Characterization and expression analysis of Staphylococcus aureus pathogenicity island 3 - Implications for the evolution of staphylococcal pathogenicity islands

We describe the complete sequence of the 15.9-kb staphylococcal pathogenicity island 3 encoding staphylococcal enterotoxin serotypes B, K, and Q. The island, which meets the generally accepted definition of pathogenicity islands, contains 24 open reading frames potentially encoding proteins of more than 50 amino acids, including an apparently functional integrase. The element is bordered by two 17-bp direct repeats identical to those found flanking staphylococcal pathogenicity island 1. The island has extensive regions of homology to previously described pathogenicity islands, particularly staphylococcal pathogenicity islands 1 and bov. The expression of 22 of the 24 open reading frames contained on staphylococcal pathogenicity island 3 was detected either in vitro during growth in a laboratory medium or serum or in vivo in a rabbit model of toxic shock syndrome using DNA microarrays. The effect of oxygen tension on staphylococcal pathogenicity island 3 gene expression was also examined. By comparison with the known staphylococcal pathogenicity islands in the context of gene expression described here, we propose a model of pathogenicity island origin and evolution involving specialized transduction events and addition, deletion, or recombination of pathogenicity island "modules.

Partial and Incremental PCMH Practice Transformation: Implications for Quality and Costs

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/102699/1/hesr12085.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/102699/2/hesr12085-sup-0002-AuthorMatrix.pd

Comparative genomic analysis of Mycobacterium avium subspecies obtained from multiple host species

<p>Abstract</p> <p>Background</p> <p><it>Mycobacterium avium </it>(<it>M. avium</it>) subspecies vary widely in both pathogenicity and host specificity, but the genetic features contributing to this diversity remain unclear.</p> <p>Results</p> <p>A comparative genomic approach was used to identify large sequence polymorphisms among <it>M. avium </it>subspecies obtained from a variety of host animals. DNA microarrays were used as a platform for comparing mycobacterial isolates with the sequenced bovine isolate <it>M. avium </it>subsp. <it>paratuberculosis </it>(MAP) K-10. Open reading frames (ORFs) were classified as present or divergent based on the relative fluorescent intensities of the experimental samples compared to MAP K-10 DNA. Multiple large polymorphic regions were found in the genomes of MAP isolates obtained from sheep. One of these clusters encodes glycopeptidolipid biosynthesis enzymes which have not previously been identified in MAP. <it>M. avium </it>subsp. <it>silvaticum </it>isolates were observed to have a hybridization profile very similar to yet distinguishable from <it>M. avium </it>subsp. <it>avium</it>. Isolates obtained from cattle (n = 5), birds (n = 4), goats (n = 3), bison (n = 3), and humans (n = 9) were indistinguishable from cattle isolate MAP K-10.</p> <p>Conclusion</p> <p>Genome diversity in <it>M. avium </it>subspecies appears to be mediated by large sequence polymorphisms that are commonly associated with mobile genetic elements. Subspecies and host adapted isolates of <it>M. avium </it>were distinguishable by the presence or absence of specific polymorphisms.</p

Early antibody response against Mycobacterium avium subspecies paratuberculosis antigens in subclinical cattle

<p>Abstract</p> <p>Background</p> <p>Our laboratories have previously reported on the experimental infection of cattle with <it>Mycobacterium avium </it>subsp <it>paratuberculosis </it>(<it>M. paratuberculosis</it>) using an intratonsillar infection model. In addition, we have recently developed a partial protein array representing 92 <it>M. paratuberculosis </it>coding sequences. These combined tools have enabled a unique look at the temporal analysis of <it>M. paratuberculosis </it>antigens within the native host. The primary objective of this study was to identify <it>M. paratuberculosis </it>antigens detected by cattle early during infection. A secondary objective was to evaluate the humoral immune response in cattle during the initial year of infection.</p> <p>Results</p> <p>Sera from two experimentally infected cattle, taken pre-inoculation and at day 70, 194 and 321 post infection, identified dynamic antibody reactivity among antigens with some showing an increased response over time and others showing declining levels of reactivity over the same time period. A <it>M. paratuberculosis </it>specific protein, encoded by MAP0862, was strongly detected initially, but the antibody response became weaker with time. The most reactive protein was a putative surface antigen encoded by MAP1087. A second protein, MAP1204, implicated in virulence, was also strongly detected by day 70 in both cattle. Subsequent experiments showed that these two proteins were detected with sera from 5 of 9 naturally infected cattle in the subclinical stage of Johne's disease.</p> <p>Conclusion</p> <p>Collectively these results demonstrate that <it>M. paratuberculosis </it>proteins are detected by sera from experimentally infected cattle as early as 70 days after exposure. These data further suggest at least two antigens may be useful in the early diagnosis of <it>M. paratuberculosis </it>infections. Finally, the construction and use of a protein array in this pilot study has led to a novel approach for discovery of <it>M. paratuberculosis </it>antigens.</p

Robust paths to net greenhouse gas mitigation and negative emissions via advanced biofuels

ACKNOWLEDGEMENTS We thank Dennis Ojima and Daniel L. Sanchez for their encouragement on this topic. The authors gratefully acknowledge partial support as follows: J.L.F., L.R.L., T.L.R., E.A.H.S., and J.J.S., the Sao Paulo Research Foundation (FAPESP grant# 2014/26767-9); J.L.F., L.R.L., K.P., and T.L.R., The Center for Bioenergy Innovation, a U.S. Department of Energy Research Center supported by the Office of Biological and Environmental Research in the DOE Office of Science (grant# DE-AC05-00OR22725); L.R.L., the Sao Paulo Research Foundation, and the Link Foundation; J.L.F. and K.P., USDA/NIFA (grant# 2013-68005-21298 and 2017-67019-26327); T.L.R., USDA/NIFA (grant# 2012-68005-19703); D.S.L. and S.P.L., the Energy Biosciences Institute. Data availability The DayCent model (https://www2.nrel.colostate.edu/projects/daycent/) is freely available upon request. Specification of DayCent model runs and automated model initialization, calibration, scenario simulation, results analysis, and figure generation were implemented in Python 2.7, using the numpy module for data processing and the matplotlib module for figure generation. Analysis code is available in a version-controlled repository (https://github.com/johnlfield/Ecosystem_dynamics). A working copy of the code, all associated DayCent model inputs, and analysis outputs are also available in an online data repository (https://figshare.com/s/4c14ec168bd550db4bad; note this URL is for accessing a private version of the repository, and will eventually be replaced with an updated URL for the public version of the repository, which will only be accessible after the journal-specified embargo date).Peer reviewedPostprintPublisher PD

Genome sequencing of ovine isolates of Mycobacterium avium subspecies paratuberculosis offers insights into host association

<p>Abstract</p> <p>Background</p> <p>The genome of <it>Mycobacterium avium </it>subspecies <it>paratuberculosis </it>(<it>MAP</it>) is remarkably homogeneous among the genomes of bovine, human and wildlife isolates. However, previous work in our laboratories with the bovine K-10 strain has revealed substantial differences compared to sheep isolates. To systematically characterize all genomic differences that may be associated with the specific hosts, we sequenced the genomes of three U.S. sheep isolates and also obtained an optical map.</p> <p>Results</p> <p>Our analysis of one of the isolates, <it>MAP </it>S397, revealed a genome 4.8 Mb in size with 4,700 open reading frames (ORFs). Comparative analysis of the <it>MAP </it>S397 isolate showed it acquired approximately 10 large sequence regions that are shared with the human <it>M. avium </it>subsp. <it>hominissuis </it>strain 104 and lost 2 large regions that are present in the bovine strain. In addition, optical mapping defined the presence of 7 large inversions between the bovine and ovine genomes (~ 2.36 Mb). Whole-genome sequencing of 2 additional sheep strains of <it>MAP </it>(JTC1074 and JTC7565) further confirmed genomic homogeneity of the sheep isolates despite the presence of polymorphisms on the nucleotide level.</p> <p>Conclusions</p> <p>Comparative sequence analysis employed here provided a better understanding of the host association, evolution of members of the <it>M. avium </it>complex and could help in deciphering the phenotypic differences observed among sheep and cattle strains of <it>MAP</it>. A similar approach based on whole-genome sequencing combined with optical mapping could be employed to examine closely related pathogens. We propose an evolutionary scenario for <it>M. avium </it>complex strains based on these genome sequences.</p

Long-term ecological research on Colorado Shortgrass Steppe

The SGS-LTER research site was established in 1980 by researchers at Colorado State University as part of a network of long-term research sites within the US LTER Network, supported by the National Science Foundation. Scientists within the Natural Resource Ecology Lab, Department of Forest and Rangeland Stewardship, Department of Soil and Crop Sciences, and Biology Department at CSU, California State Fullerton, USDA Agricultural Research Service, University of Northern Colorado, and the University of Wyoming, among others, have contributed to our understanding of the structure and functions of the shortgrass steppe and other diverse ecosystems across the network while maintaining a common mission and sharing expertise, data and infrastructure.Poster presented at the LTER All Scientists Meeting held in Estes Park, CO on September 10-13, 2012

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Transcriptional Response of Pasteurella multocida to Nutrient Limitation

Bacteria often encounter environments where nutrient availability is limited, and they must adapt accordingly. To identify Pasteurella multocida genes that are differentially expressed during nutrient limitation, we utilized whole-genome microarrays to compare levels of gene expression during growth in rich and minimal media. Our analysis showed that the levels of expression of a total of 669 genes, representing approximately one-third of the genome, were detectably altered over the course of the experiment. A large number (n = 439) of genes, including those involved in energy metabolism, transport, protein synthesis, and binding, were expressed at higher levels in rich medium, suggesting that, upon exposure to a rich environment, P. multocida immediately begins to turn on many energy-intensive biosynthetic pathways or, conversely, turns these genes off when it is exposed to a nutrient-deficient environment. Genes with increased expression in minimal medium (n = 230) included those encoding amino acid biosynthesis and transport systems, outer membrane proteins, and heat shock proteins. Importantly, our analysis also identified a large number (n = 164) of genes with unknown functions whose expression was altered during nutrient limitation. Overall, the results of our study show that a wide repertoire of genes, many of which have yet to be functionally classified, undergo transcriptional regulation in P. multocida in response to growth in minimal medium and provide a strong foundation to investigate the transcriptional response of this multispecies pathogen to growth in a nutrient-limited environment

Comparative Genomic Hybridizations Reveal Genetic Regions within the Mycobacterium avium Complex That Are Divergent from Mycobacterium avium subsp. paratuberculosis Isolates

Mycobacterium avium subsp. paratuberculosis is genetically similar to other members of the Mycobacterium avium complex (MAC), some of which are nonpathogenic and widespread in the environment. We have utilized an M. avium subsp. paratuberculosis whole-genome microarray representing over 95% of the predicted coding sequences to examine the genetic conservation among 10 M. avium subsp. paratuberculosis isolates, two isolates each of Mycobacterium avium subsp. silvaticum and Mycobacterium avium subsp. avium, and a single isolate each of both Mycobacterium intracellulare and Mycobacterium smegmatis. Genomic DNA from each isolate was competitively hybridized with DNA from M. avium subsp. paratuberculosis K10, and open reading frames (ORFs) were classified as present, divergent, or intermediate. None of the M. avium subsp. paratuberculosis isolates had ORFs classified as divergent. The two M. avium subsp. avium isolates had 210 and 135 divergent ORFs, while the two M. avium subsp. silvaticum isolates examined had 77 and 103 divergent ORFs. Similarly, 130 divergent ORFs were identified in M. intracellulare. A set of 97 ORFs were classified as divergent or intermediate in all of the nonparatuberculosis MAC isolates tested. Many of these ORFs are clustered together on the genome in regions with relatively low average GC content compared with the entire genome and contain mobile genetic elements. One of these regions of sequence divergence contained genes homologous to a mammalian cell entry (mce) operon. Our results indicate that closely related MAC mycobacteria can be distinguished from M. avium subsp. paratuberculosis by multiple clusters of divergent ORFs