The Bowman Lab

Congratulations to Luke Piszkin (now a PhD student in the Biophysics Department at the University of Notre Dame) for the first paper in the lab to be first-authored by an undergraduate! Luke’s paper is titled Extremophile enzyme optimization for low temperature and high salinity are fundamentally incompatible and appears in the journal Extremophiles. In the paper Luke explores the molecular basis underlying the intriguing observation that there appear to be very few (no?) extreme halophiles that are also extreme psychrophiles, despite the fact that there are many environments on Earth that are both cold and salty.

One of these environments is Deep Lake, Antarctica, which supports a microbial community dominated by the mesophilic archaeon Halorubrum lacusprofundi (optimal growth temperature of 36 °C). That’s rather surprising given that your typical true psychrophile conks out at about 18 °C. Like all haloarchaea, what H. lacusprofundi can do is tolerate high levels of salt, up to 4.5 M NaCl or 262 g L^-1. That level of salt tolerance is not seen among the documented true psychrophiles. Why not?

In the manuscript we posit that it comes down to the different amino acid substitutions needed to adapt a protein to high salt or low temperature conditions. High salt proteins typically have low isoelectric points, derived from more acidic amino acids. The practical implication of this is that they have a more negatively charged surface that requires a high concentration of salt for stability. This is a requirement for the “salt-in” strategists that dominate the most saline environments (such as salt crystallizer ponds). These microbes are primarily archaea but include a few bacteria, and deal with the high salinity of their environment by accumulating high intracellular concentrations of the salt KCl. This maintains their osmotic balance while excluding more harmful salts, but requires proteins that are compatible with high concentrations of KCl. By contrast most halotolerant bacteria (including psychrophiles that inhabit moderate salinity environments) are “salt-out” strategists that accumulate organic solutes to maintain osmotic balance. These solutes impose no particular requirements on intracellular proteins.

The trick is that amino acid substitutions that lead to a lower isoelectric point also decrease the flexibility of the protein. Increased flexibility is the key protein adaptation to low temperature. Thus the fundamental incompatibility between optimization to low temperature and high salinity. To test this idea Luke dusted off a model, the Protein Evolution Parameter Calculator (PEPC), that I developed many years ago in the waning days of my PhD. After updating the code from Python 2 to Python 3 and making some other improvements, Luke devised an experiment to “evolve” core haloarchaea orthologous group (tucHOG) proteins from H. lacusprofundi and the related mesophile Halorubrum salinarum. By telling the model to select for increased flexibility or decreased isoelectric point he could identify how improvements in one parameter impacted the other. As expected, likely amino acid substitutions (based on position in the protein and the BLOSUM80 substitution matrix) that increased flexibility also strongly favored an increased isoelectric point.

Congrats to Avishek Dutta for his new paper “Detection of sulfate-reducing bacteria as an indicator for successful mitigation of sulfide production” currently available as an early view in Applied and Environmental Microbiology. This was intended to be the second of two papers on a complex experiment that we participated in with BP Biosciences, but the trials and tribulations of peer review led this to be the first. We’re pretty excited about it.

Here’s the quick background. When microbes run out of oxygen the community turns to alternate electron acceptors through anaerobic respiration. One of these is sulfate, which anaerobic respiration reduces to hydrogen sulfide. In addition to smelling bad hydrogen sulfide is pretty reactive and forms sulfuric acid when dissolved in water. For industrial processes this is a problem. Sulfide can destroy products, inhibit desired reactions, and corrode pipes and equipment. To make matters worse, sulfate reducing bacteria (SRBs: those microbes that are capable of using sulfate as an alternate electron acceptor) can form tough biofilms that are hard to dislodge.

One way of dealing with undesired SRBs is to fight biology with biology and add a more preferential electron acceptor. Oxygen would of course work really well, but it typically isn’t feasible to implement oxygen injection on a really large scale. However, nitrate also works well. If nitrate is abundant nitrate reducing bacteria (NRBs) will outcompete SRBs for resources (e.g., labile carbon). Great! Now here’s the challenge… adding massive quantities of nitrate salts is expensive and likely has it’s own ecologically and environmental consequences. So we’d like to do this judiciously, adding just enough nitrate to the system to offset sulfate reduction. But how to know when you’ve added enough? In a really big system (like an oil field) the sulfide production can be happening very far from any possible sampling site so simply measuring the concentration of hydrogen sulfide doesn’t help much. But we can learn some useful things by monitoring the microbial community in the effluent.

The figure above is a schematic of the formation and decay of the biofilm before, during, and after mitigation. In our study the biofilm was presumed to be sulfidogenic and the mitigation strategy was addition of nitrate salts, but the concept applies equally well to any biofilm and any mitigation strategy. The trick – and this is one of those things that seems painfully obvious after the fact but not before – is that you’re looking for the thing you’re mitigating to appear in the effluent. Although this might seem to suggest increased abundance in the system, it actually represents decay of the biofilm and loss from the system. To take this a step further we used paprica to predict genes in the effluent and then identified anomalies in the abundance of genes involved in sulfate reduction. This anomalies provide specific markers of successful mitigation and a means to a general strategy for monitoring the effectiveness of mitigation.

Congratulations to Srishti Dasarathy for her first first-authored publication! Srishti’s paper “Multi-year Seasonal Trends in Sea Ice, Chlorophyll Concentration, and Marine Aerosol Optical Depth in the Bellingshausen Sea” is out in advance of print in JGR Atmospheres. This paper was a really long time in coming. For this study, Srishti made use of several different satellite products including measurements of marine aerosol optical depth (MOAD) derived from the CALIPSO satellite. We are not a remote sensing lab and Srishti doesn’t come from a remote sensing or physics background, so the learning curve was pretty steep. It took a couple of years, a lot of Matlab tutorials, and an internship with the CALIPSO team at NASA’s Langley Research Center just to crack the CALIPSO data and start testing hypotheses. Srishti’s main hypothesis was that MOAD would be positively correlated with ocean color and negatively correlated with sea ice, since phytoplankton are known to be a source of volatile organic compounds that can form aerosol particles. Confounding this is that sea spray – which like phytoplankton is associated with open water periods – is also a source of aerosols.

One challenge that we faced was that CALIPSO represents data with high spatial resolution along a 2D path or “curtain”, as shown above. The orbital geometry is such that not every point on the globe gets covered; the same curtains get sampled every 16 days. Thus, while spatial resolution is high along the curtain, it is poor orthogonal to the curtain, and temporal resolution is limited to 16 days. This makes it a bit challenging to capture signals associated with relatively ephemeral events (such as phytoplankton blooms).

To work around these limitations Srishti took a basin-scale view of the CALIPSO data and looked for large scale trends that would link CMOD with chlorophyll a or ice cover. This approach isn’t idea and glosses over a lot of interesting details, but it is nonetheless sufficient to reveal some interesting relationships. Most notably that MOAD and chlorophyll a are weakly but significantly correlated in a time-lagged fashion, with a delay of approximately 1 month yielded the strongest correlation. This makes sense, as the volatile organics compounds that link phytoplankton (and ice algal) communities to MAOD are thought to be maximally produced near the end of the phytoplankton bloom as the biomass starts to decay. In the near future new satellite missions like PACE and improved land/sea observing campaigns will allows us to get into the details a bit more, including direct observations of specific blooms and the time- and space-lagged MOAD response!

Last week PhD student Natalia Erazo and I were fortunate to get back into the field after a long pandemic hiatus.  Our mission was to collect mangrove propagules (essentially a detachable bud from which the mangrove seedling sprouts) from the Indian River Lagoon in Florida for an upcoming experiment on mangrove-microbe symbiosis.  Neither of us had worked in Florida before so we teamed up with Candy Feller, an emeritus scientist with the Smithsonian Marine Station in Ft. Pierce, FL.  Candy has been working on mangroves in Florida and around the world for decades and is extremely knowledgeable about the ecology of these systems.  She and husband Ray Feller allowed us to tag along as they checked on a few long-term experiments and study sites up and down the coast.

For those not familiar with Florida’s Atlantic coast, the Indian River Lagoon is a network of estuaries and barrier islands that stretch from north of Cape Canaveral to south of Port St. Lucie.  The barrier islands form a protected waterway that provides habitat for mangroves, manatees, and a variety of other species.  The Indian River Lagoon is home to quite a few people as well, and there are some issues associated with water quality. Nutrients from septic and sewage systems are cited as a cause of high phytoplankton loads and increasingly murky water, leading to a reduction in aquatic vegetation and increased manatee mortality.  Key landscape features in the Lagoon are also the result of human habitation.  For example, much of the mangrove habitat in the Ft. Pierce region exists within engineered mosquito abatement areas.  To reduce the number of mosquitos (currently a nuisance, but previously some did carry disease) berms were created around vast tracts of mangrove habitat.  These areas were then flooded, reducing the breeding success of mosquitos because they lay eggs on wet but not flooded soil. 

Unfortunately, mosquito abatement also killed the mangrove trees which, while salt tolerant and adapted to life in saturated soils, require tidal action to oxygenate the water.  Modern mosquito abatement efforts (while still energy and labor intensive) take this into account and mangroves are thriving in areas that were formerly stagnant abatement ponds.  This is a Good Thing for anyone who likes fish, crabs, shoreline stabilization, and any of the other services that mangroves are well known for providing.

A particularly interesting feature of the Indian River Lagoon is that it is oriented north to south at nearly the northernmost known extent of mangroves on the US Atlantic Coast.  This provides an excellent opportunity to study how mangroves are responding to changing climate.  It’s known that mangroves are extending their range to the north, but climate change is anything but linear, and the rise in atmospheric and sea surface temperatures are accompanied by instabilities and severe perturbations.  The most notable may be freezing events caused by deep intrusions of the now infamous polar vortex.  Such perturbations can have a bigger impact on landscape ecology than the background climate.  Mangroves are very much a tropical species but somewhat resistant to transient freeze events (at least more so than your average Florida orange tree).  How they respond physiologically to these and other stressors that they encounter in their northward progression remains to be seen.