On The Irrelevance Of Transformational Grammar To Second Language Pedagogy

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/98173/1/j.1467-1770.1969.tb00467.x.pd

High-Resolution Identification of Multiple Salmonella Serovars in a Single Sample by Using CRISPR-SeroSeq

Salmonella enterica is represented by \u3e2,600 serovars that can differ in routes of transmission, host colonization, and in resistance to antimicrobials. S. enterica is the leading bacterial cause of foodborne illness in the United States, with well-established detection methodology. Current surveillance protocols rely on the characterization of a few colonies to represent an entire sample; thus, minority serovars remain undetected. Salmonella contains two CRISPR loci, CRISPR1 and CRISPR2, and the spacer contents of these can be considered serovar specific. We exploited this property to develop an amplicon-based and multiplexed sequencing approach, CRISPR-SeroSeq (serotyping by sequencing of the CRISPR loci), to identify multiple serovars present in a single sample. Using mixed genomic DNA from two Salmonella serovars, we were able to confidently detect a serovar that constituted 0.01% of the sample. Poultry is a major reservoir of Salmonella spp., including serovars that are frequently associated with human illness, as well as those that are not. Numerous studies have examined the prevalence and diversity of Salmonella spp. in poultry, though these studies were limited to culture-based approaches and therefore only identified abundant serovars. CRISPR-SeroSeq was used to investigate samples from broiler houses and a processing facility. Ninety-one percent of samples harbored multiple serovars, and there was one sample in which four different serovars were detected. In another sample, reads for the minority serovar comprised 0.003% of the total number of Salmonella spacer reads. The most abundant serovars identified were Salmonella enterica serovars Montevideo, Kentucky, Enteritidis, and Typhimurium. CRISPR-SeroSeq also differentiated between multiple strains of some serovars. This high resolution of serovar populations has the potential to be utilized as a powerful tool in the surveillance of Salmonella species

The Role Of Extrinsic Feedback In Interlanguage Fossilization

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/98419/1/j.1467-1770.1979.tb01075.x.pd

Molecular Diversity of Bacteroidales in Fecal and Environmental Samples and Swine-Associated Subpopulations

Several swine-specific microbial source tracking methods are based on PCR assays targeting Bacteroidales 16S rRNA gene sequences. The limited application of these assays can be explained by the poor understanding of their molecular diversity in fecal sources and environmental waters. In order to address this, we studied the diversity of 9,340 partial (\u3e600bp in length) Bacteroidales 16S rRNA gene sequences from 13 fecal sources and nine feces-contaminated watersheds. The compositions of major Bacteroidales populations were analyzed to determine which host and environmental sequences were contributing to each group. This information allowed us to identify populations which were both exclusive to swine fecal sources and detected in swine-contaminated waters. Phylogenetic and diversity analyses revealed that some markers previously believed to be highly specific to swine populations are shared by multiple hosts, potentially explaining the cross-amplification signals obtained with nontargeted hosts. These data suggest that while many Bacteroidales populations are cosmopolitan, others exhibit a preferential host distribution and may be able to survive different environmental conditions. This study further demonstrates the importance of elucidating the diversity patterns of targeted bacterial groups to develop more inclusive fecal source tracking applications

Using Phosphorus Recovery from Wastewater as a Context for Teaching Sustainable Development with USEPA P3 Support

Phosphorus is an essential element required for agriculture. Current practices include one-way use of phosphorus: that is mining; production of fertilizer; land application; and ultimately loss to aquatic sediments. Once in the aquatic environment, phosphorus stimulates eutrophication resulting in the death of water bodies. With financial support from a People, Prosperity, and Planet program grant of the U.S. Environmental Protection Agency, an interdisciplinary team of faculty at the University of Cincinnati used phosphorus recovery from sewage and re-utilization as a struvite fertilizer as the context for a year-long course of study integrating graduate and undergraduate students in Environmental Engineering and Science as well as Environmental Studies. This presentation will highlight the original course format, results of student assessment from the 2004-2005 academic year, modifications incorporated for the 2005-2006 academic year, and the subsequent findings of student perceptions and learning. The challenge of integrating across two colleges within a comprehensive university system will be discussed, and our approaches for meeting the learning needs and course expectations of a diverse student population will be included

Identification of Host-Specific Bacteroidales 16S rDNA Sequences from Human Sewage and Ruminant Feces

The need to identify the source of fecal contamination of water has led to the development of various fecal source identification methods, a field known as microbial source tracking (MST). One promising method of MST focuses on fecal members of the order Bacteroidales, some of which exhibit a high degree of host-specificity. In order to identify host-specific Bacteroidales genetic markers, a ∼1060 bp section of Bacteroidales 16S rDNA was amplified from human sewage (n = 6), and bovine (n = 6) and ovine fecal (n = 5) samples and used for the generation of three clone libraries. Phylogenetic analysis of sequences from the three clone libraries revealed that the Bacteroidales species found in both human sewage and bovine and ovine feces were a highly diverse group of organisms, many of which were not represented by previously characterised 16S rDNA. Ovine and bovine feces appear to host similar populations of Bacteroidales species and these species were more diverse and less closely related to cultivated species than the Bacteroidales population found in human sewage. Species of Bacteroidales from the ruminant and human feces formed isolated clusters containing putatively host-specific sequences. These sequences were subsequently exploited for the design of host-specific primers which were used in MST studies

Comparative fecal metagenomics unveils unique functional capacity of the swine gut

<p>Abstract</p> <p>Background</p> <p>Uncovering the taxonomic composition and functional capacity within the swine gut microbial consortia is of great importance to animal physiology and health as well as to food and water safety due to the presence of human pathogens in pig feces. Nonetheless, limited information on the functional diversity of the swine gut microbiome is available.</p> <p>Results</p> <p>Analysis of 637, 722 pyrosequencing reads (130 megabases) generated from Yorkshire pig fecal DNA extracts was performed to help better understand the microbial diversity and largely unknown functional capacity of the swine gut microbiome. Swine fecal metagenomic sequences were annotated using both MG-RAST and JGI IMG/M-ER pipelines. Taxonomic analysis of metagenomic reads indicated that swine fecal microbiomes were dominated by Firmicutes and Bacteroidetes phyla. At a finer phylogenetic resolution, <it>Prevotella </it>spp. dominated the swine fecal metagenome, while some genes associated with <it>Treponema </it>and <it>Anareovibrio </it>species were found to be exclusively within the pig fecal metagenomic sequences analyzed. Functional analysis revealed that carbohydrate metabolism was the most abundant SEED subsystem, representing 13% of the swine metagenome. Genes associated with stress, virulence, cell wall and cell capsule were also abundant. Virulence factors associated with antibiotic resistance genes with highest sequence homology to genes in Bacteroidetes, Clostridia, and <it>Methanosarcina </it>were numerous within the gene families unique to the swine fecal metagenomes. Other abundant proteins unique to the distal swine gut shared high sequence homology to putative carbohydrate membrane transporters.</p> <p>Conclusions</p> <p>The results from this metagenomic survey demonstrated the presence of genes associated with resistance to antibiotics and carbohydrate metabolism suggesting that the swine gut microbiome may be shaped by husbandry practices.</p

Engineering Education Collaboration: Innovative Pedagogical Methods for High School and University Environmentalists

This paper presents an innovative teaching approach, how it is implemented, student response results of the implementation, and the assessment of impact on student learning. The findings are based on surveys given to the students after each lab lesson taught in partnership with university (Project STEP) and community members. The purpose of this paper is to showcase authentic molecular technology research methods that have been incorporated into a high school level water quality study in cooperation with a watershed restoration program. Typically, water quality studies focus on chemical analysis such as pH, dissolved oxygen, biochemical oxygen demand, orthophosphates, nitrates, temperature, turbidity, macro-invertebrate survey and fecal coliform cultures. This paper shows that by using molecular technology, the source of pollution in the watershed can be determined. Students in these high school science classes are engaged in authentic experiences to identify and analyze human impact on the environment and local ecosystems. Students also are able to collect and analyze data using computer and molecular technology. With help from the local watershed managers, the AP high school students filter bacteria, isolate their DNA, use the polymerase chain reaction (PCR) to amplify the DNA, and finally use gel electrophoresis to trace the DNA to its source (human, cow or intestinal bacteria). In this way, both AP and Physical Science students can extend the water quality study to trace the pollution to a point source. This is a unique approach to high school science laboratory activities. All watershed data is collected and organized using Microsoft Excel spreadsheets and graphing software. Students are able to form conclusions using technology that is used in today\u27s workplace. Initial findings regarding student response to this innovative teaching approach indicate that the actual application of molecular technology methods, employed to solve a problem with an unknown conclusion, is very meaningful to students. Unlike other traditional classroom labs, neither the teacher nor the students know what the results of the watershed tests are before-hand. This type of innovative teaching approach, supported by research on inquiry lessons, provides a more memorable experience for the students - actually performing technology that they would otherwise only read about in textbooks and articles. This paper will provide other instructors with a kind of roadmap, but one where there are experiences of many partners and students that highlight both successes and challenges

Bacterial Bio-indicators of Marcellus Shale Activities in Pennsylvania: A Molecular Ecology Survey

The practice of hydraulic fracking has increased over the years especially in Pennsylvania where most of the subterraneous gas-rich Marcellus Shale formations are located. Our previous work showed that headwater streams in proximity to hydraulic fracking operations have significantly different bacterial assemblages as compared to un-impacted streams in central PA. Aquatic bacterial communities are of great importance because they are often the ‘first-responders’ to environmental perturbations. We are interested in which bacteria become enriched, as this might serve as robust biomarkers of fracking, and can potentially biodegrade constituents of fracking fluids. In this study, we plan to expand upon our previous work to identify additional sentinel bacterial taxa in other areas in PA (Northeast and Southwest) heavily impacted by fracking. Water and sediment samples have been collected from Northern Pennsylvania (n=31) and Southwestern (n=11) regions upstream and downstream of fracking activities. Bacterial community profiles of these samples were generated via high-throughput sequencing of the 16S rRNA, a robust phylogenetic marker for bacterial identification. The data generated provide a snapshot of all bacteria taxa present and their relative abundance. Thus, differences in bacterial community structure between impacted and un-impacted environments can help glean which bacterial taxa are responding to environmental perturbations associated with fracking. This research can help us generate a list of potential bioindicators of nascent fracking activities and can be used to help track impacts and bioremediation potential within environmental scenarios

Specificity and sensitivity evaluation of novel and existing Bacteroidales and Bifidobacteria-specific PCR assays on feces and sewage samples and their application for microbial source tracking in Ireland

Three novel ruminant-specific PCR assays, an existing ruminant-specific PCR assay and five existing human-specific PCR assays, which target 16S rDNA from Bacteroidales or Bifidobacteria, were evaluated. The assays were tested on DNA extracted from ruminant (n = 74), human (n = 59) and non-ruminant animal (n = 44) sewage/fecal samples collected in Ireland. The three novel PCR assays compared favourably to the existing ruminant-specific assay, exhibiting sensitivities of 91 - 100% and specificities of 95 - 100% as compared to a sensitivity of 95% and specificity of 94%, for the existing ruminant-specific assay. Of the five human-specific PCR assays, the assay targeting the Bifidobacterium catenulatum group was the most promising, exhibiting a sensitivity of 100% (with human sewage samples) and a specificity of 87%. When tested on rural water samples that were naturally contaminated by ruminant feces, the three novel PCR assays tested positive with a much greater percentage (52 - 87%) of samples than the existing ruminant-specific assay (17%). These novel ruminant-specific assays show promise for microbial source tracking and merit further field testing and specificity evaluation.ERTD