forest rohwer – Jonathan Eisen's Lab

Mini journal club: staged phage attack of a humanizes microbiome of mouse

Doing another mini journal club here. Just got notified of this paper through some automated Google Scholar searches: Gnotobiotic mouse model of phage–bacterial host dynamics in the human gut

Full citation: Reyes, A., Wu, M., McNulty, N. P., Rohwer, F. L., & Gordon, J. I. (2013). Gnotobiotic mouse model of phage–bacterial host dynamics in the human gut. Proceedings of the National Academy of Sciences, 201319470.

The paper seems pretty fascinating at first glance. Basically they built on the Jeff Gordon germ free mouse model and introduced a defined set of cultured microbes that came from humans. And then they stages a phage attack on the system and monitored the response of the community to the phage attack.

Figure 1 from Reyes et al.

They (of course) also did a control – in this case with heat killed phage. And they compared what happened to the live phage. I love this concept as they are able to control the microbial community and then test dynamics of how specific phage affect that community inside a living host. Very cool.

Story behind the paper: from Jeremy Barr on "Bacteriophage and mucus. Two unlikely entities, or an exceptional symbiosis? "

I am pleased to have a guest post in my “Story behind the paper” series. This one is from Jeremy Barr in Forest Rohwer’s lab about a new PNAS paper.

Bacteriophage and mucus. Two unlikely entities, or an exceptional symbiosis?

By Jeremy J. Barr

Our recent research at The Rohwer Lab at San Diego State University investigates a new symbiosis formed between bacteriophage, viruses that only infect and kill bacteria, and mucus, that slimy stuff coating your mouth, nose, lungs and gut.

Bacteriophage, or phage for short are ubiquitous throughout nature. They are found everywhere. So it shouldn’t surprise you to learn that these phage are also found within mucus. In fact, if you actually sat down and thought about the best place you would look for phage, you might have picked mucus as a great starting point. Mucus is loaded with bacteria, and like phage, is found everywhere. Almost every animal uses mucus, or a mucus-like substance, to protect its environmentally exposed epithelium from the surrounding environment. Phage in mucus is nothing novel.

But what if there were more phage in mucus? What if the phage, immotile though they may be, were actually sticking within it?

It turns out that there are more phage in mucus, over four times more phage, and this appears true across extremely divergent animal mucosa. But this apparent increase in phage could very simply be explained by increased replication due to access to increased bacterial hosts residing within mucus layers. But this assumption alone doesn’t hold up. Applying phage T4 to sterile tissue culture cells resulted in significantly more phage sticking to the cell lines that produced a mucus layer, compared to those that did not. There were no bacterial hosts for phage replication in these experiments. Yet still, more phage accumulated in mucus.

Surely the law of mass-action could explain this apparent accumulation. The more phage we apply to an aqueous external environment, the more phage will diffuse into and enter the mucus layer, being slowed in the process due to the gel-like properties, and eventually resulting in an apparent accumulation of phage in mucus. But when we removed mass-action from the equation, and simply coated mucus-macromolecules onto a surface, still more phage stuck. Our assumptions were too simple.
Phage are ingenious. They have evolved, traded, and disseminated biological solutions to almost every biological problem, whether we are aware of it or not. So in order to solve the phage-mucus quandary, we needed to look to one of the most ubiquitous and populous families of proteins found in nature: the immunoglobulin superfamily. This protein fold is so ubiquitous that it appears in almost every form of life. Within our own bodies, it is the protein that affords us immunological protection. Bacteria utilize the protein fold to adhere to each other, to surfaces, and as a form of communication. And as it would turn out, phage make an innovative use of the same protein fold to stick to mucus.

Immunoglobulin, phage and mucus, are all pervasive throughout environments. The interaction between these three entities forms a new symbiosis between phage and their animal hosts. This symbiosis contributes a previously unrecognized immune system that reduces bacterial numbers in mucus, and protects the animal host from attack. We call this symbiosis/immunity, Bacteriophage Adherence to Mucus, or BAM for short.

Our work is open access and available through PNAS .

If you would like to read further about BAM and its implications see these two commentaries by Carl Zimmer at National Geographic and by Ed Yong at Nature News

A Forest (Rohwer that is) on Black Reefs, Shipwrecks and Coral Reef Conservation

Well Forest Rohwer is at it again. He just is always doing something I find worth paying attention to.

First, he does fascinating and pioneering science on viruses in the environment. For example, consider that he was one of if not the first to do random shotgun metagenomics from environmental samples. See his lab’s 2001 and 2002 papers on the topic (Production of shotgun libraries using random amplification and Genomic analysis of uncultured marine viral communities) which I note came out before the Sargasso and Acid Mine Drainage papers which most cite as the first environmental shotgun sequencing pubs.

Second, he studies incredibly interesting topics that take him all over the world, mostly to coastal locations. See for example – Comparative metagenomics of microbial traits within oceanic viral communities and Microbial diversity associated with four functional groups of benthic reef algae and the reef-building coral Montastraea annularis and Viral and microbial community dynamics in four aquatic environments and Metagenomic and stable isotopic analyses of modern freshwater microbialites in Cuatro Ciénegas, Mexico and more.

Third, he also develops useful bioinformatics approaches to studying metagenomes. See Phylogenetic classification of short environmental DNA fragments and Power law rank-abundance models for marine phage communities and FastGroupII: a web-based bioinformatics platform for analyses of large 16S rDNA libraries and others.

In fact, you could say in many ways we do very similar work, except he focuses on viruses. Not that we always agree mind you. I once gave a talk after him at a meeting and I changed my title to “Seeing the Forest and Missing the Trees” in a little dig at his not using phylogenetic methods and in his approach to metagenomic analysis. But I digress.

What I want to write about today is a new paper from his lab: Black reefs: iron-induced phase shifts on coral reefs.

Alas, it is not freely available as it is in ISME but is not published under their “open” option. Am working on getting a link to an available PDF … will let everyone know.

Here is the abstract:

The Line Islands are calcium carbonate coral reef platforms located in iron-poor regions of the central Pacific. Natural terrestrial run-off of iron is non-existent and aerial deposition is extremely low. However, a number of ship groundings have occurred on these atolls. The reefs surrounding the shipwreck debris are characterized by high benthic cover of turf algae, macroalgae, cyanobacterial mats and corallimorphs, as well as particulate-laden, cloudy water. These sites also have very low coral and crustose coralline algal cover and are call black reefs because of the dark-colored benthic community and reduced clarity of the overlying water column. Here we use a combination of benthic surveys, chemistry, metagenomics and microcosms to investigate if and how shipwrecks initiate and maintain black reefs. Comparative surveys show that the live coral cover was reduced from 40 to 60% to 0.75 km2). The phase shift occurs rapidly; the Kingman black reef formed within 3 years of the ship grounding. Iron concentrations in algae tissue from the Millennium black reef site were six times higher than in algae collected from reference sites. Metagenomic sequencing of the Millennium Atoll black reef-associated microbial community was enriched in iron-associated virulence genes and known pathogens. Microcosm experiments showed that corals were killed by black reef rubble through microbial activity. Together these results demonstrate that shipwrecks and their associated iron pose significant threats to coral reefs in iron-limited regions.

Forest and others have recently been studying the Line Islands because they are relatively undisturbed reefs. Here are a short video about the work there (the work in general, not this specific study per se): http://oceantoday.noaa.gov/swf/flowplayer-latest.swf

Anyway, the new paper does something very different. It focuses on shipwrecks and the impact of these wrecks on reefs. This is of particular interest because as indicated in the abstract, the reefs are very low in iron. And many shipwrecks introduce massive amounts of iron. What they conclude in this new paper is that the iron from the shipwrecks leads to algal blooms, and lead to rapid killing of / damage to the pristine reefs.

For more on the paper there is an article in National Geographic Newswatch by Enric Sala worth checking out.

Forest also wrote me some information by email. He states:

Black reefs are associated with shipwrecks or other debris in this region of the world. These sites are interesting both from a conservation and scientific point of view. As a conservation issue, they are amazingly destructive. Kingman, one of the jewels of the USA coral reefs, has lost >1 km of the lagoon in less than 3 years. An old wreck on Fanning atoll has killed about 10% of their reef.

Visually, the black reefs are some of the eeriest places I’ve ever seen. The bottom is completely covered in different algae (including cyanobacterial mats), the water is filled with marine snow, and dark precipitate on the benthos (probably sulfur). We just published a paper in ISME where we have recreate the precipitate, cloudiness, and
coral death in microcosms by combining rubble from the black reefs, with corals and an iron addition. Addition of antibiotics blocks the coral death, precipitate, and marine snow, suggesting a microbial role.

The black reefs are probably caused by iron-enrichment from the wrecks and debris. We think black reefs are specific to non-emergent coral reefs, where iron is a limiting nutrient. Our current model is that iron stimulation of algae leads to increased microbial activity and coral death. In support of this, metagenomic analysis of the microbial community showed an enrichment of iron-related pathogenicity factors.

Forest also adds a plea to help in conservation of these reefs.

If you are interested in conservation, then please help us petition Congress to support removal of the wrecks and debris. Please contact Emily Douce at the Marine Conservation Biology Institute.

I encourage people to contact her.