Final exam - Spring 1999

1. What are the 3 primary evolutionary branches of life? (5 points)

Bacteria, Archaea, and Eukarya

2. Describe the likely properties of the last common ancestor - the organism that gave rise to the 3 primary evolutionary branches of life. (5 points)

The last common ancestor must have shared the properties common to the three extant branches of life, including:

3. What is meant by the term "the RNA world"? (5 points)

The RNA World was the time (if it existed) before the evolutionary invention of DNA or protein, when RNA served as both genome and catalyst for primative living things.

4. Describe how one of the enrichment schemes we used in lab was designed to favor the growth of the targeted organisms. (5 points)

e.g. the Yeast/fungus enrichment.

Basic rich media was used, but the pH was low, favoring yeast/fungi over many types of Bacteria. In addition, a broad-spectrum antibiotic was included to kill most the the remaining Bacteria.

5. Describe how another of the enrichment schemes we used in lab was designed to favor the growth of the targeted organisms. (5 points)

e.g. the purple non-sulfur enrichment.

The media contained glycerol and bicarbonate as the only carbon sources, and cultures were incubated in sealed tubes without air but in the light. Respirative growth on glycerol quickly removes oxygen from the media, but glycerol is a non-fermentable growth substrate, so about the only way to grow left is photosynthetically. Since the media is now anerobic, eukaryotic algae and cyanobactria can't grow, and there isn't sulfide for purple sulfur Bacteria - leaving purple non-sulfurs as about the only thing that can grow.

6. What is a microenvironment? How do microenvironments contribute to the diversity of microbial communities? (5 points)

Microenvironments are just the details of the environment at small scales. For example, in a generally aerobic environment, there are certainly nooks and crannies of anaerobic conditions. The wide range of conditions present at the micro-scale support the presence of all kinds of organisms (and a broad range of interractions between them) in environments that to us don't seem approprite for their growth.

7. Why are enrichments inoculated with very dilute samples from an environment more likely to yield cultures of the predominant strains present in that environment than heavily-inoculated enrichments? (5 points)

Heavily inoculated enrichments are likely to contain large numbers of related species of a guild. The one that grows fastest in captivity will probably take over the culture, even if it represents a trivial fraction of the original sample. Dilutely-inoculated enrichments, on the other hand, should contain only those strains that are abundant in the environment, allowing their growth in the absence of competition from the 'weeds'.

8. What are the 3 general explanations for the origin of viruses.(5 points)

9. Briefly compare the 'Oparin ocean' hypothesis for the origin of life with Wäctershäuser's alternative 'surface metabolism' hypothesis. (10 points)

In the Oparin Ocean hypothesis, small organic material collected in the early ocean to high enough concentrations that spontaneous polymerization resulted in the formation of active biopolymeres - DNA, RNA, and protein - that could organize into simple living things. It is true that lots of organics must have been imported by comet influx and created de novo, but not likely so much as to create a 'primordial soup'. In any case, hydrolysis is strongly favored over polymerization in solution, so polymers wouldn't accumulate

In the surface metabolism hypothesis, charged organic compounds at low concentrations in the ocean were concentrated by adsorbtion onto pyrite or silica surfaces. On a surface polymerization is favored over hydrolysis, even using unactivated monomers, and surface chemistry is stereospecific unlike most solution chemistries.

10. Compare conjugative transfer of the F plasmid with the infective transfer of bacteriophage M13. How are these processes similar? How are they different? (10 points)

The F plasmid carries a series of genes that direct replication & conjugative transfer of the plasmid. The donor and recipient come into direct contact, and one strand of the plasmid is transfered via a vplasmid-encoded protein pore. M13 is very similar in that transfer is directed by a series of virus-encoded proteins, but transfer of the DNA is via a protein-encapsulated form rather than by direct contact.

11. As a microbiologist at the Centers for Disease Control, you are faced with an outbreak characterized by unusual and painful skin lesions. Patients do not respond to general antibiotic treatment, and attempts to culture Bacteria from lesions has failed. However, histology of the lesions shows the clear presence of Bacteria inside the human cells. How would you attempt to identify this pathogen, and how would you confirm that identification? (10 points)

I would use bacterial-specific PCR primers to amplify the ssu-rRNA gene from the pathogen from a lesion sample (or you could homogenize the tissue, purify the bacterial cells, and amplify from them using 'universal' primers). Sequencing and phylogenetic analysis of the ssu-rRNA would tell you what the organism is, or at least what it is related to. A specific fluorescently-labeled oligonucleotide probe based on this sequence could be used in in situ hybridization to make sure the sequence obtained really came from the pathogen - the bacterial cells in a sample of lesion should 'light-up' with the probe.

12. Briefly summarize one of these papers: (10 points)

e.g. the Hugenholtx paper.

DNA was extracted sediment samples of Obsidian pool. The samples were amplified using SSU-rRNA primers and 122 clones were sequenced. 54 of these were found to represent distinct bacterial sequence types (> 98% identity). 38 of these were found to be affiliated with recognized bacterial divisions. The remaining 16 were found to be unaffiliated with recognized bacterial divisions, representing 12 novel candidate bacterial divisions.

It was a surprise that no archaeal sequences were detect among the clones that were originally developed using universal PCR primers. Archaea are commonly thought to be the dominating microflora in hot springs. To test whether these findings were correct, bacterial- and archaeal-specific probes were hybridized to rDNAs from Obsidian pool to estimate the relative abundance of bacteria and archaea. Bacteria were found to dominate the Obsidian pool community by 75:1 over Archaea.

13. You have discovered that the dinoflagellate Histeria pesticida is the cause of massive fish kills and selective memory loss in low-level government officials. The organism cannot be grown in pure culture, but you have carefully described a very complex life cycle that includes an unbelievably wide range of morphological stages. Unfortunately, your competitors don't believe it, and are claiming that many of these "morphological stages" are just other species of microbes. How would you test your belief that these different morphological stages are all the same species? (10 points)

I would amplify & sequence the ssu-rRNA from an unmistakenly dinoflagellate morphological form of the organism and make a fluorescently-labeled oligonuclotide probe. Since the organism cannot be grow in pure culture, you would have to get them as 'clean' as you could, and then sort through the sequences you got for the one that looks like it comes from a dinoflagellate. In situ hybridization to cells in the different 'morphological stages' would readily tell you whether they are Histeria or not.

14. You are a microbiologist at North Carolina State University. The creation of a 'Genomics Center' on Centennial Campus was going smoothly until a competing group pointed out that nobody at NCSU was actually sequencing the genome of anything. The Dean of Research just called you up and gave you $2 million and told you to start sequencing the genome of an agriculturally-important microbe. What organism would you choose, and how would you justify spending $2 million on that genome? (10 points)

e.g. Methanosarcina barkeri

This is a critically important microbe because of its role in the conversion of small organics into methane and CO2 in the anaerobic digestion phase of wastewater treatment (both human and animal waste treatment systems). The information in this genome would tell us a lot about how the organism works, hopefully so that waste treatment could be dramatically improved.

Last updated May 6, 1999 by JWBrown
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