Rodney A. King

A Broadly Implementable Research Course in Phage Discovery and Genomics for First-Year Undergraduate Students

ABSTRACT Engaging large numbers of undergraduates in authentic scientific discovery is desirable but difficult to achieve. We have developed a general model in which faculty and teaching assistants from diverse academic institutions are trained to teach a research course for first-year undergraduate students focused on bacteriophage discovery and genomics. The course is situated within a broader scientific context aimed at understanding viral diversity, such that faculty and students are collaborators with established researchers in the field. The Howard Hughes Medical Institute (HHMI) Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) course has been widely implemented and has been taken by over 4,800 students at 73 institutions. We show here that this alliance-sourced model not only substantially advances the field of phage genomics but also stimulates students’ interest in science, positively influences academic achievement, and enhances persistence in science, technology, engineering, and mathematics (STEM) disciplines. Broad application of this model by integrating other research areas with large numbers of early-career undergraduate students has the potential to be transformative in science education and research training. IMPORTANCE Engagement of undergraduate students in scientific research at early stages in their careers presents an opportunity to excite students about science, technology, engineering, and mathematics (STEM) disciplines and promote continued interests in these areas. Many excellent course-based undergraduate research experiences have been developed, but scaling these to a broader impact with larger numbers of students is challenging. The Howard Hughes Medical Institute (HHMI) Science Education Alliance Phage Hunting Advancing Genomics and Evolutionary Science (SEA-PHAGES) program takes advantage of the huge size and diversity of the bacteriophage population to engage students in discovery of new viruses, genome annotation, and comparative genomics, with strong impacts on bacteriophage research, increased persistence in STEM fields, and student self-identification with learning gains, motivation, attitude, and career aspirations. Engagement of undergraduate students in scientific research at early stages in their careers presents an opportunity to excite students about science, technology, engineering, and mathematics (STEM) disciplines and promote continued interests in these areas. Many excellent course-based undergraduate research experiences have been developed, but scaling these to a broader impact with larger numbers of students is challenging. The Howard Hughes Medical Institute (HHMI) Science Education Alliance Phage Hunting Advancing Genomics and Evolutionary Science (SEA-PHAGES) program takes advantage of the huge size and diversity of the bacteriophage population to engage students in discovery of new viruses, genome annotation, and comparative genomics, with strong impacts on bacteriophage research, increased persistence in STEM fields, and student self-identification with learning gains, motivation, attitude, and career aspirations.

Characterization of Salmonella Bacteriophages Isolated from Swine Lagoon Effluent

Four Salmonella bacteriophages that had been originally isolated from swine manure lagoons were characterized and compared to each other and to well-known Salmonella phages P22 and Felix 01. Host ranges of the lagoon phages were similar to each other in spot tests on reference strains of Salmonella, but differed slightly from each other on a panel of Salmonella lagoon strains. In single-step growth at 35°C the lagoon phages had latent periods of 15 to 20 min and burst sizes from 100 to 230. The lagoon phages and P22 were purified by cesium chloride (CsCl) gradient centrifugation and used to produce specific antisera and DNA. The lagoon phages were indistinguishable from each other but distinct from P22 and Felix 01 in immunodiffusion and infectivity neutralization tests and in restriction digest analysis.

Comparative genomics of Cluster O mycobacteriophages.

Mycobacteriophages – viruses of mycobacterial hosts – are genetically diverse but morphologically are all classified in the Caudovirales with double-stranded DNA and tails. We describe here a group of five closely related mycobacteriophages – Corndog, Catdawg, Dylan, Firecracker, and YungJamal – designated as Cluster O with long flexible tails but with unusual prolate capsids. Proteomic analysis of phage Corndog particles, Catdawg particles, and Corndog-infected cells confirms expression of half of the predicted gene products and indicates a non-canonical mechanism for translation of the Corndog tape measure protein. Bioinformatic analysis identifies 8–9 strongly predicted SigA promoters and all five Cluster O genomes contain more than 30 copies of a 17 bp repeat sequence with dyad symmetry located throughout the genomes. Comparison of the Cluster O phages provides insights into phage genome evolution including the processes of gene flux by horizontal genetic exchange.

The HHMI National Genomics Research Initiative at Western Kentucky University

Background Western Kentucky University is a member of a select group of colleges and universities chosen to participate in a national science experiment designed to improve undergraduate science education. To introduce freshman students to authentic research, the Howard Hughes Medical Institute’s Science Education Alliance (HHMI SEA) has implemented an innovative program to introduce genomics into the classroom. The program draws on themes and techniques from biology, microbiology, molecular biology, genomics and bioinformatics. The WKU course is called “Genome Discovery and Exploration” (GDE); WKU Genome Discovery and Exploration Program [http://bioweb.wku.edu/asp/wkungri/]. During the fall semester, students isolate bacterial viruses from the environment and characterize them using a variety of techniques including DNA analysis and electron microscopy. Over the winter break, the complete DNA sequence of one of the viruses isolated by the GDE students is determined. During the spring semester, GDE students learn how use computer programs (e.g. Consed, Glimmer, Genemark, BLAST and Phamerator) to identify all the viral genes and to make genome wide comparisons to identify evolutionary relationships. All course participants and the SEA staff are connected via the internet to share and analyze data, troubleshoot, and discuss results. Near the end of the spring semester, HHMI hosts a research symposium for course faculty and one student from each institution to report on their discoveries and experiences. As unique phages are characterized on campuses nationwide, students will have the opportunity to publish their research for the broader scientific community. This hands-on experience with the tools of Bioinformatics is an outstanding training opportunity and this unique program has helped the WKU Biology Department create an introductory pipeline to research experiences that complements our existing and planned instructional programs and allows freshman to engage in the thrill of discovery.

Complete Genome Sequences of 44 Arthrobacter Phages

ABSTRACT We report here the complete genome sequences of 44 phages infecting Arthrobacter sp. strain ATCC 21022. These phages have double-stranded DNA genomes with sizes ranging from 15,680 to 70,707 bp and G+C contents from 45.1% to 68.5%. All three tail types (belonging to the families Siphoviridae, Myoviridae, and Podoviridae) are represented.

Genome Sequences of Subcluster K5 Mycobacteriophages AlleyCat, Edugator, and Guillsminger

ABSTRACT Bacteriophages AlleyCat, Edugator, and Guillsminger were isolated on Mycobacterium smegmatis mc2155 from enriched soil samples. All are members of mycobacteriophage subcluster K5, with genomes of 62,112 to 63,344 bp. Each genome contains 92 to 99 predicted protein-coding genes and one tRNA. Guillsminger is the first mycobacteriophage to carry an IS1380 family transposon.

Sequence and annotation of the Wizard007 mycobacterium phage genome

Material and methods The Wizard007 bacteriophage was isolated from a soil sample taken at N36° 49’36.8” W87° 29’42.5” in Hopkinsville, KY. It was plaque purified on Mycobacterium smegmatis and DNA was isolated using the Promega Wizard DNA purification kit. The DNA was sequenced at the Virginia Commonwealth University by Roche 454 DNA sequencing. The genome quality was confirmed in Consed [1] and found to be 51,034 bp in length. Genes were called from the assembled sequence with a workflow consisting of Glimmer [2], GeneMark [3], tRNAscan [4], and SDFinder to identify potential gene features. Gbrowse [5], Apollo [6], and BLAST [7] were used to call and annotate genes. A 10 base 3’ overhang was found at the ends by ligating the genomic DNA and sequencing a PCR product produced across the ligated ends.