The Two Faces of Photorhabdus

by Merry

Heterorhabditis bacteriophora. Source.

Nematodes—those small, nondescript roundworms—are among the most numerous animals, found in virtually every ecological niche. With an estimated 400,000 to possibly 10,000,000 species, their diversity might match that of the insects, but no one knows for sure. Most nematodes live quiet lives, unknown even by taxonomists, but one—Caenorhabditis elegans—has made it big time as a model organism. Two families of entomopathogenic nematodes have garnered much attention due to the company they keep. (More about this below.) Some 350 million years ago, each family, it is thought, independently began a mutualistic association with a lineage of enteric bacteria—one with the lineage leading to the genus Photorhabdus, the other with the Xenorhabdus lineage. These nematode-bacterial teams are deadly parasitoids that prey on diverse insects. Both teams have evolved obligate mutualisms with similar life cycles, but the underlying molecular mechanisms are distinctly different. We'll focus on Photorhabdus luminescens, the most "illuminating" example. For a review article covering Xenorhabdus, click here.

The life cycle of Photorhabdus. Source.

Although these are soil-dwelling nematodes, only one larval stage, called the infective juvenile (IJ), is found free in the soil. The name is appropriate, as the IJ is arrested at this larval stage (it is in diapause) and its sole task is to find and enter an insect host. The rest of the life cycle, including reproduction, takes place within a host.

Each IJ carries quiescent Photorhabdus in its gut. Once inside a new host, the nematode IJ migrates to the hemolymph and regurgitates the Photorhabdus. The bacteria quickly launch into a phase pathogenic to the insect, secreting toxins, enzymes, and antibiotics. Toxins effectively disable the insect's innate immune defenses; enzymes convert host tissues into a nutrient soup; antibiotics preserve the cadaver and, being effective against Xenorhabdus, limit competition from Xenorhabdus-associated nematodes. Experimental injection of less than 5 bacteria into an insect's hemolymph kills within 48-72 hours. In contrast, the insect's innate immune response can handle similar infection by >10⁶ E. coli.

Photorhabdus luminescens strain W14 on an insect
midgut under the collagen sheath. Source.

The Photorhabdus grow exponentially for the first 48 hours, reaching populations of 10⁹ per host. By then, all of the host tissue has been converted into bacterial biomass—essentially a monoculture of Photorhabdus. Meanwhile, the IJ exits from diapause, begins to feed, and continues development into an adult hermaphrodite. Photorhabdus then stops growing, and its pathogenic phase gives way to the mutualistic phase. The accumulated bacterial biomass literally nourishes the nematodes, providing most, if not all, of their nutritional requirements. The picky nematodes will feed on particular strains only.

The nematodes reproduce for several generations inside the cadaver. Then, in response to environmental signals, a final generation of eggs develops into IJs within the body of their mother nematode. They receive a vital bacterial inoculum by maternal transmission, not haphazardly from the population free in the cadaver. Thus armed, the IJs emerge from the cadaver and go forth in search of a new host, to repeat the cycle yet once again. Under optimal conditions in the lab, the entire cycle takes 10-20 days and 100,000 IJs can be produced from infection of one insect by a single IJ. Definitely a well-run operation.

Photorhabdus and Xenorhabdus have aroused considerable interest as sources of new antibiotics and broad-spectrum insect toxins for agricultural use. They are also potential model systems. Consider the possibilities. First you could compare the mechanisms used for pathogenesis and mutualism, and investigate what it takes to quickly switch from one to the other. Then there are the two similar, but distinctly different, strategies evolved by Xenorhabdus and Photorhabdus. What might they tell us about the mechanisms used by the numerous closely-related and well-studied human enteric pathogens? (Click here for a recent review discussing these possibilities.)

Around the time of insect death, the insect
corpse appears bioluminescent due to the P.
luminescens within. Source.

Photorhabdus, as the name suggests, carries out bioluminescence. It also infects wounds. This is the stuff of legends, with stories going back to the American Civil War. It seemed that soldiers with wounds that glowed in the dark were more likely to recover. Their improved prognosis has been attributed to the antibiotics produced by Photorhabdus, but this has not been experimentally confirmed. Photorhabdus is the only terrestrial bacteria known to exhibit bioluminescence. They luminesce only during their mutualistic phase, inside the insect cadaver. What use is this? There is evidence suggesting that this may be a non-functional trait, recently acquired by horizontal gene transfer from Vibrio, and in the process of being lost. And besides, Xenorhabdus gets along just fine in the dark. Makes you wonder if Photorhabdus luminescens really needs to be luminescens.

Comments

[Britt]

Bioluminescence is a fascinating and vexing topic. We know of several different and unrelated groups of fungi that do it, but why? Of course, attracting insects for spore dessemination is always suggested but it's not likely the reason, at least not for all situations. Plus, with the incredibly acute sensories of insects (tactile, olfactory, chemo-, etc), it seems unlikely that that they "need" to rely on vision. I know that sounds teleological, but no more so that having to find a reason for glowing in the dark. Maybe it is a shunt system...or maybe it just is. Who knows?

In fungi, some produce sporophores (mushrooms) that glow, but in others it's the mycelium that glows...while buried somewhere. Some argue that it attracts wood ingesting insects...I say that if you have to get that close to see the stuff, you likely already knew it was there via some other method.

After about a 10 second search, I couldn't find any previous writings on the topic by Elio...can someone hook me up with a link?

Also, the journal I edit, Fungi magazine, will publish an article on the topic in the near future. If any reader out there has info on the topic to share, I would GREATLY appreciate it, as I'm always trying to learn more. Thanks!

[Paul Orwin]

I'm really glad that you've brought this up, since nematode-bacteria interactions have become an interest of mine recently (and no, I'm pretty much completely ignorant about them, but I've never let that stop me). The literature on C. elegans as a model of infection is very interesting, although still very new. I've been thinking about how these interactions actually occur in the environment, and how that relates to the emergence of virulence (I think of legionella and mycobacteria in amoeba, and how that relates to their growth in macrophages). The added layer of complexity with insect-worm-bacteria in this complex dance makes my head spin!
Luminescence is interesting for many reasons too, of course, and I'm glad Mark reminded me that it was right here that I read about the possible role(s) of that phenotype besides the marine symbiont scenario. I'm not sure why there has to be a single reason for having a gene, though...I don't see why it can't be different things to different bacteria (a metabolic shunt, a tool for helping out fish/squid, a way to improve transmission, or maybe just something they picked up). I'd say that if we think about it, though, there are ways to approach the problem, especially now that strain genotyping can be done very rapidly and precisely.
For example, we know that there is a non-luminscent symbiont, but are there non-luminescent variants of Photorhabdus? what is their geographic range? I find it even odder that you can't find them outside the worm or the insect? Am I reading too much into the last bit?

[Mark O. Martin]

I have heard that hypothesis, about horizontal transmission via predation.

Many years ago, I worked a bit in bacterial bioluminescence with Vibrio harveyi. This wonderful little critter did not have a specific symbiosis, as seen with Vibrio fischeri with squid or the flashlight fish. V. harveyi, in fact, is commonly found in the gut of marine creatures (as well as planktonically). What is the function of bacterial bioluminescence in bacteria that colonize the gut of fish? Pretty dark in there, right? There was evidence that (sorry about this) fecal pellets produced by the fish were bioluminescent. And fish tend to like to eat "glowing items." Hence the horizontal transfer of these gut inhabitants from fish to fish. Or so we all thought.

I would not be at all surprised that glowing insect carcasses attract predators. Yet the level of luminescence caused by Photorhabdus is quite low!

This brings us all back to the true function of bioluminescence, as Elio has explored earlier in this blog. Is it metabolic? Display only? It's a fascinating if vexing topic.

It is interesting to note that Photorhabdus is the only terrestrial bioluminescent bacterium described. My guess is that, as with bioluminescent marine bacteria, it would be possible to find isolates producing VERY little light.

Merry, that was a lovely essay. I love this topic!

And see? I avoided the obvious puns throughout!

[Bob Blumenthal]

Any chance the luminescent insect corpses attract dusk-feeding or nocturnal animals, improving geographic spread of the ready-to-emerge IJs? Maybe there's some indirect selection here...

Comment on Comment
Hi Bob,
My friend Merry is better qualified to comment but your idea seems very reasonable. Look for the light.
Elio

Comment on Comment on Comment...
Interesting possibility, Bob. The information that I have on this point comes from a 2008 paper by Peat & Adams, available here:

http://mmbio.byu.edu/faculty/bja43/papers/products/adams/Peat2008Symbiosis.pdf

They write: "Photorhabdus, the bacterial symbiont of the nematode Heterorhabditis is typically confined to the gut of its host and the hemocoel of larval insects, with both hosts inhabiting soil environments. As the phase of Photorhabdus that glows is typically only found in insect cadavers, there is no intuitive benefit of glowing to attract prey or distract a predator, as all necessary resources for survival and reproduction are present in the insect cadaver."

Thus, they do not consider the factor that you suggested, i.e., improving geographic spread. However, assuming the glowing cadaver is buried within the soil, even that usefulness seems less likely. But who knows...

Merry