3.25.19 Creating a Research Question

Rationale: As the poster has been predominantly finished, the class will now move on to independent research projects. Therefore, it is necessary to develop a research question to guide our research moving forward.

Tools:

Our group narrowed down our search to focus mainly on repeats and how they may reveal functions about genes with no function. To find these repeats, we downloaded the Gepard dotplot software that will allow us to find areas of NapoleonB’s genome.

Results:

Four questions were successfully developed and submitted, and the Gepard software was successfully downloaded and usage was practiced to build confidence before research formally begins. The main question that our group would like to focus on is “Is it possible to use coding or noncoding repeats in the genomes of AM cluster Arthrobacter phages to predict the functions of genes with no known function?”. There are components of this question that will be refined as preliminary research is done, but this question seemed to our group to fit each component of a good research question.

Conclusion:

Research questions need to be specific, as initially looking at the immense genome of NapoleonB and all the options seemed a daunting task. By creating a specific question, results are likely to be focused and easier to process, which eliminates doing inefficient work that will result in less meaningful results.

Next Steps:

The Gepard software will be used to create a dot plot of NapoleonB’s genome and begin to reveal where there are repeats. These results will be analyzed and used to hone our research question.

3.20.19 Reflecting on Poster and Introducing Individual Products

Rationale: After reaching a consensus on the main points of the final poster and reaching a stage where only minor personal changes were necessary, it was found to be beneficial to reflect on the process to help learning for future posters. Also, since an independent research project is required, the project was introduced and brainstorming began.

Tools/Procedure: Google slides for presentation and Microsoft Word for QTM. As an independent project group, we finished our QTMs independently, then began discussions about the topic for our independent project and determined what should be researched to develop a proper question.

Results: The QTM was successfully completed and a topic to research was chosen.

Conclusion: Since a topic to research has been determined, it will be possible to spend more time honing a broader topic to get a developed and polished research question. Therefore, this strategy will help to ensure the results of the experiment end up being useful and interesting.

Next Steps: The next steps of the project will be to investigate the literature about this topic and later finalize the question before lab on Monday.

3.18.19 Finalizing Poster

Rationale: Since the poster had a solid beginning and an understanding of what needed to be done, it was necessary to all play roles in wrapping up the loose ends of the poster. Therefore, I worked on ensuring the formatting and shapes were all consistent and optimal for displaying the ideas of our project.

Tools and Procedure:

Google Slides was used to collaborate on the final version of the poster to ensure that everyone could access the project in real-time.

Results:

The shapes were reconfigured twice because there was a small error in the scaling that required all of the work to be redone. Also, the shape for the in silico results section was changed to a more optimal L-shape to ensure that all of the figures and data could be incorporated.

Conclusions:

Editing the formatting of posters is a very tedious, yet satisfying job that requires a lot of attention to detail. While formatting rarely makes a poster more influential or memorable (the results or procedure tend to do that), it can detract from the information being presented which can damage the overall presentation. Therefore, it was important to make sure everything was done correctly to give the poster the best possible chance to succeed.

Next Steps:

After the formatting was fixed, there are still some final tweaks that need to be added on the rest of the poster. Once these tweaks are added and inputted, the poster will be complete and ready for a final check before printing and presenting.

1. Having a state health system was particularly damaging to the consistent treatment of infectious diseases in Russia. Since the state was liable for paying for care of patients, both economic and political factors became important when determining the course of care a patient would receive. During World War II, antibiotics were presented from western countries to the Soviet Union for the first time. Their success stories were very compelling to citizens and doctors, but their high cost proved to be an issue for the Soviet Government. This led to hospitals operating with only a small portion of antibiotics that were created, and of that portion they consistently were out of types of antibiotic. This economic shortage implicated politics, as the Russian government did not want to appear weak or unable to care for their own citizens. Therefore, they created propaganda that favored alternative, or home-grown, remedies in order to encourage more citizens to use methods that did not include antibiotics. This also allowed bacteriophage therapy to coexist with the limited antibiotics that were being used, as they were less expensive to produce.
2. Across Europe, many institutes were created to further investigate bacteriophages and phage therapy. Interest with these organization lies mainly within three highlighted groups: Eliava Institute, Hirszfeld Institute, and Phage Therapy Center. The Hirszfeld and Eliava Institutes share similar struggles with Russian authorities. As examined in the first discussion, the Eliava Institute struggled with Russian politics, as Eliava himself petitioned the wrong individuals for permissions, and the politicians that were snubbed worked against him and eventually led to his arrest and execution. The Hirszfeld Institute also struggled with Russian rule in its developmental stages. Ludwik Hirszfeld constantly battled against Lysenkoism, which was staunchly supported by Stalin. Any other theory on biology was greatly discouraged by Russian authorities, and holding the beliefs that Hirszfeld did about biology was considered dangerous and a slight against authority. Therefore, Hirszfeld was not able to freely publish his work, which slowed the development of his petitions for an institute. His petitions were further damaged after a group of mainly Jewish scientists and doctors were convicted for a plot to poison government officials. Due to the Jewish linage present in Hirszfeld and his associates, there was fear that they would be soon persecuted as well. Again, this worked against the creation of a Hirszfeld institute. Thus, the Eliava and Hirszfeld Institutes both struggled under Russian rule. The third institute examined was the Phage Therapy Center. When compared with the Eliava Institute Today, it becomes apparent that while they overlap on ideas and missions, their core goals differ. In other words, the Eliava Institute contributes more towards the research and understanding of bacteriophages and their potential use in phage therapy while the Phage Therapy Center is mostly concerned with treating individuals who have conditions that are better treated by phage therapy (stated on their website as conditions localized to specific areas of lower blood flow, acute or chronic bacterial infections, or cases that implicate bacteria resistant to antibiotics). Historically, however, the Eliava Institute was very involved with the treatment of bacterial infections and diseases with bacteriophages.
3. A large challenge that Merril had when he attempted to use phage therapy as a treatment was that the majority of the bacteriophages used were instantly filtered out as by the liver and spleen. This led to the development of the serial passage technique. First, Merril and his team chose lambda phage, a very well-known and understood bacteriophage because of the work by Delbruck, for the experiment. This allowed them to control for variables that would otherwise have been unknown. Next, the group injected infected blood with a large amount of bacteriophage. The bacteriophage that made it through the mice was harvested seven hours later, and the process was repeated many times until a large amount of bacteriophage that was viable could be used to combat the disease in the organism. The comparison between bacteriophages that had undergone serial passage and those that had not was clearly shown in a figure of the PNAS paper to have approximately twice the concentration after 25 hours of being in the infected mouse. This portion proved that a phage resistant to being filtered out by internal organs could be obtained. The next step was to prove that mice with E. coli could actually be saved by the phage treatment. Therefore, they created an experiment where some mice with E. coli received phage treatment and others did not. When the phage treatment worked, they concluded that the newly passaged phages were effective in treating a relatively straightforward disease. These results were displayed in the PNAS paper in a line graph that compared symptoms to time; mice that received phage treatment with passaged phages Argo1 and Argo2 were healed within 100 hours, and those that were not died due to the infection. To continue to investigate the magnitude of the discovery, they investigated how phages could combat a more dangerous type of bacteria (VRE). The group proved that the phages were capable of treating this strain of more severe bacterial infection. Lastly, and of perhaps greater concern to critics of phage therapy and the FDA, the group managed to show that bacteriophage, not the immune system, was the agent that was causing the results by introducing heat-killed bacteriophages that would stimulate the immune response, but not kill any of the bacterial cells. When the same effectiveness was not shown with the heat-killed phages, it was concluded that the phages had some effect in combating the infections.
4. GangaGen was described in the text as a company originating in India due to the finding of bacteriophages in the Ganges River (which resulted in the name of the company). Due to the attractiveness of phages, the company attracted investors before any real ideas had been developed. The other companies, like GangaGen, are focused on developing products that can help to treat disorders with phages rather than other treatments and changing bacteriophages from an alternative treatment to a tool that can be used to combat modern medical problems. These treatments could be particularly useful, as antibiotic-resistant strains of bacteria are becoming more prevalent and cannot be easily solved with current technologies. Experiments proving bacteriophage effectiveness in combatting modern antibiotic-resistant strains along with other conditions where bacteriophages would be less expensive than antibiotics would allow doctors and individuals being treated to have more options, which can often lead to better care. The additional options would be achieved by the specificity of bacteriophages, as doctors or patients would not need to rely on extrapolating an antibiotic to try to combat an evolved strain of bacteria.

3.6.19 Poster Presentations

Rationale: Since each group has created a poster that is ready to display for the class, it is necessary for the class to decide which format is chosen to create the final poster.

Procedure:

Each group stood in front of the class and gave a brief presentation. The presentations were followed by discussions about each posters. After the discussions, a class-wide vote and conclusion was reached about the poster style.

Results:

One poster was selected from the entire class, but aspects of each poster were chosen to be added to that poster. Therefore, even though only one group’s base design will be chosen for the entire poster, each group/individual will have the opportunity to have a role in creating a product representative of the year’s work.

Conclusions:

With so many people working on one project, splitting into groups was especially helpful in ensuring that each person had their ideas heard, and that they had a chance to be seen on a poster, even if it was a draft.

Next Steps: The poster will be refined until there is a solid final product that is ready to be submitted to URSA.

3.4.19 Poster Creation

Rationale: Since the posters were half-done, the next step was to refine/revise the posters and combine with another group.

Procedure:

Powerpoint was used to finish the poster. Email was used to transfer information between individuals in the group. Personally, I contributed to the In Silico Results section by making graphics in excel and taking screenshots of the programs.

Results:

A complete poster that was fused with another group’s was created. This poster is a rough draft, and still needs to be revised in the future. However, it will provide a basis for the rest of the lab and deciding which poster to use and proceed with.

Conclusions:

Making posters is more difficult in a shortened/expedited time frame. Therefore, the poster cannot be expected to be perfected in two days, so additional collaboration and classwork will need to happen before a final product will be ready to submit.

Next Steps:

On Wednesday, the posters with the main groups will be presented so the class can evaluate the posters and determine which poster should be edited into a final version. The poster that results from this process will be submitted for the research symposium.

2.27.19 Creating PowerPoint Poster

Rationale: After the paper sketch was returned, the next step was to transfer the ideas to a PowerPoint slide (the typical tool used to make scientific posters). Therefore, today was dedicated to beginning the process of creating a PowerPoint version of what had been established on Monday.

Procedure:

Sketch was analyzed and revised. Work was split up amongst the three group members  and different sections were created today. The sections were worked upon until the end of lab.

Results:

A PowerPoint slide with general content was created today. The time permitted was not nearly enough to compile all of the content needed for a complete poster, so only some of the content was present on the submitted copy.

Conclusions:

Creating a poster is very difficult to do in an hour and a half. The process is far longer and requires a lot of attention and planning. In the planning process, it would have been more beneficial to focus more on content and less on how the information would look on the slide. However, a good beginning to a slide was achieved today and will provide a foundation for future work.

Next Steps:

Information that was put on the slide today will need revision, and the rest of the information that had yet to be added will need to be analyzed and added during the next opportunity.

2.25.19 Sketching Poster

Rationale: Since a poster is the most common form of presentation for URSA, it was necessary to learn more about what successful posters look like and to begin to brainstorm what our own posters may look like eventually. Therefore, we looked at many past posters and were assigned into the groups we brainstormed our own posters in.

Procedure:

As a class, we reviewed many posters for coloring and content. We discussed things we liked and those we didn’t. Then, we were assigned into groups and in those groups, we discussed and planned important content we wanted to include on the posters. Finally, we created a visual representation on paper of what we would like our posters to look like.

Results:

This lab session resulted in a more complete understanding of what a research poster entails. In addition, this period also resulted in us developing a sketch about what our own posters would look like in small groups.

Conclusions:

Posters have far more moving parts than initially anticipated and take a lot of thought and effort. They are benefitted by careful planning and analysis before action, and many review processes are required before a final product can be submitted.

Next Steps:

On Wednesday, a PowerPoint version of the poster brainstormed will be created. This is the next step in beginning to create a poster; the product will need to be reviewed and changed many times before it can be fully submitted.

2.20.19 Checking NapoleonB – Part 2

Rationale: Since part of the annotation process is checking work, it is necessary to return to a complete annotation with fresh eyes and examine it again. Therefore, the class looked over the majority of the genes in NapoleonB that had been called once more to check to ensure annotations were properly done.

Tools/Procedure:

• Genemark
• DNA Master
• NCBI BLAST
• PhagesDB Blast

1. Opened PhageNotes and DNA Master
2. Checked and discussed each gene that was questionable
3. Ran a separate Genemark with the correct species

Results:

The time in the lab today resulted in portions of calls being changed for a few genes that had been mistyped or had errors with the calls. The class had failed to use the proper species when using Genemark for the first time, so the correct species was used and some results were changed because of this.

Conclusions:

The main conclusion from today’s lab time was that the species is significant in the Genemark system. This was a previously forgotten fact, and it would have been detrimental to the annotations if it had gone unnoticed. Furthermore, today proved that one check was not enough, as the class found more discrepancies that were missed the first time around.

Next Steps:

During the next lab time, the independent projects will be examined more closely along with poster & abstract ideas for Scholar’s Week. Furthermore, it would be beneficial to examine NapoleonB with more detail than was done today.

2.18.19 NapoleonB Annotation Checking & Abstracts

Rationale: Since the genome annotations were completed for NapoleonB, it was possible to check the annotations that had been done. Therefore, it was pertinent to do so to ensure that no incomplete or inaccurate information was being placed in the database. Furthermore, abstracts were also presented to the group to begin to finalize the submission to the Scholar’s Week conference.

Tools/Procedure:

• DNA Master
• HHPred
• PhageNotes
• NCBI Blast
• PhagesDB Blast

In the lab, genes that were listed to have questionable calls were checked over. For my group, genes 50, 1, and 2 were examined. A new gene was added and confirmed with Lathan. After these calls had been made, the abstracts were edited and submitted for the group rather than individually.

Results:

Gene 50 was determined to have been called correctly. Genes 1 & 2 had a large gap which was examined and eventually determined that there was supposed to be a reverse gene to account for the large gap. This gene’s annotation is found below. Apart from these two calls, the genes appeared to have been done correctly.

SSC:372 – 569, CP:No No coding potential was present, SCS:Neither, ST:NA, BLAST-Start:Aligns with circum gp2 NCBI BLAST q1:s1 100 2E-41, Aligns with Tribby gp2 PhagesDB BLAST q1:s1 1 6E-35, Gap:first gene, LO:Yes, RBS:Kibbler7 and Karlin Medium 3.263 -2.059 Yes, F:NKF, SIF-BLAST:NKF, SIF-HHPred:NKF, SIF-Syn:NKF

Conclusions:

Since the genome was checked in part by everyone, it can be assumed that NapoleonB has been fully sequenced and is close to being ready to submit with annotations. The abstract was also successfully assembled, which means that the group is closer to developing Scholar’s Week work.

Next Steps: Scholar’s Week work will be continued and abstracts/posters will continue to be developed throughout the week.