Zetaproteobacteria: The Journey Continues

Lauren Rodgers, Rutgers University

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Findings: This past summer, we have been working furiously with one goal in mind: measuring the concentrations of iron in the sediments around Charleston to identify if they would be a good habitat for Zetaproteobacteria. We scoured Charleston for the perfect muddy sampling sights and battled the pluff, almost losing a few boots in the process, while also gaining lots of bug bites. We then spent hours in the lab, making solutions, extracting iron, running the spectrophotometer, and collecting data.  And now, after 10 long weeks, it is coming to an end.

This summer of researching Zetaproteobacteria with Heather, Alejandra, and Sarg has taught me so many things. One of which is that science does not happen overnight. You may have an idea of how you are going to do something, but when it comes to actually carrying it out, odds are that it will not go as you think. Science is a dynamic process. You are trying things out, failing, brainstorming for other ways to do things, revising methods, and most of all learning. And this part of the process is what makes it exciting. I experienced this first hand when it came to conducting the ferrozine assay. Much of the research detailing methods for a ferrozine assay were only written for liquid samples, not for sediment samples, so we had to come up with methodology for extracting iron from the sediments. As you can imagine, this took a lot of trial and error. Trial and error such as figuring out the hard way that the water you are using to make up the solutions is actually contaminated with iron, or that glass cuvettes tend to contain their own concentrations of iron in them as well. It was frustrating at times,  but this process was so important for me because it was one of the first times that I was able to take in ideas from many different sources and develop something of my own. It was hard work, but it was all worth it in the end. We were able to collect samples, successfully extract the iron, and measure the iron concentrations, gaining some very exciting results in the end.

What’s next for Zetaproteobacteria?

Now that we have optimized the ferrozine assay for measuring the concentrations of iron in the sediments, we can continue our research on Zetaproteobacteria. The first objective that we will work towards is identifying if Zetaproteobacteria are actually present in the sediments. If they are found, we will then quantify how many Zetaproteobacteria are actually present.

If Zetaproteobacteria are found in the sediments around Charleston, it could have many implications. The first implication is that they could be affecting the local iron cycle around Charleston through their transformations of Fe(II). Zetaproteobacteria have also been shown to be able to live on solid metal and use the Fe(II) present in it, quickening the metal’s rate of rusting. If they are found in Charleston, they could be speeding up the rusting of ships or even metal pipes. Lastly, their presence in Charleston would add evidence to their potential worldwide distribution.

The project from this summer may be finished, but the Zetaproteobacteria journey has just begun!

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The Fullerton Lab Team stopping to pose while sampling at Kearns Park on the Wando River. (p.c. Heather Fullerton)


I would like to thank my mentor, Dr. Heather Fullerton, for guiding me through this research, and my Fullerton Lab members for assisting me in the field and in the lab throughout the summer. I would also like to thank the National Science Foundation for funding this research as well as the College of Charleston and Grice Marine Lab for their support.


 

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Journey to the Center of the Pluff

Lauren Rodgers, Rutgers University

Version 2The Approach: In my previous blog post I discussed the importance of iron in ocean ecosystems. Because so many living things rely on iron to live and grow, it is important for us to understand how iron cycles, as it enters the ocean, exits the ocean, and changes from one form to another. Zetaproteobacteria are marine bacteria that rely on iron to create energy for themselves, but in this process, they also turn dissolved iron into solid iron. So these bacteria make rust as they grow. Unfortunately, rust isn’t very good for other organisms, and the Zetaproteobacteria effectually remove iron from the ocean. But still, these organisms are one half of the iron cycle and therefore play an prominent role. With our research, we aim to determine whether these bacteria are present in Charleston’s estuaries, and extrapolate how they might be impacting the local iron cycle.

Now, you most likely have one thing on your mind: How are they going to study all of this!? From our lofty research aims, we must simplify those down to into bite sized goals so we can have a successful summer of sampling.

Our Goals:

  1. Identify whether Zetaproteobacteria can be found in the sediments around Charleston.
  2. Measure the amount of Fe(II) and Fe(III) in the sediments

The first thing that we did in order to accomplish these goals is pick sampling sites. We wanted to sample the sediments for these Zetaproteobacteria, so we chose muddy regions close to tidal rivers that empty into the ocean. We wanted tidal rivers because Zetaproteobacteria live in salty waters, and these rivers mix with salt water from the ocean. We decided to look for these muddy regions along the Ashley River, Wando River, Stono River, and Cooper River, picking easily accessible sites far up the river where the water is fresher, midway down the river where the salt content is at a mid-range, and low down on the rivers, near the ocean, where the water is salty.

 

After identifying the sites that we wanted to sample at, we needed to figure out how to sample. We wanted to sample the mud at different depths, so we decided to use syringes to suck up the mud.

 

Once all of the samples were collected it was time to get back into the lab to analyze the data. In order to confirm the presence of Zetaproteobacteria we conducted PCR, which is a process that tells us if there was any DNA belonging to the Zetaproteobacteria in the samples.

 

 

 

To analyze the iron a ferrozine assay was conducted. In a ferrozine assay, different chemicals are added to the samples, which then turn different shades of purple depending on how much iron is present in them.

 

While we have already completed a lot of the data collection, we still have more to do. In the next few weeks we will focus on collecting the last few samples and analyzing them in the lab. Soon all of the results will be ready for interpretation!


I would like to thank my mentor, Dr. Heather Fullerton, for guiding me through this research. I would also like to thank the National Science Foundation for funding this research as well as the College of Charleston and Grice Marine Lab for their support.

Bacteria in the Ocean? That Eat Iron??

Lauren Rodgers, Rutgers University

Version 2The problem: Have you ever asked yourself, what is iron? It is an element? A rock? Some weird orange-ish substance? Is it the tool that you use to get the wrinkles out of clothes? And what does iron even do? Does it just sit there? Does anything eat it? Can we make things out of it? Iron is one of the most abundant elements on earth, yet not many people know much about the important role it plays in our lives.

Iron is more than just an element, or something found within a rock. It’s a nutrient, something necessary for the growth and metabolism of almost every living organism on Earth (Hedrich & Johnson, 2011). In the ocean, iron is found in two different forms, ferrous iron or Fe(II), which is soluble in water, and ferric iron or Fe(III), which is insoluble in water (Hedrich & Johnson, 2011). Because ferrous iron is soluble it is the form of iron that can be used by most organisms in the water (Hedrich & Johnson, 2011). This ferrous iron, however, is limited in the ocean despite its abundance in the Earth’s crust. In fact, Fe(II) is present only in incredibly small concentrations, making it a major limiting factor of growth for all of the plants and algae in the ocean. This is important because these plants and algae serve as the base of many food chains, so if there is a limitation on the growth of these organisms, it affects every other organism throughout the food chain. Though iron is an extremely important nutrient for many living organisms, it is still not well understood. One of the least understood aspects is how iron specifically cycles through different marine environments. Does it ever change form? Does anything add iron to the ocean? Does anything take iron out of the ocean? These questions bring us to Zetaproteobacteria.

Zetaproteobacteria is a recently discovered class of iron-oxidizing microbes. This just means that the bacteria eat iron in the form of Fe(II) and produce Fe(III) as a waste product (Emerson et al., 2007; Chiu et al., 2017). In fact, these waste products can take on the form of hollow tubes, also called tubular sheaths, or twisted stalks that you can see under the microscope!

 

Zetaproteobacteria were initially described in 2007 near hydrothermal vents, utilizing the large concentrations of Fe(II) that were present in the fluid that spewed from the vents (Emerson et al., 2007).

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Iron mat composed of Zetaproteobacteria on a lava rock near the submarine Loihi volcano. (A. Malahoff, Hawaii, Loihi Volcano, July 1988)

How do Zetaproteobacteria relate to the cycling of iron? 

Zetaproteobacteria, with their role in eating iron and transforming it from its soluble Fe(II) state into its insoluble Fe(III) form may have an important role in the cycling of iron through the environment, functioning as an important source of iron removal.

Since their discovery, Zetaproteobacteria have also been observed in many other habitats, including coastal estuarine habitats with lower levels of iron, similar to that of Charleston, SC. (Laufer et al., 2017; Chiu et al., 2017). Our study will try to identify if these Zetaproteobacteria are present in the muddy soils around Charleston, as well as measure the levels of Fe(II) and Fe(III) in the rivers where these bacteria may be found.

 

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Hopefully, through the study of the distribution of Zetaproteobacteria across the globe, including the chemical characteristics of the different environments that they inhabit, we may get a clearer picture of how iron cycles in aquatic environments and the role that these Zetaproteobacteria play.


I would like to thank my mentor, Dr. Heather Fullerton, for guiding me through this research. I would also like to thank the National Science Foundation for funding this research as well as the College of Charleston and Grice Marine Lab for their support.


References 

Chiu, B. K., Kato, S., McAllister, S. M., Field, E. K., & Chan, C. S. (2017). Novel pelagic iron-oxidizing Zetaproteobacteria from the Chesapeake Bay oxic-anoxic transition zone. Frontiers in Microbiology, 8(JUL), 1–16. https://doi.org/10.3389/fmicb.2017.01280

Emerson, D., Rentz, J. A., Lilburn, T. G., Davis, R. E., Aldrich, H., Chan, C. S., & Moyer, C. L. (2007). A novel lineage of proteobacteria involved in formation of marine Fe-oxidizing microbial mat communities. PLoS ONE, 2(8), e667. https://doi.org/10.1371/journal.pone.0000667

Hedrich, S., Schlömann, M., & Johnson, D. B. (2011). The iron-oxidizing proteobacteria. Microbiology,157(6), 1551–1564.

Laufer, K., Nordhoff, M., Halama, M., Martinez, R. ., Obst, M., Nowak, M., … Kappler, A. (2017). Microaerophilic Fe(II)-oxidizing Zetaproteobacteriaisolated from low-Fe marine coastal sediments – physiology and characterization of their twisted stalks. Applied and Environmental Microbiology, 83(February), AEM.03118-16. https://doi.org/10.1128/AEM.03118-16

Mori, J. F., Scott, J. J., Hager, K. W., Moyer, C. L., Küsel, K., & Emerson, D. (2017). Physiological and ecological implications of an iron- or hydrogen-oxidizing member of the Zetaproteobacteria, Ghiorsea bivora, gen. nov., sp. Nov. ISME Journal, 11(11), 2624–2636. https://doi.org/10.1038/ismej.2017.132