Improved water vapour spectroscopy in the 4174-4300 cm-1 region and its impact on SCIAMACHY HDO/H2O measurements

The relative abundance of the heavy water isotopologue HDO provides a deeper insight into the atmospheric hydrological cycle. The SCanning Imaging Absorption spectroMeter for Atmospheric CartograpHY (SCIAMACHY) allows for global retrievals of the ratio HDO/H2O in the 2.3 micron wavelength range. However, the spectroscopy of water lines in this region remains a large source of uncertainty for these retrievals. We therefore evaluate and improve the water spectroscopy in the range 4174–4300 cm−1 and test if this reduces systematic uncertainties in the SCIAMACHY retrievals of HDO/H2O. We use a laboratory spectrum of water vapour to fit line intensity, air broadening and wavelength shift parameters. The improved spectroscopy is tested on a series of ground-based high resolution FTS spectra as well as on SCIAMACHY retrievals of H2O and the ratio HDO/H2O. We find that the improved spectroscopy leads to lower residuals in the FTS spectra compared to HITRAN 2008 and Jenouvrier et al. (2007) spectroscopy, and the retrievals become more robust against changes in the retrieval window. For both the FTS and SCIAMACHY measurements, the retrieved total H2O columns decrease by 2–4% and we find a negative shift of the HDO/H2O ratio, which for SCIAMACHY is partly compensated by changes in the retrieval setup and calibration software. The updated SCIAMACHY HDO/H2O product shows somewhat steeper latitudinal and temporal gradients and a steeper Rayleigh distillation curve, strengthening previous conclusions that current isotope-enabled general circulation models underestimate the variability in the near-surface HDO/H2O ratio.

Genetic stability of Schmallenberg virus in vivo during an epidemic, and in vitro, when passaged in the highly susceptible porcine SK-6 cell line

Schmallenberg virus (SBV), an arthropod-borne orthobunyavirus was first detected in 2011 in cattle suffering from diarrhea and fever. The most severe impact of an SBV infection is the induction of malformations in newborns and abortions. Between 2011 and 2013 SBV spread throughout Europe in an unprecedented epidemic wave. SBV contains a tripartite genome consisting of the three negative-sense RNA segments L, M, and S. The virus is usually isolated from clinical samples by inoculation of KC (insect) or BHK-21 (mammalian) cells. Several virus passages are required to allow adaptation of SBV to cells in vitro. In the present study, the porcine SK-6 cell line was used for isolation and passaging of SBV. SK-6 cells proved to be more sensitive to SBV infection and allowed to produce higher titers more rapidly as in BHK-21 cells after just one passage. No adaptation was required. In order to determine the in vivo genetic stability of SBV during an epidemic spread of the virus the nucleotide sequence of the genome from seven SBV field isolates collected in summer 2012 in Switzerland was determined and compared to other SBV sequences available in GenBank. A total of 101 mutations, mostly transitions randomly dispersed along the L and M segment were found when the Swiss isolates were compared to the first SBV isolated late 2011 in Germany. However, when these mutations were studied in detail, a previously described hypervariable region in the M segment was identified. The S segment was completely conserved among all sequenced SBV isolates. To assess the in vitro genetic stability of SBV, three isolates were passage 10 times in SK-6 cells and sequenced before and after passaging. Between two and five nt exchanges per genome were found. This low in vitro mutation rate further demonstrates the suitability of SK-6 cells for SBV propagation.

Ramucirumab versus placebo as second-line treatment in patients with advanced hepatocellular carcinoma following first-line therapy with sorafenib (REACH): a randomised, double-blind, multicentre, phase 3 trial

BACKGROUND VEGF and VEGF receptor-2-mediated angiogenesis contribute to hepatocellular carcinoma pathogenesis. Ramucirumab is a recombinant IgG1 monoclonal antibody and VEGF receptor-2 antagonist. We aimed to assess the safety and efficacy of ramucirumab in advanced hepatocellular carcinoma following first-line therapy with sorafenib. METHODS In this randomised, placebo-controlled, double-blind, multicentre, phase 3 trial (REACH), patients were enrolled from 154 centres in 27 countries. Eligible patients were aged 18 years or older, had hepatocellular carcinoma with Barcelona Clinic Liver Cancer stage C disease or stage B disease that was refractory or not amenable to locoregional therapy, had Child-Pugh A liver disease, an Eastern Cooperative Oncology Group performance status of 0 or 1, had previously received sorafenib (stopped because of progression or intolerance), and had adequate haematological and biochemical parameters. Patients were randomly assigned (1:1) to receive intravenous ramucirumab (8 mg/kg) or placebo every 2 weeks, plus best supportive care, until disease progression, unacceptable toxicity, or death. Randomisation was stratified by geographic region and cause of liver disease with a stratified permuted block method. Patients, medical staff, investigators, and the funder were masked to treatment assignment. The primary endpoint was overall survival in the intention-to-treat population. This study is registered with ClinicalTrials.gov, number NCT01140347. FINDINGS Between Nov 4, 2010, and April 18, 2013, 565 patients were enrolled, of whom 283 were assigned to ramucirumab and 282 were assigned to placebo. Median overall survival for the ramucirumab group was 9·2 months (95% CI 8·0-10·6) versus 7·6 months (6·0-9·3) for the placebo group (HR 0·87 [95% CI 0·72-1·05]; p=0·14). Grade 3 or greater adverse events occurring in 5% or more of patients in either treatment group were ascites (13 [5%] of 277 patients treated with ramucirumab vs 11 [4%] of 276 patients treated with placebo), hypertension (34 [12%] vs ten [4%]), asthenia (14 [5%] vs five [2%]), malignant neoplasm progression (18 [6%] vs 11 [4%]), increased aspartate aminotransferase concentration (15 [5%] vs 23 [8%]), thrombocytopenia (13 [5%] vs one [<1%]), hyperbilirubinaemia (three [1%] vs 13 [5%]), and increased blood bilirubin (five [2%] vs 14 [5%]). The most frequently reported (≥1%) treatment-emergent serious adverse event of any grade or grade 3 or more was malignant neoplasm progression. INTERPRETATION Second-line treatment with ramucirumab did not significantly improve survival over placebo in patients with advanced hepatocellular carcinoma. No new safety signals were noted in eligible patients and the safety profile is manageable. FUNDING Eli Lilly and Co.

The need for second-line antiretroviral therapy in adults in sub-Saharan Africa up to 2030: a mathematical modelling study.

BACKGROUND The number of patients in need of second-line antiretroviral drugs is increasing in sub-Saharan Africa. We aimed to project the need of second-line antiretroviral therapy in adults in sub-Saharan Africa up to 2030. METHODS We developed a simulation model for HIV and applied it to each sub-Saharan African country. We used the WHO country intelligence database to estimate the number of adult patients receiving antiretroviral therapy from 2005 to 2014. We fitted the number of adult patients receiving antiretroviral therapy to observed estimates, and predicted first-line and second-line needs between 2015 and 2030. We present results for sub-Saharan Africa, and eight selected countries. We present 18 scenarios, combining the availability of viral load monitoring, speed of antiretroviral scale-up, and rates of retention and switching to second-line. HIV transmission was not included. FINDINGS Depending on the scenario, 8·7-25·6 million people are expected to receive antiretroviral therapy in 2020, of whom 0·5-3·0 million will be receiving second-line antiretroviral therapy. The proportion of patients on treatment receiving second-line therapy was highest (15·6%) in the scenario with perfect retention and immediate switching, no further scale-up, and universal routine viral load monitoring. In 2030, the estimated range of patients receiving antiretroviral therapy will remain constant, but the number of patients receiving second-line antiretroviral therapy will increase to 0·8-4·6 million (6·6-19·6%). The need for second-line antiretroviral therapy was two to three times higher if routine viral load monitoring was implemented throughout the region, compared with a scenario of no further viral load monitoring scale-up. For each monitoring strategy, the future proportion of patients receiving second-line antiretroviral therapy differed only minimally between countries. INTERPRETATION Donors and countries in sub-Saharan Africa should prepare for a substantial increase in the need for second-line drugs during the next few years as access to viral load monitoring improves. An urgent need exists to decrease the costs of second-line drugs. FUNDING World Health Organization, Swiss National Science Foundation, National Institutes of Health.

Use of a novel DNA melting profile assay for the identification of PCR-amplified genomic sequences encoding for variant-specific surface proteins from the clonal GS/M-83-H7 line of Giardia lamblia.

During infections, Giardia lamblia undergoes a continuous change of its major surface antigens, the variant-specific surface proteins (VSPs). Many studies on antigenic variation have been performed using G. lamblia clone GS/M-83-H7, which expresses surface antigen VSP H7. The present study was focused on the identification and characterization of vsp gene sequences within the genome of the clonal G. lamblia GS/M-83-H7 line. For this purpose, we applied a PCR which specifically amplified truncated sequences from the 3'-terminal region of the vsp genes. Upon cloning, most of the vsp gene amplification products were shown to be approximately identical in size and thus could not be distinguished from each other by conventional gel electrophoresis. In order to pre-estimate the sequence complexity within the large panel of vsp clones isolated, we elaborated a novel concept which facilitated our large-scale genetic screening approach: PCR products from cloned DNA molecules were generated and then subjected to a DNA melting profile assay based on the use of the LightCycler Instrument. This high-throughput assay system proved to be well suited to monitor sequence differences between the amplification products from closely related vsp genes and thus could be used for the primary, sequence-related discrimination of the corresponding clones. After testing 50 candidates, vsp clones could be divided into five groups, each characterized by an individual DNA melting profile of the corresponding amplification products. Sequence analysis of some of these 50 candidates confirmed data from the aforementioned assay in that clones were demonstrated to be identical within, but different between, the distinct groups. The nucleotide and deduced amino acid sequences of five representative vsp clones showed high similarities both among each other and also with the corresponding gene segment of the variant-specific surface antigen (VSP H7) expressed by the original GS/M-83-H7 variant type. Furthermore, three of the genomic vsp sequences turned out to be identical to vsp sequences that represented previously characterized transcription products from in vivo- or in vitro-switched GS/M-83-H7 trophozoites. In conclusion, the DNA melting profile assay seems to be a versatile tool for the PCR-based genotyping of moderately or highly diversified sequence orthologues.