Microbiome profiling by Illumina sequencing of combinatorial sequence-tagged PCR products

We developed a low-cost, high-throughput microbiome profiling method that uses combinatorial sequence tags attached to PCR primers that amplify the rRNA V6 region. Amplified PCR products are sequenced using an Illumina paired-end protocol to generate millions of overlapping reads. Combinatorial sequence tagging can be used to examine hundreds of samples with far fewer primers than is required when sequence tags are incorporated at only a single end. The number of reads generated permitted saturating or near-saturating analysis of samples of the vaginal microbiome. The large number of reads al- lowed an in-depth analysis of errors, and we found that PCR-induced errors composed the vast majority of non-organism derived species variants, an ob- servation that has significant implications for sequence clustering of similar high-throughput data. We show that the short reads are sufficient to assign organisms to the genus or species level in most cases. We suggest that this method will be useful for the deep sequencing of any short nucleotide region that is taxonomically informative; these include the V3, V5 regions of the bac- terial 16S rRNA genes and the eukaryotic V9 region that is gaining popularity for sampling protist diversity.Comment: 28 pages, 13 figure

Serum HCoV-spike specific antibodies do not protect against subsequent SARS-CoV-2 infection in children and adolescents

SARS-CoV-2 infections in children are generally asymptomatic or mild and rarely progress to severe disease and hospitalization. Why this is so remains unclear. Here we explore the potential for protection due to pre-existing cross-reactive seasonal coronavirus antibodies and compare the rate of antibody decline for nucleocapsid and spike protein in serum and oral fluid against SARS-CoV-2 within the pediatric population. No differences in seasonal coronaviruses antibody concentrations were found at baseline between cases and controls, suggesting no protective effect from pre-existing immunity against seasonal coronaviruses. Antibodies against seasonal betacoronaviruses were boosted in response to SARS-CoV-2 infection. In serum, anti-nucleocapsid antibodies fell below the threshold of positivity more quickly than anti-spike protein antibodies. These findings add to our understanding of protection against infection with SARS-CoV-2 within the pediatric population, which is important when considering pediatric SARS-CoV-2 immunization policies

Matrix Metalloproteinase-10 Is Required for Lung Cancer Stem Cell Maintenance, Tumor Initiation and Metastatic Potential

Matrix metalloproteinases (Mmps) stimulate tumor invasion and metastasis by degrading the extracellular matrix. Here we reveal an unexpected role for Mmp10 (stromelysin 2) in the maintenance and tumorigenicity of mouse lung cancer stem-like cells (CSC). Mmp10 is highly expressed in oncosphere cultures enriched in CSCs and RNAi-mediated knockdown of Mmp10 leads to a loss of stem cell marker gene expression and inhibition of oncosphere growth, clonal expansion, and transformed growth in vitro. Interestingly, clonal expansion of Mmp10 deficient oncospheres can be restored by addition of exogenous Mmp10 protein to the culture medium, demonstrating a direct role for Mmp10 in the proliferation of these cells. Oncospheres exhibit enhanced tumor-initiating and metastatic activity when injected orthotopically into syngeneic mice, whereas Mmp10-deficient cultures show a severe defect in tumor initiation. Conversely, oncospheres implanted into syngeneic non-transgenic or Mmp10−/− mice show no significant difference in tumor initiation, growth or metastasis, demonstrating the importance of Mmp10 produced by cancer cells rather than the tumor microenvironment in lung tumor initiation and maintenance. Analysis of gene expression data from human cancers reveals a strong positive correlation between tumor Mmp10 expression and metastatic behavior in many human tumor types. Thus, Mmp10 is required for maintenance of a highly tumorigenic, cancer-initiating, metastatic stem-like cell population in lung cancer. Our data demonstrate for the first time that Mmp10 is a critical lung cancer stem cell gene and novel therapeutic target for lung cancer stem cells

Design, development and validation of a model of problem solving for Egyptian Science classes

Educators and policymakers envision the future of education in Egypt as enabling learners to acquire scientific inquiry and problem-solving skills. In this article, we describe the validation of a model for problem solving and the design of instruments for evaluating new teaching methods in Egyptian science classes. The instruments were based on an established model for problem solving and were designed to assess seventh grade students’ problem solving, experimental strategy knowledge, achievement and motivation towards science. The test for assessing students’ knowledge has been developed based on the topic, density and buoyancy which will be taught in seventh grade in a later intervention study. The instruments were partly self-developed and partly adapted from newly performed studies on strategy knowledge and problem solving in Germany. All instruments were translated into Arabic; the translation process and quality control are described. In order to determine the quality of the instruments, 44 students in Egypt completed the questionnaires and tests. The study’s aim to develop and validate the instruments did require an ad hoc and typical sample which was drawn from an accessible population. Accordingly, the characteristics of the sample are described. Data were analysed according to the classical test theory, but to underpin the results, the instruments were additionally analysed using the even stronger Rasch model. The findings demonstrated the reliability of the items and aspects of validity. In addition, this study showed how test items can be successfully developed and adapted in an international study and applied in a different language

Panel 7: otitis media:treatment and complications

Objective: We aimed to summarize key articles published between 2011 and 2015 on the treatment of (recurrent) acute otitis media, otitis media with effusion, tympanostomy tube otorrhea, chronic suppurative otitis media and complications of otitis media, and their implications for clinical practice. Data Sources: PubMed, Ovid Medline, the Cochrane Library, and Clinical Evidence (BMJ Publishing). Review Methods: All types of articles related to otitis media treatment and complications between June 2011 and March 2015 were identified. A total of 1122 potential related articles were reviewed by the panel members; 118 relevant articles were ultimately included in this summary. Conclusions: Recent literature and guidelines emphasize accurate diagnosis of acute otitis media and optimal management of ear pain. Watchful waiting is optional in mild to moderate acute otitis media; antibiotics do shorten symptoms and duration of middle ear effusion. The additive benefit of adenoidectomy to tympanostomy tubes in recurrent acute otitis media and otitis media with effusion is controversial and age dependent. Topical antibiotic is the treatment of choice in acute tube otorrhea. Symptomatic hearing loss due to persistent otitis media with effusion is best treated with tympanostomy tubes. Novel molecular and biomaterial treatments as adjuvants to surgical closure of eardrum perforations seem promising. There is insufficient evidence to support the use of complementary and alternative treatments. Implications for Practice: Emphasis on accurate diagnosis of otitis media, in its various forms, is important to reduce overdiagnosis, overtreatment, and antibiotic resistance. Children at risk for otitis media and its complications deserve special attention

Distortion Isomerism of Cu(II) Chloride Adducts with Bis(2-benzimidazolyl)ethane. Synthesis, Characterization, X-ray Structures and Spectroscopy of Four Different Isomers

Metals in Catalysis, Biomimetics & Inorganic Material