Chipping away at the tip of the iceberg

Through out 2011, we have been featuring more weird and wonderful parasites as we have done in 2010, but on top of that we have also been featuring some of the latest pieces of research being published on all manners of parasitic and infectious organisms which may not have been covered elsewhere in the blogosphere.

Just this year we saw a molecular study that revealed the transmission pathway of a great white shark tapeworm via dolphin blubber, a koala blood parasite named after Steve Irwin, a nematode which infiltrate pine trees via borrowed genes from fungi, a parasitic plant which disperse its seeds via beetles, and a little digenean fluke which changed the face of an entire mudflat, just to name a handful out of dozens that we have featured.

So in 2011, we have gone beyond merely showing you the parasites, but also told you about the research which are being conducted behind the scene to work out how they live. At the same time, you also got a bit of insight into the scientific process, and how knowledge accumulate and grow over time.

In 2012, we will continue to bring you new and exciting research on parasites and parasitism - publications which we found interesting but might not have received their share of fanfare and press releases (as I type this up, I have at least another 4 paper waiting in line to write up. And no doubt I'll find another 4 to write about by the time I'm done with those).

Of course, research on a particular host-parasite system does not simply enter suspended animation after a single study has been published. For many lab groups, the parasite we have featured on this blog form the basis of their scientific research, and the newly published paper we have chosen to feature here merely represents a thin cross-section of their ongoing research program. So in 2012 we might also be revisiting some of the parasites that we have previously featured on the blog, and fill you in on the latest updates as they hit the press.

Of course, you can now also find us on other forms of social media where we will be posting about updates to the blog. Susan is on Twitter , and you can find myself on Google Plus.

Here's to another year of parasitism - the most common way of life on this planet!

Hematodinium sp.

Today's parasite, Hematodinium sp., infects blue crabs and causes a disease known as "bitter crab". While the name may sound just slightly nauseating for your palate, for the afflicted crabs, its symptom is down right horrific. The parasite causes the crab's hepatopancreas (equivalent of our liver and pancreas) to malfunction, it starts suffocating, and its muscles eventually dissolve within its exoskeleton. Crabs that are experimentally infected start dying about 2 weeks after initial exposure, and this deadly parasite may have even contributed to the recent decline of blue crabs in Chesapeake Bay.

Hematodinium and related species are dinoflagellates, and while most dinoflagellate are free-living, this species belongs to a group which have evolved to be parasites, with many different species infecting a wide variety of hosts. While several different stages of the parasite have been isolated from the blood of infected crabs, little is known about how they are transmitted between hosts, nor the inner life of those different stages in the hosts. Because many parasites live enclosed within the body of their hosts, it is almost impossible to directly observe how they live and grow the way you might be able to observe a fish or a bird. Ideally, if you can isolate a parasite out of its host, put in it a clear container which closely mimics the conditions found within its host, and still have it complete its life-cycle, then you can find out a lot more about how it lives.

Recently, a group of researchers from Virginia were able to successfully complete the life-cycle of Hematodinium - in vitro - which means they were able to grow it in a culture of chemical broth that sustained the parasite's every need, without any host animals involved. This was accomplished through a painstaking series of transfers, starting with isolating the parasite from infected crabs, then moving each stage into different culture mixes as it grew, all while keeping the conditions as sterile as possible. Out of the 10 isolates they attempted to grow, only 4 successfully completed their life-cycle in vitro. The researchers also found out that the parasite grows best in the dark, and indeed light exposure kills them within weeks, which makes sense given that it is pretty dark inside a crab (a variation on the Marx Brother joke).

Through this in vitro technique, they were able observe the different parasitic stages of Hematodinium directly, and view them as they would have been while floating in the blood and organs of a blue crab. They noted that when Hematodinium cells first enter the crab as "dinospores," they turn into a worm-shaped form called a "filamentous trophont" (see the accompanying photo which was from a figure in the paper). About a month after that, the cells begin transforming into clumps that are composed of multiple clones of the original infection stage. These clumps then grow into a stage called an "arachnoid trophont," which resembles a blob with numerous tendrils around its fringe (which would be embedded in the hepatopancreas of the crab). These clumps tend to merge and form larger blobs as they come into contact with each other. When those "arachnoid trophonts" fully develop, the cells in the middle of the blob start producing spores that eventually turn into the infective dinospores that escape from the crab to infect new hosts, starting the life-cycle anew.

Reference:
Li, C., Miller, T.L., Small, H.J. and Shields, J.D. (2011) In vitro culture and developmental cycle of the parasitic dinoflagellate Hematodinium sp. from the blue crab Callinectes sapidus. Parasitology 138:1924-1934.

Postscript: Three days after this post went up, I was contacted by Peter Coffey, who used to work on this species of parasite with a bit of additional information/correction: I just have one quick comment on the first sentence in your post. In blue crabs we don't see the same bitter flavor that we do in Alaskan Tanner and Snow Crabs, so we haven't been calling infections in blue crabs BCD.
Thanks Peter!

Lepeophtheirus acutus

Today, we are featuring a paper which reported on a grey reef shark (Carcharhinus amblyrhynchos) at Burger's Zoo in the Netherlands that had to be euthanized. "Wait a sec!" you think, "Isn't this supposed to be a blog about parasites? I didn't come here for dead sharks!" Well, just calm down before you close your browser tab in outrage. This particular shark actually succumbed due to a heavy infection of today's parasite - Lepeophtheirus acutus. This parasite is in the same genus as other fish lice that we have previously featured on this blog, but very little is known about this particular species. Prior to this incident, it has only been reported once from the wild, and it was found on the back of a ribbon-tailed stingray (Taeniura lymma), not a shark and certainly nothing was known about how harmful it can be to its host.

From what the staff at the aquarium could work out, this deadly little crustacean was introduced to the facility by an infected male zebra shark (Stegostoma fasciatum) collected off Cairns, Australia on the Great Barrier Reef, which appeared perfectly healthy at the time and passed quarantine. However, about 2 weeks after he was introduced into the aquaria with the other fishes, he started acting weird. At the same time, the grey reef shark mentioned at the start of this post became lethargic and ceased to eat regularly, and about a month after that, both sharks were afflicted with swollen and opaque eyes. Despite the best efforts of the staff to put the infected sharks in quarantine, filter the water with activated carbon, and give them anti-parasite drugs, they were unable to save the grey reef shark, by which time it was swimming with its mouth wide open, not eating at all, and its eyes had deteriorated even further, so the decision was made to euthanize the long-suffering shark.

A necropsy revealed the identity of the killer - a parasitic copepod - most of which were found around the shark's eyes which caused them to become swollen and covered in mucus, and the mouth which led to bleeding gums. The parasite was also found on a female zebra shark and a shovelnose ray (Glaucostegus typus) which shared the aquaria with the deceased grey reef shark. Notably, the blacktip reef shark (Carcharhinus melanopterus) and blacktip sharks (Carcharhinus limbatus) which swam in the same water alongside those infected sharks did not become infected, nor did the many different species bony fishes sharing the same tanks and water. This indicates that L. acutus does display some selectivity in the type of host it infects, with a particular preference for elasmobranchs (sharks and rays), and even then only certain species within that group.

Other than the dead grey reef shark, the other infected sharks survived and recovered fully after treatment. However, this incident shows how outbreaks of infectious diseases can be a big problem for animals in the confined conditions of captivity. In the case of L. acutus, its small size, semi-transparent body, its tendency to infect parts of the host that are difficult to inspect (for example, inside the mouth), and the fact that nothing is know about its ecology meant that the staff had not anticipated such an outbreak. It was the first documented case of infection by a parasitic copepod that led to a shark dying in captivity. This case also illustrates the importance of thorough quarantine procedures, especially when introducing new animals into any facility, as captive conditions can seriously alter the transmission dynamics and pathology of relatively harmless parasites.

Image from figure in the paper.

Reference:

Kik, M.J.L., Janse, M., Benz, G.W. (2011) The sea louse Lepeophtheirus acutus (Caligidae, Siphonostomatoida, Copepoda) as a pathogen of aquarium-held elasmobranchs. Journal of Fish Diseases 34: 793-799,

Rhinanthus minor

Many parasites can have substantial effects on their hosts, but their impact can often extend to other organisms in the environment. Today's parasite is one of the more pretty-looking ones which we have featured in a while - as opposed to the usual worms and lice, today we are featuring a flowering plant - the yellow rattle. But don't let its pretty yellow flowers fool you, Rhinanthus minor is a ruthless parasite.

It is a hemiparasite (like the mistletoe , which becomes rather popular during this time of the year). The plant overwinters as seed in the soil and germinates during spring, penetrating into the roots of its host plants where it can suck out nutrients and water from the plant's xylem tissue. The yellow rattle is a fast growing plant - it flowers 12 weeks after germination and 3 weeks after that it produces seeds that are loosely held in dry capsules, which gives the plant its name. The yellow rattle often share its host plants with a range of insects, so a group of researchers in the UK decided to look at how this hemiparasite can affect those insects. Specifically, they looked at the effects of R. minor on insects that exploit plants in different ways; the aphid that feeds on the sugary sap flowing in the plant's phloem, the spittle bug, which taps into the xylem that transports water and other nutrients, and the grasshopper, which simply chews on leaves.

The researchers predicted that over the course of its growth, the yellow rattle would affect those insects differently. They were expecting that it would negatively impact on the spittle bug, because that insect and the hemiparasite both draw their nutrients from the host's xylem. But as is often the case in science, they found something unexpected.

First of all, they found that the effect R. minor had on those insects depended upon the parasite's growth stage, and it becomes most pronounced when the yellow rattle reaches its peak biomass and begins setting seeds. However in contrast to what they were expecting, spittle bugs actually preferred plants parasitised by R. minor. But the insect that benefited the most from the hemiparasite's presence were the aphids. Not only did they prefer sharing a host plant with the yellow rattle (there were three times as many aphids on plants with R. minor compared to uninfected plants), they also tend to breed more on infected plants. What about the grasshoppers? Grasshoppers were not all that affected by the presence of the yellow rattle either way.

The mechanism behind why the yellow rattle makes its host more attractive to plant-feeding insects is currently unknown. However, it may have something to do with the hemiparasite altering the water content of the host plant, or changing the composition of the phloem sap, which makes it more nutritious to aphids. Either way, it seems that at least for some insects, sharing a plant with a hemiparasite might actually be a good thing.

Image from the Wikipedia.

Reference:

Ewald, N.C., John, E.A. and Hartley, S. (2011) Responses of insect herbivores to sharing a host plant with a hemiparasite: impacts on preference and performance differe with feeding guild. Ecological Entomology 36: 596-604.

Ophiocordyceps unilateralis

Have you ever been so intoxicated that you start walking erratically, stumble away from your friends, stagger around in circles, clamber onto things that you wouldn't normally be seen near, and the next thing you know, you are strapped down in unfamiliar surroundings, unable to extricate yourself? Well, that pretty much describe what happens to ants which become infected with the famous "zombie ant" fungus - Ophiocordyceps unilateralis.

Much has been written about this famous fungus which turns ants into zombies - it is a parasite which captures the same part of our psyche as the monstrosities of horror movies, and there is evidence to suggest that these fungi have been tormenting ants for at least tens of millions of years. But despite all that attention, few people have actually witnessed or documented the sequence of behaviour leading up to the infected ant's paralysis and death. But in a paper published this year, a group of researchers followed the behaviour of ants infected with the famous "zombie"-inducing fungus and compare them to their uninfected brethren.

They noticed a few peculiarities with the behavioural repertoire of infected ants which stood out. While healthy ants studiously stick to the usual lanes of ant traffic, climbing into the canopy to forage with all the other busy worker ants, "zombie ants" are loners which meander around in the understory by themselves, are unresponsive to most stimuli, and frequently stumble and fall from the branches they are walking on. Essentially, the ants act absolutely drunk, indeed, the researchers described the behaviour of the "zombie ants" as a "drunkard's walk" in their paper. Another weird thing that infected ants start doing is their tendency to crawl all over and bite into leaves - something which healthy ants don't tend to do. There's a good reason why the fungus steers the ant towards leaves and afflict it with this oral fixation - it is preparing it for the next step in the fungus' development.

For the fungus to successfully reproduce, the ant must die - but it must die in a particular position to maximise the viability and dispersal of the fungal spores, specifically in the humid understory, hanging from the underside of a leaf, about 25 cm (about 10 inches) above the ground. But once the fungus maneuver the ant into position, how does it get the host to comply and stay there? The researchers made fine histological cross-section of the infected ant's head and found that once the fungus has made the ant locks its mandible in place, it busily gets to work dissolving the muscles which control those mandibles, ensuring that the ant will be locked in a death grip forevermore. A few days after the ant dies while gripping onto, the fungal stalk emerges from the head of the ant, ready to spray its spores down to the soil below to create more drunken "zombie ants".

Image from the Wikipedia.

Reference:

Hughes DP, Andersen SB, Hywel-Jones NL, Himaman W, Billen J, Boomsma JJ. (2011) Behavioral mechanisms and morphological symptoms of zombie ants dying from fungal infection. BMC Ecology 11:13

Postscript: A few hours after I wrote this post, I found out that Carl Zimmer has already written about this study (why, of course! *facepalm*), so if you want to read his version instead, you can see it here.

Polypocephalus sp.

Today's parasite might be thought of as an "aquatic Toxoplasma" in that it also induces behavioral changes in its hosts. Polypocephalus is a genus of tapeworms that infects both shrimp (Litopenaeus setiferus) and then likely rays such as the Atlantic stingray, (Dasyatis sabina). The larvae of the cestode invade the neural tissue of the shrimp hosts, particularly in the abdominal ganglia. Studies recently showed that the more larval tapeworms a shrimp had, the more time these hosts spent walking on the substrate, as opposed to sitting still or swimming. Although the authors had predicted that they would see an increase in swimming behavior because that might expose them to predation more readily, perhaps just the increased activity in general is enough to promote transmission. Nonetheless, this was an exciting insight into a potentially new system for studying parasite manipulation of their hosts.

Source: Carreon, N., Z. Faulkes, and B. L. Fredensborg. 2011. Polypocephalus sp. infects the nervous system and increases activity of commercially harvested shrimp. Journal of Parasitology 97:755-759

Image from figure of that paper.

Bursaphelenchus xylophilus

Today's parasite is the nematode Bursaphelenchus xylophilus, a well-known tree-killer responsible for the devastating plant disease known as pine wilt. Originating in North America, it has since been spread over much of Asia, and has recently been introduced to Europe. This nematode is transported by longhorn beetles known as "pine sawyers", and gain initial access to the tree through the feeding wound created by that insect. So the arrival of a B. xylophilus-laden beetle pretty much amounts to a death sentence for a pine tree. While pine trees in North America have coevolved with B. xylophilus and developed resistance or tolerance for the parasite, it has caused widespread wilting and death to the pine trees of Japan. So how can such a tiny worm bring down an entire pine forest?

For B. xylophilus, or any other plant parasites for that matter, a tree is a formidable fortress - protected by walls and scaffolding of tough cellulose, and canals of deadly resin. Plant cell wall presents the main barrier to any plant parasites - it is a tough material to break down, and most animals are incapable of doing so without the aid of symbiotic microbes. In addition, the vascular tissue of many coniferous plants like pine are saturated with resin - a thick, sticky cocktail of aromatic chemicals (from which we derive many useful substances including solvents, varnishes, adhesive and perfume) which would overwhelm and kill most invaders. Yet none of those defenses seem to deter B. xylophilus - not only can it break through the thick cellulose barrier of the pine tree, it actually lives within the resin canals of its host, which is practically the most lethal place within the tree. It would be akin to living in a moat of toxic tar.

A recent study published in PLoS Pathogens on the genome of B. xylophilus offers vital clues to how this nematode exploits its pine tree host. One of the most important enzymes for plant exploitation is cellulase - it is used to break down cellulose structures and allow potential parasites to enter and navigate through the host. Bursaphelenchus xylophilus is able to produce a unique combination of 34 enzymes for breaking down cellulose and carries a diverse suite of genes for producing enzymes that detoxify the aromatic compounds found in resin. So how did this tree killer acquire the necessary molecular machinery to invade and disarm its host?

The wide range of detoxifying genes in the B. xylophilus genome appear to be multiple duplication of pre-existing genes which are also found in other nematodes, such as the well-known standard lab worm Caenorhabditis elegans - B. xylophilus just happen to have more of copies of those genes to cope with the wider array of toxins it encounters. However, the cellulase genes have a much more unusual origin. Out of the 34 cellulase enzymes produced by B. xylophilus, 11 of those enzymes are not found in any other nematode, but are most similar to those produced by fungi. So how does a nematode end up producing fungal enzymes?

The answer might be through horizontal gene transfer (HGT). The closest living relatives of B. xylophilus are fungi-eating worms which are transported by beetles to dead and dying trees. Once they reach their destination, they disembark from their beetle vectors and feed on the fungi which have colonised the dead trees. In a case of you are what you eat, the ancestors of B. xylophilus appeared to have incorporated a whole suite of useful genes from their food, allowing them to bypass the process of feeding on fungi which are growing on dead trees and just go straight to breaking down live plant tissue.

Image from figure of the paper.

Reference:

Kikuchi T, Cotton JA, Dalzell JJ, Hasegawa K, Kanzaki N, et al. (2011) Genomic Insights into the Origin of Parasitism in the Emerging Plant Pathogen Bursaphelenchus xylophilus. PLoS Pathog 7(9): e1002219. doi:10.1371/journal.ppat.1002219

Nicothoë astaci

The parasite we are featuring today is Nicothoë astaci, the "lobster louse." Despite its name, it is not a "louse" (true lice are insects) as such, but rather a copepod (a type of crustacean), just like the salmon lice we have previously featured on this blog. But whereas salmon lice are well-studied due to their economic impact on salmonid fisheries (especially on farmed fishes), far less is know about the lobster louse. Despite having been recorded on the European lobster (Homarus gammarus) since the 1950s, to this day there is very little known about this parasite, including the type of pathology it causes, its complete life-cycle, or even what the male of the species looks like (parasitic copepods often have cryptic or dwarf males which are very elusive).

The paper we are looking at today is taking the first step to rectifying that situation. The photo (from the paper itself) depicts larval stages of N. astraci, with the arrows indicating the oral cone,the structure this parasite uses (along with its front pairs of legs) to attach itself to the host's gill filament and feed on its blood. While the larval stage looks like a rather ordinary copepod, as it matures into an adult, it morphs into what looks like a miniature boomerang with a pair of stretched out "wings" on either side, and a pair of bulbous egg sacs dangling from its rear end. The attachment and feeding activity of the lobster louse can cause pronounced physical damage to the lobster's gill filaments.

As with any kind of infection, you would expect to see some kind of cellular response. While the innate immune systems of invertebrates like lobsters are not as sophisticated as the adaptive immune system of vertebrate animals such as ourselves, they can present a formidable challenge to any would-be intruder (to see an example of what the cellular defence of a crustacean can do to a parasite, click here). Basically, the crustacean's equivalent of blood cells wrap themselves around the parasite or pathogen and initiate the process of melanization, where the intruder becomes entombed in a hardened capsule of melanin (the pigment which determines our skin colour). The researcher did find signs of melanization and other cellular disruption throughout the gills of infected lobsters, but none of it was near the lobster louse's attachment point.

So the lobster's immune system recognizes the presence of an intruder, but is unable to pinpoint and focus its wrath on the parasite. The authors of this paper suggest that this indicates the lobster louse is able to somehow interfere with the lobster's defensive mechanism so that it can blood-feed in peace. The mechanism through which the lobster louse disrupts this particular aspect of host physiology is yet to be uncovered, along with much of the parasite's ecology and life-cycle. Hopefully, with further research on this host-parasite system, this situation will change in the future.

Image from the paper.

Reference:

Wootton EC, Pope EC, Vogan CL, Roberts EC, Davies CE, Rowley AF. (2011) Morphology and pathology of the ectoparasitic copepod, Nicothoë astaci ('lobster louse') in the European lobster, Homarus gammarus. Parasitology 138:1285-1295.

Maritrema subdolum

This is a story about two little crustaceans and a parasitic fluke. Corophium volutator and Corophium arenarium are amphipods that make a living tunneling in the mudflats on the coast of the Danish Wadden Sea. They are also host to many species of parasitic flukes, and one of them is the parasite we are featuring today - Maritrema subdolum. Last year, we featured a related species from New Zealand - M. novaezealandensis - and much like its Kiwi cousin, M. subdolum is the bane of the local crustacean population. However, M. subdolum does not affect all of its crustacean hosts equally and this has some important ecological consequences.

Out on the Danish mudflats, C. volutator is king. Of the two Corophium species, it is the stronger competitor, reaching much higher abundances and generally making life difficult for C. arenarium. But along comes M. subdolum, which evens out the playing field. Living alongside those little crustaceans are mud snails, and during early spring they can become extremely abundant, with over 25000 snails per square metre. By summer, almost half of those snails are infected with M. subdolum, which turn them into little parasite factories, each cloning a massive reserve of parasite larval stages call cercariae that are then unleashed into the environment to infect the amphipods. The main trigger for releasing these cercariae is high temperature, and during 1990 there was a bout of unusually high temperature during summer that has been linked to North Atlantic Oscillation (NAO), a phenomenon that plays a major role in determining climatic conditions in the northern hemisphere.

All of this combined into a perfect storm that devastated the C. volutator population. Triggered by high temperature, all the infected snails released their payload of parasites into the surrounding waters. Each infected snail can release hundreds of cercariae, and with thousands of snails per square metre, the shallow waters of the mudflats turned into a seething parasite soup. To the amphipod, this amounted to being in a shooting range, as each cercaria is armed with glands of digestive enzymes and a scalpel-like organ call a stylet with that they use to puncture the amphipod's exoskeleton. For many C. volutator, the outcome of being attacked by a swarm of these little horrors was terminal, and this resulted in a dramatic decline in their population. However, for some reason we still don't know, the C. arenarium population was able to weather the M. subdolum storm unscathed, either because they tolerated the parasite swarm, or because they were simply not the preferred target. Either way, with the collapse of the C. volutator population, in the next season, C. arenarium was able to succeed them and become the dominant amphipod species on the Danish mudflat.

And this dramatic ecological change was ultimately brought about by a little parasite.

Photo by Kim Mouritsen

Reference:

Larsen, M.H., Jensen, K.T. and Mouritsen, K.M. (2011) Climate influences parasite-mediated competitive release. Parasitology 138: 1436-1441

Metarhizium acridum

The locust in the photo is covered in a fine layer of green mold - that is because it was killed by the parasite we're featuring today, Metarhizium acridum. Metarhizium acridum is a pathogenic fungus which specifically infects and kills grasshoppers, locust and other insects in the Orthoptera order. Because of the pest status of some orthopterans (think locust plagues), M. acridum is mass-produced as type of environmentally-friendly, biological alternative to most insecticide. But while M. acridum only targets locust and grasshoppers, its close relative, M. robertsii, is far less picky, capable of infecting hundreds of different insect species.

So why is M. acridum so picky while its close relative is so indiscriminate? Amazingly, it appears to come down to a single gene call Mest1 - a gene present in M. robertsii, but is absent in M. acridum. To find out the function of this gene, a group of researchers in China created a mutant M. robertsii strain which has a non-functioning copy of Mest1. This mutant lost its ability to infect most insects - except grasshoppers and locusts - which happens to be the speciality of M. acridium. In parallel, the researchers also inserted functional copies of Mest1 into M. acridum. The insertion of this single gene allowed M. acridium to infect a wider range of insects.

What is so special about Mest1? In M. robertsii, Mest1 is expressed during spore germination, and plays an important role in initiating the infection process. Mest1 expression can be triggered by a range of stimuli including nutrient poor conditions or contact with insect cuticle. Metarhizium acridium has other genes playing the role of Mest1, but they are triggered by substances which are present only in the waxy coating of grasshoppers and locusts. So if its spores land on other insects such as caterpillars which have a different type of coating, M. acridum fails to germinate because the appropriate stimuli are absent. Thus, the insertion of Mest1 into M. acridium allows the fungus to bypass those usual stimuli and begin germinating under a wider range of conditions

Host specificity is one of the central question in the evolutionary biology of parasitic organisms. In this case, we can see how a single gene can changed this otherwise specialist pathogen into a broad-spectrum generalist.

Image from the Wikipedia.

Wang S, Fang W, Wang C, St. Leger RJ (2011) Insertion of an Esterase Gene into a Specific Locust Pathogen (Metarhizium acridum) Enables It to Infect Caterpillars. PLoS Pathog 7(6): e1002097. doi:10.1371/journal.ppat.1002097