Moselio Schaechter


  • The purpose of this blog is to share my appreciation for the width and depth of the microbial activities on this planet. I will emphasize the unusual and the unexpected phenomena for which I have a special fascination... (more)

    For the memoirs of my first 21 years of life, click here.

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April 17, 2014

Terms of Biology – Syntrophy

by Elio

Figure1 Figure 1. An example of syntrophy. Here, benzoate degradation is carried out by a model syntrophic consortium. Dashed blues lines indicate additional sources of cell carbon for the methanogens. Source.

Syntrophy or “eating together” is a kind of symbiosis widespread in biology and of great importance for key biochemical transactions on this planet. Indeed, the cycling of carbon in many of its aspects depends on microbial syntrophy. Thus, the breakdown of waste material critically depends on chemical modifications brought about by microbial syntrophic associations. The phenomenon basically, requires reverse electron transfer, i. e., the input of energy to drive critical redox reactions.

The term syntrophy was first used in microbiology to describe the association between chemotrophic sulfate reducing and phototrophic sulfur oxidizing bacteria. Syntrophy exists between two species of bacteria, two of archaea, or one of each. Lucky for us, a fine essay on this subject was written a few years ago by McInerney, Sieber, and Gunsalus. A later, more detailed review by the same authors is found here, and by others here as well. You will encounter a large variety of fascinating syntrophic associations as well as learning of the ingenious ways that microbes have evolved to squeeze out the last drop of energy from thermodynamically unfavorable reactions. A classic example of symbiotic syntrophy involves an archaeon that can convert methane into CO2 and hydrogen by virtue of a sulfate-reducing bacterium siphoning off the hydrogen byoxidizing into into H2S. And more complex systems abound. For instance, in a tripartite relationship, bacteria in anaerobic environments degrade polymers into acetate, long chain fatty acids, and other compounds, other microbes convert these into hydrogen, formate, and acetate, and a third group use these to make methane and CO2.

The authors pose the question of how do syntrophic cell partners communicate? The answers are not obvious and require further investigation. But a great deal is known already. In the authors’ words: Genomic analysis has revealed the genetic blueprints of model syntrophic substrate-utilizing microorganisms, which in turn has provided insights into the metabolic pathways operative for carbon and electron flow to the end products acetate, hydrogen, and formate. The emerging bio- chemical paradigm includes the need for reverse electron transfer via membrane-bound as well as cytoplasmic confurcation-type enzymes that dispose of electrons as hydrogen and/or formate. Syntrophic strategies truly exist at the thermodynamic limits of life."

March 10, 2014

The Cold Side Of Microbial Life

by Gemma Reguera

The Cold Side Of The Earth

Figure1 Figure 1. The Arctic and boreal regions of the Earth (or Northern lands) are shown in color, each color corresponding to a different type of above-ground vegetation. Source.

In the midst of one of the worst Michigan winters on record, I felt inspired to learn more about how microbes cope with the cold (it is true: misery does indeed like company). So it happens that the polar vortex that has visited our northern states a few times this season has given us a glimpse at the subzero temperatures that prevail in the northern lands of our planet, sometimes all year round. The most northern areas are what we generally know as the Arctic, a treeless region that is either barren of vegetation or limited to tundra (shrubs, grass, lichens, etc.). As you continue going south, you enter the boreal region, characterized by its thick coniferous forests. The soils in these northern lands are called permafrost because they remain frozen for long periods of time. The almost permanent frozen state of these soils limits microbial activity and the turnover of organic matter. Not surprisingly, permafrost soils store approximately 25% of all of the planet’s soil organic matter.

For all their frigid circumstances, do not assume that permafrost microbes are in a perpetual state of hibernation. These frozen soils harbor a broad diversity of microbes. Furthermore, bulk measurements indicate that there is microbial metabolic activity despite the subzero temperature regimes. Just how much, we don’t know. Scientists fear that global warming will continue to thaw large areas of permafrost, promoting microbial activities and the turnover of the large quantities of organic matter trapped in the soils. This could lead to the release of substantial amounts of carbon into the atmosphere in the form of carbon dioxide and the greenhouse gas methane. However, with the limited information we have, it is difficult to predict how the permafrost microbial communities will respond to global warming.

Continue reading "The Cold Side Of Microbial Life" »

February 17, 2014

The Fungus That Killed Darwin’s Frog

by Gemma Reguera

A Mouthful of Kids

Figure1 Figure 1. A ‘pregnant’ male Darwin frog carries its babies in the vocal pouch (left) until they are big enough to be spat out (right). Sources here and here.

In his second expedition to South America, Darwin discovered many new species of animals and plants. The field observations obtained throughout this 5-year expedition provided the intellectual framework for the maturation of his ideas on evolution. It also introduced the world to a tiny (2-3 cm in length) frog known as Darwin’s frog. The group includes the northern (Rhinoderma rufum) and the southern (Rhinoderma darwinii) species, which inhabit the central and southern forests of Chile (and adjacent areas of Argentina), respectively. As in many other amphibians, fecundation is external. However, Darwin’s frogs do not leave the fecundated eggs on the ground and exposed to environmental insults and predators. The males scoop them with their mouths and incubate them in their vocal sac. The dedicated dads feed their offspring after the eggs hatch, producing secretions analogous to milk that allow the tadpoles to grow in a protected environment, sometimes until they have fully developed into froglets. When the young are mature enough to fend for themselves, the male frog literally spits them out. You can see a short video describing this amazing reproductive strategy following this link. This behavior, generally known as neomelia, allows the male ‘surrogates’ to care for the eggs and then the young, maximizing survival throughout the critical tadpole stage. Unfortunately, deforestation in the regions inhabited by these frogs has resulted in vast habitat losses, leaving Darwin’s frogs in precarious conditions. The last sight of a northern Darwin frog was reported in 1980, leading researchers to suspect that this particular species went extinct years ago. The species has been tagged as ‘possibly extinct’. The southern species, R. darwinii, which has traditionally occupied a much larger region, has been able to survive, but population numbers have declined dramatically.

Continue reading "The Fungus That Killed Darwin’s Frog" »

November 18, 2013

Antarctica's Deep Lake: A Frigid Home for Steadfast Archaea

by Elio

Some time ago, we asked this Talmudic Question: Can you think of a place on Earth where there is free water but no microbes? (A sterile flask of nutrient broth in a lab, the insides of the body, or an IV bag in a hospital don't count.) Someone answered that perhaps deep in Antarctica there would be a buried lake that was sterile. Not so. The early returns from the deep drilling of the buried Vostok Lake indicated the presence of microbes even there. But other sites in that continent come close to being just such an environment because their microbial population is unusually thin. So, let’s rephrase the question: “Can you think of natural places where there is free water but, comparatively speaking, few microbes?” (Pick your own limit, but <105 cells/ml seems thin to me.)

Figure1 Fig. 1. Deep Lake. Credit: University of New South Wales. Source.

In answer, I point to Deep Lake, a most unusual environment in East Antarctica. Despite its low temperature, it never freezes. The reason? Its waters contain 27% salt (w/v, almost 5M of NaCl), just a tad below the salinity of the Dead Sea (33.7% w/v) and about ten times that of the oceans. That salinity keeps the lake liquid despite temperatures being as low as -40° C for the ambient air and -20° C throughout its depths. What stalwart microbes do you find here? Mainly, some unusual salt-loving Archaea, aka haloarchaea. They have been studied in some detail, with some unexpected findings. Four species make up about three quarters of the total population. They belong to a different genus each, all within the family Halobacteriaceae (the ‘-bacteriaceae’ being a holdover from older, “pre-archaeal” taxonomy). Like other haloarchaea, some of these isolates harbor two or three chromosomes, or replicons, totaling nine replicons among them. Each species has its own predilection for particular depths. They can be cultivated, as long as you have a lot of patience. One of them grows at temperatures from -1° C to 40° C, fastest at 33° C. When cultured in the laboratory at -1°C, the temperature of the top of the lake, the cells double six times per year. At that rate, the authors calculate that it would take 10 years for a single cell to fully populate the lake.

Continue reading "Antarctica's Deep Lake: A Frigid Home for Steadfast Archaea" »

November 08, 2013

A Whiff of Gastronomy

by Elio

Figure1 Fig. 1. The black truffle Tuber melanosporum. Source.

Ah, truffles! They are the gourmet’s celebration, the cook’s inspiration, the common man’s anticipation. And they demand quite a price, which at last count hovered around $2,000 a pound for Italian white truffles, the French black truffles being cheaper, but still not within the reach of most people. And their production is decreasing. So it befalls few mortals to dine on these delicacies. But there is hope. As many of you know, truffle oil is used quite widely in restaurants. So, where does truffle oil come from? Did it ever see a truffle? Before going there, let’s agree that the price is right. You can find all sorts of truffle oil on the Internet for as little as $5 per ounce. It even comes in a kosher variety.

 

Figure2
Fig. 2. 2,4 Dithiapentane.

Alas, truffle oil has not been within smelling distance of a real truffle but is simply some sort of regular vegetable oil, often olive oil, that has been doctored by the addition of the ether, 2,4 dithiapentane. This is one of the main odor-producing compounds in truffles and, to a debatable approximation, it emulates the real aroma. Many expert food connoisseurs disagree and maintain that it doesn’t resemble the real thing. Also, some are bothered by the artificial nature of this concoction. In defense of truffle oil, the flavor it imparts to food is quite impressive. It does remind me of the taste of the few truffles I have been lucky enough to eat—metallic, pungent, earthy, and very distinctive. It may not be the real thing but it does contribute a nice bite to otherwise uninspired dishes. I, for one, am on the side of the folks who enjoy it. The only allowance needed is to agree that even an imitation may taste good.

Continue reading "A Whiff of Gastronomy" »

October 28, 2013

Programmed Cell Death

by S. Marvin Friedman

Gone are the days when bacteria were thought to just grow and divide and not bother to converse with one another. That simple idea has produced mountains of data and most of what we know about bacterial physiology is based on this notion. It turns out, as we know now, that this is an oversimplification. Lots more goes on in the life of bacteria, much of it dealing with communication with other cells. One widespread, if menacing-sounding, mode of communication consists of killing individuals of a population, apparently for the benefit of other of its members. A prevalent mechanism that accomplishes this is the toxin-antitoxin system for programmed cell death. Here, some seemingly altruistic cells are killed from within whereas others get dispatched after receiving extracellular signals. A lot is known about both mechanisms, this being a very active field of research. The signals involved are the subject of this piece.

Figure1
Figure 1. Toxin–antitoxin (TA) module. Antitoxins bind their cognate toxins and inhibit toxin activity. In E. coli, the toxin is activated by degradation of antitoxin either by the Lon or the ClpAP protease. Activated toxins inhibit cell growth and/or lead to cell death. Source.

The Loaded Gun

Many, possibly most bacterial genomes carry toxin-antitoxin modules consisting of two genes in an operon, one that codes for a stable toxin, the other for a labile antitoxin. When the antitoxin vanishes for whatever reason, the toxin can go to work and kill the cell that made it or, through extracellular signals, induce the killing of adjacent cells. Most bacteria contain multiple versions of these modules. Bioinformatic analysis of the tubercle bacillus genome has come up with an astounding 88 putative toxin-antitoxin candidates!

A widely studied system (and the earliest discovered in a bacterial chromosome) is mazEF in Escherichia coli. Stressful conditions that turn off the expression of the mazEF module result in reduction of antitoxin MazE within the cell, allowing toxin MazF to act freely. Such conditions include inhibition of transcription and/or translation, severe amino acid starvation leading to overproduction of the alarmone ppGpp and DNA damage. Engelberg-Kulka’s laboratory, where this phenomenon was first discovered in the bacterial chromosome (it had been found earlier in plasmids by Yarmolinsky) had shown that mazEF-mediated programmed cell death is triggered in other cells of the population by an extracellular quorum sensing factor, EcEDF (for E. coli Extra-Cellular Death Factor). It was identified as the fairly simple pentapeptide NNWNN and acts as a quorum-sensing molecule. In the present report, Kumar and coworkers now extended this story to other species, investigating the EDFs of Bacillus subtilis and Pseudomonas aeruginosa.

Cross Talk

Prior work had shown that in E. coli mazEF-mediated cell death takes place in dense cultures and that the supernatant from such a culture induced cell death when added to dilute cultures of E. coli. Now, supernatants from B. subtilis and P. aeruginosa were added separately to dilute E. coli cultures that had been treated with rifampin to induce mazEF. Both the B subtilis and P. aeruginosa supernatants killed these E. coli cells. Thus, both B. subtilis and P. aeruginosa supernatants contain EDFs. To purify them, the authors ran supernatants through high performance liquid chromatography (HPLC). The only compound (BsEDF) isolated from B. subtilis turned out to be a hexapeptide with the amino acid sequence RGQQNE. Three active compounds were isolated from P. aeruginosa, the nonapeptide INFQTVVTK (PsEDF-1) and the hexadecapeptides VEVSDDGSGGNTSLSQ (PsEDF-2) and PKLSDGAAAGYVTKA (PsEDF-3). Synthetic identical peptides also displayed EDF activity.

Figure2
Figure 2. The effect of the supernatants from dense cultures of B. subtilis or P.aeruginosa on mazEF-mediated cell death in E. coli. Wild-type and ΔmazEF E. coli cultures were not diluted (Dense) or diluted to a density of 3 x 104 cells/ml in M9 medium (M9) or in a supernatant of a dense culture (SN) of B. subtilis or P. aeruginosa. The samples were incubated without shaking at 37°C for 10 min and for another 10 min with rifampin (10 µg/ml). Source.

Programmed cell death in B. subtilis is mediated by the ydcDE operon that codes for a toxin-antitoxin module and belongs to the mazEF family. The authors found considerable cross-talk, as each of the EDFs from E. coli, B. subtilis or P .aeruginosa added separately to dilute cultures of B. subtilis resulted in ydcDE-mediated cell death. Next, they determined which amino acids of BsEDF are essential to cause cell death. The first (arginine) was not required to kill E. coli, but the second one (glycine) was essential. To kill B subtilis, however, glycine, the third (glutamine), fourth (glutamine) and sixth (glutamic acid) amino acids were absolutely required, and the fifth (asparagine) partially required.

How Does the Toxin Work?

It has been known that MazF works as an endoribonuclease by cleaving RNA and that EcEDF enhances this activity. Would BsEDP and each of the PsEDFs also enhance MazF endonucleolytic activity in E. coli? The authors assayed this activity using a continuous flurorometric assay with substrates consisting of one or five RNA bases flanked by DNA nucleotides labeled with a fluorophore at their 5’ ends and a quencher molecule at their 3’ ends. Processing of such a substrate leads to a great increase in fluorescence. Adding BsEDF or any of the PsEDFs led to a concentration-dependent enhancement of MazF activity. The PsEDFs were, however, less efficient than EcEDF or BsEDF. Which amino acids in BsEDP are critical to stimulate MazF activity in vitro? These workers found that the second (glycine) and fifth (asparagine) amino acids are absolutely required, whereas the fourth (glutamine) and sixth (glutamic acid) are partially required and the first (arginine) and third (glutamine) acids are not necessary. Thus, for both the in vivo effect of BsEDF on cell death in E. coli and the in vitro effects of BsEDF on MazF activity, the second and fifth amino acids (though different) are absolutely required and the first amino acid is dispensable.

Figure3
Figure 3. BsEDF and the PaEDFs enhance the in vitro MazF endoribonucleolytic activity. (A) How the endoribonucleolytic activity of MazF was measured. Cleavage of the chimeric fluorescent oligonucleotides by MazF increases fluorescence emission of the fluorophore FAM. (B) The time course of the effect of BsEDF (RGQQNE) on MazF activity (Left panel). The relative (%) increase of MazF activity induced by BsEDF (Right panel). (C) Ditto for the effect of PaEDF-1 on MazF activity. Source.

Using affinity columns, the authors showed that BsEDF binds directly to MazF but not to MazE. The BsEDF mutant with alanine substituted for glycine in the second position did not bind to MazE. Previously, work from this laboratory showed that EcEDF can overcome the antagonistic effect of the antitoxin MazE on the E. coli MazF toxin, thus freeing the toxin from its inhibitor and enhancing its activity. The way this works, it is thought, is by EDF displacing MazE from the MazE-MazF complex. Would BsEDF and PsEDFs also do this? BsEDF did overcome the inhibitory effect of MazE on MazF, but none of the PsEDFs displayed this activity. However, EcEDF was more efficient than BsEDF in overcoming the inhibitory effect of MazE. The same amino acids of BsEDF were required to overcome the inhibitory effect of MazE and for stimulating the E. coli MazF activity in vitro.

A variant form of EcEDF with the sequence NNGNN (named EciEDF) interferes with the ability of WT EcEDF to stimulate the activity of E. coli MazF. Likewise, the variant BsEDF, RAQQNE (named BsiEDF) interferes with the enhancement of E. coli MazF activity by WT BsEDF. Importantly, they found that iEDFs from each species prevented the activity of the EDF of its own species but not that of other species. This result strongly suggests that EDFs from E. coli, B. subtilis and P. aeruginosa interact at different sites on E. coli MazF.

All In The Family?

In E. coli, the quorum-sensing peptide EcEDF triggers a MazF-induced downstream pathway that leads to death of most of the population, leaving a small subpopulation to survive. It is interesting to note that in contrast to the situation in E. coli and B. subtilis, cell death mediated by P. aeruginosa is population-independent, suggesting that its EDFs are used to kill other bacteria but not itself. The present study of EDFs from B. subtilis and P. aeruginosa indicates that programmed cell death is a much broader phenomenon that also involves interspecies killing and that could play a major role in the regulation of mixed bacterial populations. As the authors say: ” ….the production and the role of the QS EDF peptides in interspecies bacterial cell death predicts the existence of an ‘EDF family’ that would also be found in other bacterial species.” An exciting ramification of this research is the possibility of using EDFs as a novel class of antibacterial agents. Could this spell welcomed troubles for those hard-to-treat multiple antibiotic-resistant strains?

Friedman, Marvin_sm

S. Marvin Friedman is Professor Emeritus, Department of Biological Sciences, Hunter College of CUNY, New York City and an Associate Blogger for Small Things Considered.

ResearchBlogging.org


Kumar S, Kolodkin-Gal I, & Engelberg-Kulka H (2013). Novel quorum-sensing peptides mediating interspecies bacterial cell death. mBio, 4 (3) PMID: 23736285

September 30, 2013

E. coli Keeps Its Powder Dry

by S. Marvin Friedman

Figure1
Figure 1. Left: Dictyostelium discoideum aggregating into a fruiting body. Source. Right: Adult Caenorhabditis elegans male. Source.

Whether inhabiting soil, fresh water, or marine ecosystems, bacteria are constantly facing the threat of numerous and effective predators such as protists, nematodes, or phages. To defend against such predation, bacteria have evolved a number of strategies, including getting larger in size, moving faster, producing defensive secondary metabolites, and forming biofilms. Because of its anthropocentric appeal, one strategy stands out—keeping a locked armory of genetic weapons. The strategy involves breaking open the armory when confronted by perils—activating the genes that were locked mostly to save the cost of expressing them when not needed.

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September 23, 2013

The Microscopic Flash Mob

by Melissa Wilks

Every day we see animals migrating through the air, across plains, and in the oceans, in beautifully coordinated patterns; starlings flock together in the thousands while sardines swim together in enormous shoals. These social behaviors are important in allowing animals to socialize, avoid predators, and find refuge and food. But what about smaller organisms? Although more difficult to visualize, microorganisms can perform these coordinated behaviors as well. As is true for birds and fish, figuring out how and why bacterial cells communicate to organize their movements will help understand the behavior of bacteria in the environment.

Figure0
Figure 0. Ripples on the Camas Prairie of northern Montana. Source.

Myxococcus xanthus is a soil dwelling microbe that has attracted the attention of microbiologists in part due to its predatory activity. To survive in environments where nutrients are scarce, it kills and breaks open other bacteria and feeds on the released nutrients. During times of starvation, M. xanthus cells self-organizes into fruiting bodies, macroscopic dome-shaped mounds that contain over 100,000 cells each. In these structures, the cells differentiate into metabolically inactive spores that remain dormant and allow survival until times get better. Since the 1960s, M. xanthus cells have also been known for organizing their movements across a solid surface to produce waves; this phenomenon, called rippling, resembles the waves produced when you throw a rock into a pond, and can be seen in nature on a scale ranging from the microscopic to the geologic.

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August 26, 2013

Mycorrhizal Fungi: The World’s Biggest Drinking Straws And Largest Unseen Communication System

by Elio

Figure1
Figure 1. Mycorrhizal fungal filaments around a plant root. Source.

Quick, which is the biggest symbiotic association on Earth? Did you guess the mycorrhizae? They are the huge symbioses between fungi and the roots of most terrestrial plants. Their total size is not easy to measure because not all the fungal filaments in soils are mycorrhizal nor are the mycorrhizal ones always mycorrhizing (a new verb?). Even so, one can make the case that the combined mass of the participating fungi and plant roots is colossal. A friend, the soil microbiologist David Lipson, tells me that the mass of mycorrhizal fungi on the planet is estimated to be somewhere around 1-3 x 1010 tons (that’s between 1.4 and 4 tons per person). We are obviously talking about a topic that deserves much more of our attention than it has received to date.

Plants depend on mycorrhizal fungal filaments to supply them with a stunning proportion of their needed water and minerals. In some forests, these fungi provide the plants with up to 80% of their nitrogen and 90% of their phosphorus. The fungi, in turn, depend on plants to provide them with organic compounds needed for their own growth. Although it had been stated that this traffic in carbon is unidirectional—plants-to-fungi—some 400 species of non-photosynthetic plants (e.g., Indian pipes) that lack chlorophyll are proof to the contrary. For these mycoheterotrophs, mycorrhizae are the primary suppliers of essential carbon compounds.

Continue reading "Mycorrhizal Fungi: The World’s Biggest Drinking Straws And Largest Unseen Communication System" »

August 19, 2013

Oddly Microbial: Programmed Cell Death

by Marcia Stone

The lab was a crime scene; dead and dying cells were everywhere—but did they commit mass suicide or were they murdered? The burly young post doc who discovered the corpses sat in the corner sobbing softly, his PI by his side. She was trying in vain to comfort her most promising young scientist in decades, hoping this wouldn’t drive him into medical school or even worse, into engineering. At a loss, Sarah Sweltweather made an emergency call to cell-death expert, Detective Chief Inspector Douglas Green at St Jude Children’s Research Hospital in Memphis. Their cells were immortalized, she told him, they weren’t supposed to die in such huge numbers; thus her post doc was unprepared for the morning’s grisly discovery. “Please,” she implored Green, “help us make some sense out of what’s happened.”

Cool as ever, Green reminded the distraught PI that dying is something cells naturally do a lot of and most cell biologists eventually get used to it. Finding out how cells die and why is our job, he added noting that cell death is rarely passive, citing “suicide, murder or sabotage” as the causes.

Suicides, continued Green, are most often carried out by apoptosis and murders committed with reactive oxygen species (ROS) or other toxic molecules released by “killer” cells. “Sabotage is a bit more subtle: here derailing some normal cellular processes kills the cell. The best known example of cell sabotage is mitotic inhibition or ‘mitotic catastrophe’ where DNA damage keeps the cell from dividing and this leads to its death.

It would be best if “I examined the crime scene myself,” cautioned Green. “Don’t touch or move anything, I’ll catch the next train out.”

Continue reading "Oddly Microbial: Programmed Cell Death" »

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