Final Exam : In-class questions | MB 451 Microbial Diversity · Final Exam : Take-home questions | MB 451 Microbial Diversity The Rules : You are allowed to use any book or online

Final Exam : In-class questions | MB 451 Microbial Diversity

Honor pledge: “I have neither given nor received unauthorized aid on this test.”

Name : ____________________________________________________________ Date : __________________________

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

Multiple-choice questions (1 points each)

5. ____ How did Reysenbach (and co-workers) confirm that the sequence EM17 really was the pink filamentous organism?

A. Denaturing gradient gel electrophoresis

B. Fluorescent in situ hybridization

C. terminal restriction fragment length polymorphism

D. real-time polymerase chain reaction

E. all of the above

6. ____ In Denaturing Gradient Gel Electrophoresis (DGGE), DNA molecules are separated by...

A. G+C content

B. denaturation point

C. length

D. sequence complexity

E. none 0f the above

7. ____ The carbon-fixing symbionts in the scaly snail reside...

A. in their gills

B. in the surface layer of their scales

C. in their liver and spleen

D. in their muscular foot

E. in their esophageal glands

8. ____ Which of the following is not one of the four phyla most abundant in/on the human body?

A. Bacteroids

B. Proteobacteria

C. Firmicutes

D. Cyanobacteria

E. Actinobacteria

9. ____ Which of the following human microbiomes cannot be distinguished?

A. male from female

B. dominant hand from non-dominant hand

C. oral cavity from gut

D. one individual from another

E. all of these can be distinguished

10. ____ If you want to determine in a Stable Isotope Probing (SIP) experiment who in an environment is fixing nitrogen, the stable-isotope probe would be ...

A. 13CO2

B. a 32P-labeled rRNA primer

C. 15N2

D. 133Cs-tetrafluoroacetate

E. none of the above

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11. ____ What approach did Yim (and co-workers) use to examine the microbial communities from the Tibetan hot spring?

A. Denaturing gradient gel electrophoresis

B. Fluorescent in situ hybridization

C. terminal restriction fragment length polymorphism

D. real-time polymerase chain reaction

E. all of the above

12. _____ Proteorhodopsin is a …

A. sensory opsin

B. light-driven chloride pump

C. light-driven proton pump

D. light-detecting calcium channel

E. pseudogene

13. ____ Which of the following unicellular eukaryotes (protists) was found by Moreno (and co-workers) to preferentially feed on autotrophic ammonia-oxidizing microbes in wastewater sludge?

A. Arcella

B. Chaos

C. Epistylus

D. Spumella

E. Hartmanella

14. ____ What was the primary method used by Sakamoto (and co-workers) to to compare oral microbial populations before and after periodontal treatment?

A. FISH

B. DGGE

C. tRFLP

D. rtPCR

E. SIP

Short-answer questions (5 points each)

15. What is “horizontal gene transfer” and list 3 ways horizontally transferred genes be detected.

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16. Describe in detail the most interesting thing you learned from any of the papers we reviewed in class. Make sure to include why you think this is interesting - what does it mean to you? This is a substantial question - please give a substantial answer.

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XKCD.com/1664/

17. Describe one of the following technologies: stable-isotope probing (SIP), terminal RFLP (tRFLP), or denaturing gradient gel electrophoresis (DGGE).

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Essay question (10 points)

18. Describe in detail any paper we’ve discussed in this course. You are allowed to use papers we reviewed in the Discussion session if you wish.

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Papers reviewed in class:

• Reysenbach AL, Wickham GS & Pace NR 1994 Phylogenetic analysis of the hyperthermophilic pink filament community in Octopus Spring, Yellowstone National Park. Appl. Env. Microbiol. 60:2133-2199

• Huber R, et al. 1998 Thermocrinus ruber, gen. nov., sp. nov., a pink-filament-forming hyperthemophilic bacterium isolated from Yellowstone National Park. Appl. Env. Microbiol. 64:3576-3583

• Goffredi SK, Waren A, Orphan VJ, Van Dover CL & Vrijenhoek RC 2004 Novel forms of structural integration between microbes and a hydrothermal vent gastropod from the Indian Ocean. Appl Env Microbiol 70:3082-3090

• Costello EK, Lauber CL, Hamady M, Fierer N gordon JI & Knight R 2009 Bacterial community variation in human body habitats across space and time. Science 326:1694-1697

• Yim LC, Hongmei J, Aitchison JC & Pointing SB 2006 Highly diverse community structure in a remote central Tibetan geothermal spring does not display monotonic variation to thermal stress. FEMS Microbiol Ecol. 57:80-91

• Sakamoto M, Huang Y, Ohnishi M, Umeda M, Ishikawa I & Benno Y. 2004 Changes in oral microbial profiles after peridontal treatment as determined by molecular analysis of 16S rRNA genes. J. Med. Microbiol. 53:563-571.

• Moreno Am, Matz C, Kjelleberg S & Manefield M 2010 Identification of ciliate grazers of autotrophic Bacteria in ammonia-oxidizing activated sludge by RNA stable isotope probing. AEM 76:2203-2211

• Beja O, et al. 2000 Bacterial rhodopsin: Evidence for a new type of phototrophy in the sea. Science 289:1902-1906.

• Nelson KE, . . . Venter JC, Fraser CM. 1999. Evidence for lateral gene transfer between Archaea and bacteria from genome sequence of Thermotoga maritima. Nature 399:323-329.

• Venter JC, et al., 2004 Environmental genome shotgun sequencing of the Sargasso sea. Science 304:66-74.

• Rasmussen, B. 2000 Filamentous microfossils in a 3,235-million-year-old volcanogenic massive sulphide deposit. Nature 405:676-679

Papers reviewed in the Discussion sessions this semester (you can use these if you wish):

• Murgia C, Pritchard JK, Kim SY, Fassati A, and RA Weiss 2006 Clonal origin and evolution of a transmissible cancer. Cell 126:477-487

• Rault D, et al. 2008 Nanobacteria are mineralo fetuin complexes. PLOS Pathogens 4:e41

• Giovannoni SJ, et al. 2005 Genome streamlining in a cosmopolitan oceanic bacterium. Science 309:1242

• Fierer N, Hamady M, Lauber CL and R Knight 2008 The influence of sex, handedness, and washing on the diversity of hand surface bacteria. PNAS 105:17994-17999

• Cyanobacteria use micro-optics to sense light direction. 2016. Scourgers N, et al. eLife 5:e12620

• RASMUSSEN ET AL., EARLY DIVERGENT STRAINS OF YERSINA PESTIS IN EURASIA 5,000 YEARS AGO. CELL 2015 163:571-582

• Hug LA, et al. 2016. A new view of the tree of life. Nat. Microbiol. 48: Article 16048 (11 April 2016)

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Final Exam : Take-home questions | MB 451 Microbial Diversity

The Rules : You are allowed to use any book or online resources to help answer these questions. Your are not allowed to talk with or work with anyone else (whether they are students in this course or not) about these questions.

Name : __________________________________________________________________ Date : ________________________________

1. Imagine you have woodlice (also known as sow bugs, pill bugs, roly-polys) in Microbial Diversity lab. Droppings from these bugs examined under a microscope reveal large numbers of filamentous organisms that look like Bacillus (not Clostridium), complete with endospores. When you plate bug droppings out on PYD and incubate aerobically, you get lots of identical-looking Bacillus colonies, but the cells are typical individual cells, or at most pairs. The ssu-rRNA sequences from these colonies are a perfect match to B. cereus, a common soil Bacillus species. Your TA tells you to keep trying, looking for a different colony type, until you get the filamentous Bacillus to grow on plates. But it doesn’t seem to work, you keep getting B. cereus and random non-Bacillus things, no filamentous organisms. You think that the filamentous Bacillus might not be able to grow on the media you’re using, and so the only thing you get on plates is easy-to-grow B. cereus that’s also in the droppings. But Dr. Brown suggests that B. cereus might be pleomorphic; that it might grow as filaments in bug intestines (and so in the droppings) but as individual cells in culture; in other words that the individual-celled B. cereus you’re growing might actually be the the same thing as the filamen-tous Bacillus in the bug droppings. You decide to figure it out once and for all in a research lab the following semester. How would you go about this? What would the results look like either way? Are there other alternatives? Be sure to provide important experimental details. (20 points)

2. Imagine you’ve discovered a novel species of fish in a deep-sea methane (CH4) seep. This fish completely lacks a digestive tract, including both gullet and anus - the mouth opens to the gills but no further. However, it gets along just fine, swimming around in methane-infused water. You hypothesize that it’s absorbing both methane and oxygen from the water with its gills, and is living by methane oxidation (CH4 + O2 -> CO2 + H2O) rather than eating. Recognizing that the fish also needs an organic nitrogen source (which would normally be acquired in the diet), you further hypothesize that the fish is somehow getting nitrogen from the N2 also present in significant amounts in the seep environment. You dissect one of these fish, and discover a grossly enlarged liver filling the space where the GI tract normally would be. Microscopic examination of the cells of this organ show both spherical and rod-shaped bacterial endosymbionts, and a rod-shaped bacterial symbiont in the interstitial spaces. (1) How would you iden-tify these apparent symbionts, and (2) how would you determine which (if any) of these are carrying out methane oxidation and/or fixing nitrogen? There is no need to describe how the general techniques involved work, but be sure to provide the specifics required to make these techniques provide the answers you’re looking for. Note that neither the bacteria nor the fish can live in the lab, and so stable-isotope probing (SIP) is not an option. Be sure to tell me exactly what you’re looking for in your experi-ments - be specific. (20 points)

3. In the paper “Changes in oral microbial profiles after periodontal treatment as determined by molecular analysis of 16S rRNA genes” by Sakamoto, et al, the authors use t-RFLP, with some help from realtime PCR and traditional ssu-rRNA clone-and-se-quence approaches, to see how periodontal microflora in periodontal disease patients changed after treatment. This paper is now more than 10 years old. How would you do this experiment today with modern techniques? (20 points).

The common wood louse

(suborder Oniscidea)

Bacillus sp. (?) observed in bug feces

Bacillus cereus cultivated from bug feces

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Question #1

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Don’t Panic!

Question #2

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Question #3

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