Sequential conformational changes in the morbillivirus attachment protein initiate the membrane fusion process.

Despite large vaccination campaigns, measles virus (MeV) and canine distemper virus (CDV) cause major morbidity and mortality in humans and animals, respectively. The MeV and CDV cell entry system relies on two interacting envelope glycoproteins: the attachment protein (H), consisting of stalk and head domains, co-operates with the fusion protein (F) to mediate membrane fusion. However, how receptor-binding by the H-protein leads to F-triggering is not fully understood. Here, we report that an anti-CDV-H monoclonal antibody (mAb-1347), which targets the linear H-stalk segment 126-133, potently inhibits membrane fusion without interfering with H receptor-binding or F-interaction. Rather, mAb-1347 blocked the F-triggering function of H-proteins regardless of the presence or absence of the head domains. Remarkably, mAb-1347 binding to headless CDV H, as well as standard and engineered bioactive stalk-elongated CDV H-constructs treated with cells expressing the SLAM receptor, was enhanced. Despite proper cell surface expression, fusion promotion by most H-stalk mutants harboring alanine substitutions in the 126-138 "spacer" section was substantially impaired, consistent with deficient receptor-induced mAb-1347 binding enhancement. However, a previously reported F-triggering defective H-I98A variant still exhibited the receptor-induced "head-stalk" rearrangement. Collectively, our data spotlight a distinct mechanism for morbillivirus membrane fusion activation: prior to receptor contact, at least one of the morbillivirus H-head domains interacts with the membrane-distal "spacer" domain in the H-stalk, leaving the F-binding site located further membrane-proximal in the stalk fully accessible. This "head-to-spacer" interaction conformationally stabilizes H in an auto-repressed state, which enables intracellular H-stalk/F engagement while preventing the inherent H-stalk's bioactivity that may prematurely activate F. Receptor-contact disrupts the "head-to-spacer" interaction, which subsequently "unlocks" the stalk, allowing it to rearrange and trigger F. Overall, our study reveals essential mechanistic requirements governing the activation of the morbillivirus membrane fusion cascade and spotlights the H-stalk "spacer" microdomain as a possible drug target for antiviral therapy.

Enantiomeric 3-deaza-1',6'-isoneplanocin and its 3-bromo analogue: Synthesis by the Ullmann reaction and their antiviral properties.

The 1',6'-isomer of neplanocin A possesses biological properties that have not been optimised through rationally conceived analogues. In that direction, this Letter reports the use of the Ullmann reaction to achieve enantiomeric 3-deaza-1',6'-isoneplanocin and 3-bromo-3-deaza-1',6'-isoneplanocin. These four compounds showed significant Ebola activity that is not specifically due to their inhibition of S-adenonosylhomocysteine hydrolase, as might have been expected for 3-deazaadenine carbocyclic nucleosides. For some members of this group, antiviral activity was also found against human cytomegalovirus, hepatitis B, norovirus, and measles.

Determinants of vaccine coverage and timeliness in a northern Pakistani village.

The incidence of vaccine preventable disease in Pakistan remains high despite a long-standing Expanded Program on Immunization (EPI). We describe vaccine completeness, timeliness and determinants of coverage from a remote rural cohort (2012-2014). Vaccination histories were taken from EPI records. Vaccination was complete if all doses were received according to the EPI schedule and timely if doses were not ≥3 days early or ≥ 28 days late. Three models are presented: a multivariable logistic regression of household demographic and socioeconomic factors associated with complete vaccination, a multivariable mixed effects logistic regression assessing whether or not the vaccine was administered late (versus on-time), and a mixed effects multivariable Poisson regression model analysing the interval (in days) between vaccine doses. Of 959 enrolled children with full vaccination histories, 88.2 and 65.1% were fully vaccinated following either the pentavalent or DPT/HBV schedules if measles was excluded; coverage dropped to 50.0 and 27.1% when both doses of measles were included. Sixty-four (6.7%) were unvaccinated. Coverage and timeliness declined with subsequent doses. Migrating into the village after 1995 (95%CI 1.88 to 5.17) was associated with late vaccination. Being male, having an older father, and having parents with at least some formal education reduced the likelihood of a late dose. The interval between doses was consistent at 5 weeks (compared with the 4 weeks recommended by EPI). None of the socio-demographic variables were related to the likelihood of receiving full coverage. Vaccine coverage in Oshikhandass was higher than national averages. Measles vaccine coverage and timeliness were low; special consideration should be paid to this vaccine. The local vaccination schedule differed from the EPI, but the consistency suggests good local administration.

Interpreting the transmissibility of measles in two different post periods of supplementary immunization activities in Hubei, China.

Although evidence has shown that supplementary immunization activity (SIA) campaigns greatly reduce the incidence of measles, their effects on disease transmissibility have seldom been monitored. A great decrease in the number of cases may be a false signal of early success towards measles elimination to policy makers. By interpreting the transmissibility in two different post-SIA periods in Hubei, China, the current study showed sustained measles transmissions despite a reduced number of cases. Two population-based cross-sectional serological surveys of measles antibodies were conducted in Hubei province in mid-2010 and mid-2011 after the implementation of SIAs. Immunoglobulin G (IgG) antibodies against measles were measured by enzyme-linked immunosorbent assay (ELISA). Based on the estimated age-specific susceptibility levels, the effective reproduction number (R), a key indicator of disease transmissibility, was determined by the next generation matrix in transmission model. The results revealed an overall IgG seroprevalence of 88.0% (95% confidence interval [CI]: 85.6-90.4%) and 89.6% (95%CI: 88.0-91.2%), respectively, in the two different periods. Comparatively lower seroprevalence rates were observed among children less than 24months of age and young adults 15 to 19years of age in 2011. The Rs were 0.76 and 1.53 for the two study periods. In conclusion, even though the incidence was reduced to below 1/100,000 in both 2010 and 2011, the reproduction number in 2011 indicates a high risk for sustained measles transmission. This finding was potentially due to a lower seropositivity rate among young adults that had not been covered in the first SIA. Thus, implementation of SIA targeted to appropriate age groups is recommended. Regular monitoring of seroprevalence is also suggested to track disease transmissibility and to align SIA with the appropriate age groups.

Organization, Function, and Therapeutic Targeting of the Morbillivirus RNA-Dependent RNA Polymerase Complex.

The morbillivirus genus comprises major human and animal pathogens, including the highly contagious measles virus. Morbilliviruses feature single stranded negative sense RNA genomes that are wrapped by a plasma membrane-derived lipid envelope. Genomes are encapsidated by the viral nucleocapsid protein forming ribonucleoprotein complexes, and only the encapsidated RNA is transcribed and replicated by the viral RNA-dependent RNA polymerase (RdRp). In this review, we discuss recent breakthroughs towards the structural and functional understanding of the morbillivirus polymerase complex. Considering the clinical burden imposed by members of the morbillivirus genus, the development of novel antiviral therapeutics is urgently needed. The viral polymerase complex presents unique structural and enzymatic properties that can serve as attractive candidates for druggable targets. We evaluate distinct strategies for therapeutic intervention and examine how high-resolution insight into the organization of the polymerase complex may pave the path towards the structure-based design and optimization of next-generation RdRp inhibitors.

[Antiviral activity of vegetable triterpens, their derivatives and ribavirin phosphonates]. / Protivovirusnaia aktivnost' triterpenov rastitel'nogo proiskhozhdeniia, ikh proizvodnykh i fosfonatov ribavirina.

The purpose of the study was to investigate, in the Vero cell culture, the antiviral activity of vegetable tritrpens derivatives and ribavirin analogues against the viruses of measles, herpes simple (type 1), cytomegaloviruses and filoviruses. The toxicity and antiviral activity of compounds were determined after coloring of cells with crystal violate. Additionally, the combined action of triterpens' derivatives and ribavirin was investigated. The studied compounds showed relatively low antiviral activity, nonetheless, further research of vegetable triterpens and their derivatives as well as ribavirin analogues would be promising.

[The morphology of the micro-capsulated polyacrylic acid-based formulation of measles vaccine].

The morphology and virus localization were studied in the microcapsulated measles vaccine formulation involving polyacrylic acid (PAA) copolymers as a matrix. Transmission electron microscopy and phosphotungsic staining at a pH value of 2 to 7 showed that the morphology of microparticles was related to the value of pH and to the concentration of a polymer in the matrix. In the neutral medium, the microcapsules had the sizes of 0.5 to 10 microm, which were optimal for transport through the intestinal wall when immunization was orally used. Immunocytochemistry revealed measles virus antigen within the microparticles. The specific activity of the microcapsulated formulation of measles virus was as high as 3.36 and 4.31 lg of TCD50/0.5 ml for the samples containing 1 and 0.1% polymer, respectively. The findings suggest that the microparticles of the vaccine contain live measles virus.

Substituted benzimidazoles with nanomolar activity against respiratory syncytial virus.

A cell-based assay was used to discover compounds inhibiting respiratory syncytial virus (RSV)-induced fusion in HeLa/M cells. A lead compound was identified and subsequent synthesis of >300 analogues led to the identification of JNJ 2408068 (R170591), a low molecular weight (MW 395) benzimidazole derivative with an EC(50) (0.16 nM) against some lab strains almost 100,000 times better than that of ribavirin (15 microM). Antiviral activity was confirmed for subgroup A and B clinical isolates of human RSV and for a bovine RSV isolate. The compound did not inhibit the growth of representative viruses from other Paramyxovirus genera, i.e. HPIV2 and Mumps Virus (genus Rubulavirus), HPIV3 (genus Respirovirus), Measles virus (genus Morbillivirus) and hMPV. Efficacy in cytopathic effect inhibition assays correlated well with efficacy in virus yield reduction assays. A concentration of 10nM reduced RSV production 1000-fold in multi-cycle experiments, irrespective of the multiplicity of infection. Time of addition studies pointed to a dual mode of action: inhibition of virus-cell fusion early in the infection cycle and inhibition of cell-cell fusion at the end of the replication cycle. Two resistant mutants were raised and shown to have single point mutations in the F-gene (S398L and D486N). JNJ 2408068 was also shown to inhibit the release of proinflammatory cytokines IL-6, IL-8 and Rantes from RSV-infected A549 cells.

Evaluation of an expanded case definition for vaccine-modified measles in a school outbreak in South Korea in 2010.

In this study, we have described the clinical characteristics of vaccine-modified measles to assess the performance of an expanded case definition in a school outbreak that occurred in 2010. The sensitivity, specificity, and the positive and negative predictive values were evaluated. Among 74 cases of vaccine-modified measles, 47 (64%) met the original case definition. Fever and rash were observed in 73% (54/74); fever was the most common (96%, 71/74) presenting symptom, and rash was noted in 77% (57/74) of the cases. The original case definition showed an overall sensitivity of 63.5% and a specificity of 100.0%. The expanded case definition combining fever and rash showed a higher sensitivity (72.9%) but a lower specificity (88.2%) than the original. The presence of fever and one or more of cough, coryza, or conjunctivitis scored the highest sensitivity among the combinations of signs and symptoms (77.0%), but scored the lowest specificity (52.9%). The expanded case definition was sensitive in identifying suspected cases of vaccine-modified measles. We suggest using this expanded definition for outbreak investigation in a closed community, and consider further discussions on expanding the case definition of measles for routine surveillance in South Korea.

Virucidal activity of a scorpion venom peptide variant mucroporin-M1 against measles, SARS-CoV and influenza H5N1 viruses.

Outbreaks of SARS-CoV, influenza A (H5N1, H1N1) and measles viruses in recent years have raised serious concerns about the measures available to control emerging and re-emerging infectious viral diseases. Effective antiviral agents are lacking that specifically target RNA viruses such as measles, SARS-CoV and influenza H5N1 viruses, and available vaccinations have demonstrated variable efficacy. Therefore, the development of novel antiviral agents is needed to close the vaccination gap and silence outbreaks. We previously identified mucroporin, a cationic host defense peptide from scorpion venom, which can effectively inhibit standard bacteria. The optimized mucroporin-M1 can inhibit gram-positive bacteria at low concentrations and antibiotic-resistant pathogens. In this investigation, we further tested mucroporin and the optimized mucroporin-M1 for their antiviral activity. Surprisingly, we found that the antiviral activities of mucroporin-M1 against measles, SARS-CoV and influenza H5N1 viruses were notably increased with an EC50 of 7.15 µg/ml (3.52 µM) and a CC50 of 70.46 µg/ml (34.70 µM) against measles virus, an EC50 of 14.46 µg/ml (7.12 µM) against SARS-CoV and an EC50 of 2.10 µg/ml (1.03 µM) against H5N1, while the original peptide mucroporin showed no antiviral activity against any of these three viruses. The inhibition model could be via a direct interaction with the virus envelope, thereby decreasing the infectivity of virus. This report provides evidence that host defense peptides from scorpion venom can be modified for antiviral activity by rational design and represents a practical approach for developing broad-spectrum antiviral agents, especially against RNA viruses.