The human membrane trafficking siRNA library targeting 122 genes and the corresponding transfection reagents were purchased from Dharmacon (#G-005500; Dharmacon Research Inc, Lafayette, CO, USA). For the screen, the HeLa-prME cells were seeded in eight 96-well plates with 10,000 cells per well. 24 hours later, 10 pmol of each siRNA was added to each well together with the transfection reagent. siRNA targeting DV1 prME (target sequence: AGATC-CAGCTGACCGATT) and non-targeting siRNA (NT) (Dharmacon Research Inc) were used as controls. Each plate contained in triplicates 15-16 siRNAs from the library, as well as positive (DV1 prME) and negative (NT) siRNA controls. Culture medium was changed two days post-transfection, the supernatant from each well was harvested after an additional 48 hour incubation and then cleared by centrifugation at 4000 rpm for 15 min. 40 ml of supernatant from each well were used to measure by CLDB the levels of RSPs secreted by siRNA transfected HeLa-prME. Two-sided Student's t test was used to assess the statistical significance of differences between each sample and the non-targeting control. Differences were considered statistically significant when P,0.05. Levels of E protein in cell supernatant were expressed in relative luminescence units and ratios of experimental conditions to controls, set as unity, were calculated. Figure S1 The four optimized DV prME sequences. Each optimized prME gene has a BamH I restriction enzyme site, a kozak sequence GCCACC, a signal sequences from VSV-G, and a Xho I restriction enzyme site. Found at: doi:10.1371/journal.pone.0008325.s001 (0.03 MB DOC) Breaking the Waves: Modelling the Potential Impact of Public Health Measures to Defer the Epidemic Peak of Novel Influenza A/H1N1 BACKGROUND: On June 11, 2009, the World Health Organization declared phase 6 of the novel influenza A/H1N1 pandemic. Although by the end of September 2009, the novel virus had been reported from all continents, the impact in most countries of the northern hemisphere has been limited. The return of the virus in a second wave would encounter populations that are still nonimmune and not vaccinated yet. We modelled the effect of control strategies to reduce the spread with the goal to defer the epidemic wave in a country where it is detected in a very early stage. METHODOLOGY/PRINCIPAL FINDINGS: We constructed a deterministic SEIR model using the age distribution and size of the population of Germany based on the observed number of imported cases and the early findings for the epidemiologic characteristics described by Fraser (Science, 2009). We propose a two-step control strategy with an initial effort to trace, quarantine, and selectively give prophylactic treatment to contacts of the first 100 to 500 cases. In the second step, the same measures are focused on the households of the next 5,000 to 10,000 cases. As a result, the peak of the epidemic could be delayed up to 7.6 weeks if up to 30% of cases are detected. However, the cumulative attack rates would not change. Necessary doses of antivirals would be less than the number of treatment courses for 0.1% of the population. In a sensitivity analysis, both case detection rate and the variation of R0 have major effects on the resulting delay. CONCLUSIONS/SIGNIFICANCE: Control strategies that reduce the spread of the disease during the early phase of a pandemic wave may lead to a substantial delay of the epidemic. Since prophylactic treatment is only offered to the contacts of the first 10,000 cases, the amount of antivirals needed is still very limited. On June 11, 2009 , the World Health Organization (WHO) declared Pandemic Phase 6 based on sustained community transmission in more than one WHO region [1] . By the end of September 2009, in most European countries the impact has been limited, which is likely due to the dampening effect of the summer months. What the future holds is difficult to predict as past pandemics have been rather variable because of the complex interactions of immunity, pathogenicity, season and other factors [2] . Many countries have established a stockpile of antivirals for the treatment of the (severely) sick. We examine how a control strategy in the very beginning could defer the peak of the epidemic by using only a very small part of the stored antivirals for prophylactic treatment.