Genetic variation of a bacterial pathogen within individuals with cystic fibrosis provides a record of selective pressures

Advances in sequencing have enabled the identification of mutations acquired by bacterial pathogens during infection1-10. However, it remains unclear whether adaptive mutations fix in the population or lead to pathogen diversification within the patient11,12. Here, we study the genotypic diversity of Burkholderia dolosa within people with cystic fibrosis by re-sequencing individual colonies and whole populations from single sputum samples. Extensive intra-sample diversity reveals that mutations rarely fix within a patient's pathogen population—instead, diversifying lineages coexist for many years. When strong selection is acting on a gene, multiple adaptive mutations arise but neither sweeps to fixation, generating lasting allele diversity that provides a recorded signature of past selection. Genes involved in outer-membrane components, iron scavenging and antibiotic resistance all showed this signature of within-patient selection. These results offer a general and rapid approach for identifying selective pressures acting on a pathogen in individual patients based on single clinical samples

University–industry collaboration: using meta-rules to overcome barriers to knowledge transfer

This is the final version of the article. Available from Springer Verlag via the DOI in this record.University–industry knowledge transfer is an important source wealth of creation for all partners; however, the practical management of this activity within universities is often hampered by procedural rigidity either through the absence of decision-making protocols to reconcile conflicting priorities or through the inconsistent implementation of existing policies. This is problematic, since it can impede operational effectiveness, prevent inter-organisational knowledge-creation and hamper organisational learning. This paper addresses this issue by adopting a cross-discipline approach and presenting meta-rules as a solution to aid organisational decision making. It is proposed that meta-rules can help resolve tensions arising from conflicting priorities between academics, knowledge transfer offices and industry and help facilitate strategic alignment of processes and policies within and between organisations. This research contributes to the growing debate on the strategic challenges of managing knowledge transfer and presents meta-rules as a practical solution to facilitate strategic alignment of internal and external stakeholder tensions. Meta-rules has previously only been applied in a computer intelligence context however, this research proves the efficacy of meta rules in a university–industry knowledge transfer context. This research also has practical implications for knowledge transfer office managers who can use meta-rules to help overcome resource limitations, conflicting priorities and goals of diverse internal and external stakeholders

Reversion of the ELISPOT test after treatment in Gambian tuberculosis cases

BACKGROUND: New tools are required to improve tuberculosis (TB) diagnosis and treatment, including enhanced ability to compare new treatment strategies. The ELISPOT assay uses Mycobacterium tuberculosis-specific antigens to produce a precise quantitative readout of the immune response to pathogen. We hypothesized that TB patients in The Gambia would have reduced ELISPOT counts after successful treatment. METHODS: We recruited Gambian adults with sputum smear and culture positive tuberculosis for ELISPOT assay and HIV test, and followed them up one year later to repeat testing and document treatment outcome. We used ESAT-6, CFP-10 and Purified Protein Derivative (PPD) as stimulatory antigens. We confirmed the reliability of our assay in 23 volunteers through 2 tests one week apart, comparing within and between subject variation. RESULTS: We performed an ELISPOT test at diagnosis and 12 months later in 89 patients. At recruitment, 70/85 HIV-negative patients (82%) were ESAT-6 or CFP-10 (EC) ELISPOT positive, 77 (90%) were PPD ELISPOT positive. Eighty-two cases (96%) successfully completed treatment: 44 (55%; p < 0.001) were EC ELISPOT negative at 12 months, 17 (21%; p = 0.051) were PPD ELISPOT negative. Sixty (73%) cured cases had a CFP-10 ELISPOT count decrease, 64 (78%) had an ESAT-6 ELISPOT count decrease, 58 (70%) had a PPD ELISPOT count decrease. There was a mean decline of 25, 44 and 47 SFU/2 × 10(5 )cells for CFP-10, ESAT-6 and PPD respectively (p < 0.001 for all). Three of 4 HIV positive patients were cured, all 3 underwent ELISPOT reversion; all 4 not cured subjects (3 HIV-negative, 1 HIV positive) were ESAT-6, CFP-10 and PPD ELISPOT positive at 12 months. CONCLUSION: Successful tuberculosis treatment is accompanied by a significant reduction in the M. tuberculosis-specific antigen ELISPOT count. The ELISPOT has potential as a proxy measure of TB treatment outcome. Further investigation into the decay kinetics of T-cells with treatment is warranted

A High-Throughput Screen Identifies a New Natural Product with Broad-Spectrum Antibacterial Activity

Due to the inexorable invasion of our hospitals and communities by drug-resistant bacteria, there is a pressing need for novel antibacterial agents. Here we report the development of a sensitive and robust but low-tech and inexpensive high-throughput metabolic screen for novel antibiotics. This screen is based on a colorimetric assay of pH that identifies inhibitors of bacterial sugar fermentation. After validation of the method, we screened over 39,000 crude extracts derived from organisms that grow in the diverse ecosystems of Costa Rica and identified 49 with reproducible antibacterial effects. An extract from an endophytic fungus was further characterized, and this led to the discovery of three novel natural products. One of these, which we named mirandamycin, has broad-spectrum antibacterial activity against Escherichia coli, Pseudomonas aeruginosa, Vibrio cholerae, methicillin-resistant Staphylococcus aureus, and Mycobacterium tuberculosis. This demonstrates the power of simple high throughput screens for rapid identification of new antibacterial agents from environmental samples

Characterising the CI and CI-like carbonaceous chondrites using thermogravimetric analysis and infrared spectroscopy

The CI and CI-like chondrites provide a record of aqueous alteration in the early solar system. However, the CI-like chondrites differ in having also experienced a late stage period of thermal metamorphism. In order to constrain the nature and extent of the aqueous and thermal alteration, we have investigated the bulk mineralogy and abundance of H2O in the CI and CI-like chondrites using thermogravimetric analysis and infrared spectroscopy. The CI chondrites Ivuna and Orgueil show significant mass loss (28.5–31.8 wt.%) upon heating to 1000 °C due to dehydration and dehydroxylation of abundant phyllosilicates and Fe-(oxy)hydroxides and the decomposition of Fe-sulphides, carbonates and organics. Infrared spectra for Ivuna and Orgueil have a prominent 3-μm feature due to bound −OH/H2O in phyllosilicates and Fe-(oxy)hydroxides and only a minor 11-μm feature from anhydrous silicates. These characteristics are consistent with previous studies indicating that the CI chondrites underwent near-complete aqueous alteration. Similarities in the total abundance of H2O and 3 μm/11 μm ratio suggest that there is no difference in the relative degree of hydration experienced by Ivuna and Orgueil. In contrast, the CI-like chondrites Y-82162 and Y-980115 show lower mass loss (13.8–18.8 wt.%) and contain >50 % less H2O than the CI chondrites. The 3-μm feature is almost absent from spectra of Y-82162 and Y-980115 but the 11-μm feature is intense. The CI-like chondrites experienced thermal metamorphism at temperatures >500 °C that initially caused dehydration and dehydroxylation of phyllosilicates before partial recrystallization back into anhydrous silicates. The surfaces of many C-type asteroids were probably heated through impact metamorphism and/or solar radiation, so thermally altered carbonaceous chondrites are likely good analogues for samples that will be returned by the Hayabusa-2 and OSIRIS-REx missions

Two Origins for the Gene Encoding α-Isopropylmalate Synthase in Fungi

BACKGROUND: The biosynthesis of leucine is a biochemical pathway common to prokaryotes, plants and fungi, but absent from humans and animals. The pathway is a proposed target for antimicrobial therapy. METHODOLOGY/PRINCIPAL FINDINGS: Here we identified the leuA gene encoding alpha-isopropylmalate synthase in the zygomycete fungus Phycomyces blakesleeanus using a genetic mapping approach with crosses between wild type and leucine auxotrophic strains. To confirm the function of the gene, Phycomyces leuA was used to complement the auxotrophic phenotype exhibited by mutation of the leu3+ gene of the ascomycete fungus Schizosaccharomyces pombe. Phylogenetic analysis revealed that the leuA gene in Phycomyces, other zygomycetes, and the chytrids is more closely related to homologs in plants and photosynthetic bacteria than ascomycetes or basidiomycetes, and suggests that the Dikarya have acquired the gene more recently. CONCLUSIONS/SIGNIFICANCE: The identification of leuA in Phycomyces adds to the growing body of evidence that some primary metabolic pathways or parts of them have arisen multiple times during the evolution of fungi, probably through horizontal gene transfer events

Epidemiology and risk factors for Staphylococcus aureus colonization in children in the post-PCV7 era

<p>Abstract</p> <p>Background</p> <p>The incidence of community-associated methicillin-resistant <it>Staphylococcus aureus </it>(MRSA) has risen dramatically in the U.S., particularly among children. Although <it>Streptococcus pneumoniae </it>colonization has been inversely associated with <it>S. aureus </it>colonization in unvaccinated children, this and other risk factors for <it>S. aureus </it>carriage have not been assessed following widespread use of the heptavalent pneumococcal conjugate vaccine (PCV7). Our objectives were to (1) determine the prevalence of <it>S. aureus </it>and MRSA colonization in young children in the context of widespread use of PCV7; and (2) examine risk factors for <it>S. aureus </it>colonization in the post-PCV7 era, including the absence of vaccine-type <it>S. pneumoniae </it>colonization.</p> <p>Methods</p> <p>Swabs of the anterior nares (<it>S. aureus</it>) were obtained from children enrolled in an ongoing study of nasopharyngeal pneumococcal colonization of healthy children in 8 Massachusetts communities. Children 3 months to <7 years of age seen for well child or sick visits in primary care offices from 11/03–4/04 and 10/06–4/07 were enrolled. <it>S. aureus </it>was identified and antibiotic susceptibility testing was performed. Epidemiologic risk factors for <it>S. aureus </it>colonization were collected from parent surveys and chart reviews, along with data on pneumococcal colonization. Multivariate mixed model analyses were performed to identify factors associated with <it>S. aureus </it>colonization.</p> <p>Results</p> <p>Among 1,968 children, the mean age (SD) was 2.7 (1.8) years, 32% received an antibiotic in the past 2 months, 2% were colonized with PCV7 strains and 24% were colonized with non-PCV7 strains. The prevalence of <it>S. aureus </it>colonization remained stable between 2003–04 and 2006–07 (14.6% vs. 14.1%), while MRSA colonization remained low (0.2% vs. 0.9%, p = 0.09). Although absence of pneumococcal colonization was not significantly associated with <it>S. aureus </it>colonization, age (6–11 mo vs. ≥5 yrs, OR 0.39 [95% CI 0.24–0.64]; 1–1.99 yrs vs. ≥5 yrs, OR 0.35 [0.23–0.54]; 2–2.99 yrs vs. ≥5 yrs, OR 0.45 [0.28–0.73]; 3–3.99 yrs vs. ≥5 yrs, OR 0.53 [0.33–0.86]) and recent antibiotic use were significant predictors in multivariate models.</p> <p>Conclusion</p> <p>In Massachusetts, <it>S. aureus </it>and MRSA colonization remained stable from 2003–04 to 2006–07 among children <7 years despite widespread use of pneumococcal conjugate vaccine. <it>S. aureus </it>nasal colonization varies by age and is inversely correlated with recent antibiotic use.</p