp interactome." Understanding the structure, function, and interactions of the RNA-synthesizing machinery of coronaviruses will be key to rationalizing their evolutionary success and the development of improved control strategies.
This review focuses on the monophyletic group of animal RNA viruses united in the order
Nidovirales. The order includes the distantly related coronaviruses, toroviruses, and roniviruses, which ...possess the largest known RNA genomes (from 26 to 32
kb) and will therefore be called ‘large’ nidoviruses in this review. They are compared with their arterivirus cousins, which also belong to the
Nidovirales despite having a much smaller genome (13–16
kb). Common and unique features that have been identified for either large or all nidoviruses are outlined. These include the nidovirus genetic plan and genome diversity, the composition of the replicase machinery and virus particles, virus-specific accessory genes, the mechanisms of RNA and protein synthesis, and the origin and evolution of nidoviruses with small and large genomes. Nidoviruses employ single-stranded, polycistronic RNA genomes of positive polarity that direct the synthesis of the subunits of the replicative complex, including the RNA-dependent RNA polymerase and helicase. Replicase gene expression is under the principal control of a ribosomal frameshifting signal and a chymotrypsin-like protease, which is assisted by one or more papain-like proteases. A nested set of subgenomic RNAs is synthesized to express the 3′-proximal ORFs that encode most conserved structural proteins and, in some large nidoviruses, also diverse accessory proteins that may promote virus adaptation to specific hosts. The replicase machinery includes a set of RNA-processing enzymes some of which are unique for either all or large nidoviruses. The acquisition of these enzymes may have improved the low fidelity of RNA replication to allow genome expansion and give rise to the ancestors of small and, subsequently, large nidoviruses.
The genome organization and expression strategy of the newly identified severe acute respiratory syndrome coronavirus (SARS-CoV) were predicted using recently published genome sequences. Fourteen ...putative open reading frames were identified, 12 of which were predicted to be expressed from a nested set of eight subgenomic mRNAs. The synthesis of these mRNAs in SARS-CoV-infected cells was confirmed experimentally. The 4382- and 7073 amino acid residue SARS-CoV replicase polyproteins are predicted to be cleaved into 16 subunits by two viral proteinases (bringing the total number of SARS-CoV proteins to 28). A phylogenetic analysis of the replicase gene, using a distantly related torovirus as an outgroup, demonstrated that, despite a number of unique features, SARS-CoV is most closely related to group 2 coronaviruses. Distant homologs of cellular RNA processing enzymes were identified in group 2 coronaviruses, with four of them being conserved in SARS-CoV. These newly recognized viral enzymes place the mechanism of coronavirus RNA synthesis in a completely new perspective. Furthermore, together with previously described viral enzymes, they will be important targets for the design of antiviral strategies aimed at controlling the further spread of SARS-CoV.
The epidemiology of hepatitis E virus (HEV) is not fully understood. In this study, we assessed putative risk factors for HEV seropositivity in various study populations in the Netherlands.
Data and ...samples from five different study populations were analysed: (A) blood donors (n = 5,239), (B) adults reporting a vegetarian life style since the age of 12 years (n = 231), (C) residents of Amsterdam, the Netherlands, with different ethnic backgrounds (n = 1,198), (D) men who have sex with men (MSM) (HIV positive and HIV negative) (n = 197), and (E) persons who use drugs (PWUD) (HIV positive and HIV negative) (n = 200). Anti-HEV immunoglobulin M (IgM) and immunoglobulin G (IgG) testing was performed using ELISA test (Wantai).
HEV IgM seroprevalence was low across all study populations (<1% to 8%). The age and gender-adjusted HEV IgG seroprevalence was 24% among blood donors (reference group) and 9% among the vegetarian group (adjusted Relative Risk aRR:0.36, 95%CI:0.23-0.57). Among participants of different ethnic backgrounds, the adjusted HEV IgG seroprevalence was 16% among participants with a Dutch origin (aRR:0.64, 95%CI:0.40-1.02), 2% among South-Asian Surinamese (aRR:0.07, 95%CI:0.02-0.29), 3% among African Surinamese (aRR:0.11, 95%CI:0.04-0.34), 34% among Ghanaian (aRR:1.53, 95%CI:1.15-2.03), 19% among Moroccan (aRR:0.75, 95%CI:0.49-1.14), and 5% among Turkish (aRR:0.18, 95%CI:0.08-0.44) origin participants. First generation Moroccans had a higher risk for being IgG HEV seropositive compared to second generation Moroccan migrants. The statistical power to perform these analyses in the other ethnic groups was too low. In the MSM group the IgG HEV seroprevalence was 24% (aRR:0.99, 95%CI:0.76-1.29), and among PWUD it was 28% (aRR:1.19, 95%CI:0.90-1.58). The number of sexual partners in the preceding six months was not significantly associated with IgG HEV seropositivity in MSM. The association between HIV status and HEV seropositivity was significant in PWUD, yet absent in MSM. HIV viral load and CD4 cell count were not associated with HEV seropositivity in HIV positive MSM and PWUD.
Vegetarians were significantly less often HEV seropositive. Ethnic origin influenced the risk for being IgG HEV seropositive. MSM and PWUD were not at higher risk for being IgG HEV seropositive than blood donors.
We observe the unconventional photon blockade effect in quantum dot cavity QED, which, in contrast to the conventional photon blockade, operates in the weak coupling regime. A single quantum dot ...transition is simultaneously coupled to two orthogonally polarized optical cavity modes, and by careful tuning of the input and output state of polarization, the unconventional photon blockade effect is observed. We find a minimum second-order correlation g^{(2)}(0)≈0.37, which corresponds to g^{(2)}(0)≈0.005 when corrected for detector jitter, and observe the expected polarization dependency and photon bunching and antibunching; close by in parameter space, which indicates the abrupt change from phase to amplitude squeezing.
A novel coronavirus is the causative agent of the current epidemic of severe acute respiratory syndrome (SARS). Coronaviruses are exceptionally large RNA viruses and employ complex regulatory ...mechanisms to express their genomes. Here, we determined the sequence of SARS coronavirus (SARS-CoV), isolate Frankfurt 1, and characterized key RNA elements and protein functions involved in viral genome expression. Important regulatory mechanisms, such as the (discontinuous) synthesis of eight subgenomic mRNAs, ribosomal frameshifting and post-translational proteolytic processing, were addressed. Activities of three SARS coronavirus enzymes, the helicase and two cysteine proteinases, which are known to be critically involved in replication, transcription and/or post-translational polyprotein processing, were characterized. The availability of recombinant forms of key replicative enzymes of SARS coronavirus should pave the way for high-throughput screening approaches to identify candidate inhibitors in compound libraries.
Fat distribution as measured by waist-to-hip ratio has been shown to be an important independent predictor of glucose intolerance. Few studies, however, have considered the contributions of the waist ...and hip circumferences independently. The aim of this study was to investigate the independent associations of waist and hip circumference with diabetes in a large population-based study, and to investigate whether they also apply to other major components of the metabolic syndrome (hypertension and dyslipidemia). In addition, as previous studies were performed in older persons, we investigated whether these associations were present across adult age groups.
Weight, height, waist and hip circumferences were measured in 11 247 participants of the nationally representative Australian Diabetes, Obesity and Lifestyle (AusDiab) Study. HDL-cholesterol, triglycerides, fasting and 2-h postload glucose were determined, and diastolic and systolic blood pressure was measured. After exclusion of persons already known to have diabetes, hypertension or dyslipidemia, logistic and linear regression were used to study cross-sectional associations of anthropometric variables with newly diagnosed diabetes, hypertension and dyslipidemia, and with continuous metabolic measures, all separately for men (n=3818) and women (n=4582). Analyses were repeated in the same population stratified for age.
After adjustment for age, body mass index and waist, a larger hip circumference was associated with a lower prevalence of undiagnosed diabetes (odds ratio (OR) per one s.d. increase in hip circumference 0.55 (95% CI 0.41-0.73) in men and 0.42 (0.27-0.65) in women) and undiagnosed dyslipidemia (OR 0.58 (0.50-0.67) in men and 0.37 (0.30-0.45) in women). Associations with undiagnosed hypertension were weaker (OR 0.80 (0.69-0.93) in men and 0.88 (0.70-1.11) in women). As expected, larger waist circumference was associated with higher prevalence of these conditions. Similar associations were found using continuous metabolic variables as outcomes in linear regression analyses. Height partly explained the negative associations with hip circumference. When these analyses were performed stratified for age, associations became weaker or disappeared in the oldest age groups (age > or =75 y in particular), except for HDL-cholesterol.
We found independent and opposite associations of waist and hip circumference with diabetes, dyslipidemia and less strongly with hypertension in a large population-based survey. These results emphasize that waist and hip circumference are important predictors for the metabolic syndrome and should both be considered in epidemiological studies. The associations were consistent in all age groups, except in age > or =75 y. Further research should be aimed at verifying hypotheses explaining the 'protective' effect of larger hips.
The recently identified etiological agent of the severe acute respiratory syndrome (SARS) belongs to Coronaviridae (CoV), a family of viruses replicating by a poorly understood mechanism. Here, we ...report the crystal structure at 2.7-Å resolution of nsp9, a hitherto uncharacterized subunit of the SARS-CoV replicative polyproteins. We show that SARS-CoV nsp9 is a single-stranded RNA-binding protein displaying a previously unreported, oligosaccharide/oligonucleotide fold-like fold. The presence of this type of protein has not been detected in the replicative complexes of RNA viruses, and its presence may reflect the unique and complex CoV viral replication/transcription mechinery.
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