Viral pneumonia is an important cause of death and morbidity among infants worldwide. Transmission of non-influenza respiratory viruses in households can inform preventative interventions and has not been well-characterised in South Asia. From April 2011 to April 2012, household members of pregnant women enrolled in a randomised trial of influenza vaccine in rural Nepal were surveyed weekly for respiratory illness until 180 days after birth. Nasal swabs were tested by polymerase chain reaction for respiratory viruses in symptomatic individuals. A transmission event was defined as a secondary case of the same virus within 14 days of initial infection within a household. From 555 households, 825 initial viral illness episodes occurred, resulting in 79 transmission events. The overall incidence of transmission was 1.14 events per 100 person-weeks. Risk of transmission incidence was associated with an index case age 1–4 years (incidence rate ratio (IRR) 2.35; 95% confidence interval (CI) 1.40–3.96), coinfection as initial infection (IRR 1.94; 95% CI 1.05–3.61) and no electricity in household (IRR 2.70; 95% CI 1.41–5.00). Preventive interventions targeting preschool-age children in households in resource-limited settings may decrease the risk of transmission to vulnerable household members, such as young infants.

Long-chain n-3 PUFA (LCn-3PUFA) including DHA and EPA, are known to decrease inflammation by inhibiting arachidonic acid (AA) metabolism to eicosanoids, decreasing the production of pro-inflammatory cytokines and reducing immune cell function. The aim of this study was to determine if EPA and DHA reduced the release of inflammatory mediators from airway epithelial cells infected with rhinovirus (RV). Airway epithelial cells (Calu-3) were incubated with EPA, DHA and AA for 24 h, followed by rhinovirus infection for 48 h. IL-6, IL-8 and interferon-γ-induced protein-10 (IP-10) released by cells were measured using ELISA. Viral replication was measured by serial titration assays. The fatty acid content of cells was analysed using GC. Cellular viability was determined by visual inspection of cells and lactate dehydrogenase release. DHA (400 μm) resulted in a significant 16 % reduction in IL-6 release after RV-43 infection, 29 % reduction in IL-6 release after RV-1B infection, 28 % reduction in IP-10 release after RV-43 infection and 23 % reduction in IP-10 release after RV-1B infection. Cellular DHA content negatively correlated with IL-6 and IP-10 release. None of the fatty acids significantly modified rhinovirus replication. DHA supplementation resulted in increased cellular content of DHA at the cost of AA, which may explain the decreased inflammatory response of cells. EPA and AA did not change the release of inflammatory biomarkers significantly. It is concluded that DHA has a potential role in suppressing RV-induced airway inflammation.

The factors that influence the enhanced stability observed experimentally of human rhinovirus 14 (HRV14) upon binding a hydrophobic antiviral drug have been investigated by molecular dynamics. Simulations centered about the HRV14 drug-binding pocket allow the reliable assessment of differences in capsid protein motions of HRV14 and drug-bound HRV14. We propose that the experimentally observed stabilization of the ligated virus arises from higher entropy, rather than enthalpy. Time-averaged interaction energies between the viral protein and molecules occupying the pocket are less favorable in the presence of the drug, consistent with the proposal that the observed stability arises from entropic effects. Interaction energies characterizing subunit–subunit contacts within one viral protomer are found to be substantially stronger than those between two protomers. Such distinction in subunit interaction would have clear implications on assembly and disassembly. Drug binding is found to affect large-scale, collective properties, while leaving local atomic properties unperturbed. Specifically, the simulations reveal a weakening of long-range correlations in atomic motions upon drug binding. On the other hand, neither the fast time scale RMS fluctuations of individual atomic positions nor the fluctuation build-up curves from the capsid β-sandwich forming the drug-binding pocket show a consistent distinction between the drug-bound and drug-free viral simulations. Collectively, the detailed description available from the simulations provides an understanding of the experimental observations on the drug-induced changes in thermal stability and protease sensitivity reported for picornaviruses. The predicted significance of binding entropy can be explored experimentally and should be considered in the design of new antiviral compounds.

The internal ribosome entry segment (IRES) of picornaviruses consists of ∼450 nt of 5′-untranslated region, terminating at the 3′ end with an ∼25 nt element consisting of an absolutely conserved UUUC motif followed by a more variable pyrimidine-rich tract and G-poor spacer, and finally an AUG triplet, which is considered to be the actual ribosome entry site. Events following entry at this site differ among picornaviruses: in encephalomyocarditis virus (EMCV) virtually all ribosomes initiate translation at this site (AUG-11); in foot-and-mouth-disease virus (FMDV), one-third of the ribosomes initiate at this AUG (the Lab site), and the rest at the next AUG 84 nt downstream (Lb site); and in poliovirus (PV), the AUG at the 3′ end of the IRES (at nt 586 in PV type 1) is considered to be a silent entry site, with all ribosomes initiating translation at the next AUG downstream (nt 743). To investigate what determines this different behavior, chimeras were constructed with a crossover at the conserved UUUC motif: the body of the IRES, the sequences upstream of this UUUC motif, was derived from one species, and the downstream sequences from another. When the body of the FMDV or PV IRESes was replaced by that of EMCV, there was a marked increase in the absolute and relative frequency of initiation at the upstream AUG, the Lab site of FMDV and 586AUG of PV, respectively. In contrast, when the body of the EMCV IRES was replaced by that of PV, initiation occurred with no preference at three AUGs: the normal site (AUG-11), AUG-10 situated 8 nt upstream, and AUG-12, which is 12 nt downstream. Thus although the context of the AUG at the 3′ end of the IRES may influence initiation frequency at this site, as was shown by improving the context of 586AUG of PV, the behavior of the ribosome is also highly dependent on the nature of the upstream IRES. Delivery of the ribosome to this AUG in an initiation-competent manner is particularly efficient and accurate with the EMCV IRES.

Cis-acting RNA signals are required for replication of positive-strand viruses such as the picornaviruses. Although these generally have been mapped to the 5′ and/or 3′ termini of the viral genome, RNAs derived from human rhinovirus type 14 are unable to replicate unless they contain an internal cis-acting replication element (cre) located within the genome segment encoding the capsid proteins. Here, we show that the essential cre sequence is 83–96 nt in length and located between nt 2318–2413 of the genome. Using dicistronic RNAs in which translation of the P1 and P2–P3 segments of the polyprotein were functionally dissociated, we further demonstrate that translation of the cre sequence is not required for RNA replication. Thus, although it is located within a protein-coding segment of the genome, the cre functions as an RNA entity. Computer folds suggested that cre sequences could form a stable structure in either positive- or minus-strand RNA. However, an analysis of mutant RNAs containing multiple covariant and non-covariant nucleotide substitutions within these putative structures demonstrated that only the predicted positive-strand structure is essential for efficient RNA replication. The absence of detectable minus-strand synthesis from RNAs that lack the cre suggests that the cre is required for initiation of minus-strand RNA synthesis. Since a lethal 3′ noncoding region mutation could be partially rescued by a compensating mutation within the cre, the cre appears to participate in a long-range RNA–RNA interaction required for this process. These data provide novel insight into the mechanisms of replication of a positive-strand RNA virus, as they define the involvement of an internally located RNA structure in the recognition of viral RNA by the viral replicase complex. Since internally located RNA replication signals have been shown to exist in several other positive-strand RNA virus families, these observations are potentially relevant to a wide array of related viruses.