Termite gut microforay - excavate time!

A couple days ago a local basal winged termite crossed my path. Then, like two microtubules approaching each other below the threshold angle, our paths zippered up together and the poor sucker had its parabasalian jewels strewn exposed on the slide. No worries, the termite lives on, albeit as a carbon source for other life...

Now I have rather limited experience working with gut endosymbionts (my second time), so I suck. Also, I didn't have the right solutions for keeping the denizens from exploding osmotically. So my images suck too. Kind of like this poor exploding trichonympha:

Actually, they seemed to be less exploded later when mounted in 5% glycerol (or perhaps they managed to die in the termite while I was working on the first sample, and experienced less osmotic shock that way somehow...) The glycerol may have helped out in the refractive index matching a bit, although perhaps I just magically came across better DIC settings later... (I need to stop speaking in strings of hypotheses. [fundie]I believe the glycerol helped my imaging, and I shall guard my faith until forever! Yeah, take that, 'evidence'! In other news, termite endosymbiosis is older than the earth itself. [/fundie])

First thing I saw was some mystery nematode: (was wiggling about way too much for a decent image) How do you key out nematodes anyway???

Top right: Some mystery...thing. Looks like an ex-organism, judging by the characteristic bubbles that tend to form when anaerobic protists die. Then again, those may well be artefacts of something else. Any ideas?
Bottom:Trichomitopsis! (it actually looks really cute when not completely mutilated)

(the scalebar on the second one is wrong; that would be about 12.5um, forgot to adjust for 40x settings...)
A slightly better shitty image of streblomastix; how many flagella can you count at the anterior end?

And fresh from the Crap on the Bottom of the Slide department, some mystery flagellates(?), or pieces of dirt sitting on top of bacteria (Or elongated pieces of crap). Hey, identifying whether crap is/was alive or not is kind of difficult sometimes! Although I'm pretty sure (B) is a real thing. (D) also looks kind of flagellated... any ideas?

Edit 18.08.09: With Opisthokont's help, possible IDs: A - hexamita?; B, E - monocercomonoides; C- a thing with an axostyle, or hexamita)

Then I had fun with the carnival ride that is a prism and and couple polarisers (aka DIC):

Trees in the autumn sky...

...roots on the autumn ground?

Shiny!

Omg, pseudo-darkfield! (no it's not)


The helical spiraly thing is a piece of xylem, methinks; termites eat wood right? Well, trichonympha eat wood inside the termite... so you get pieces of plant matter in them. And then you find random stomata floating about, which I'm pretty sure are not some weird contamination from my arabidopsis screening stuff (kinda reused the slide...)

Those things freaking haunt me, even when not doing research! Amazing how it survived intact...stomata can be quite fragile. Although perhaps that's specifically when you need them not to be...
Actually, wait, question: how the hell do stomata get in there if termites eat wood? While plants do have stomata on their stems, 1. termites don't munch on epidermal tissue all that muchl; 2. I doubt woody plants have any once bark is formed. Maybe it -is- some weird contamination from the previous sample...luckily, not actually doing any real science here.

Speaking of stomata, I has works to do. We're kind of like plant dentists - working on 'mouthes' all day! Especially with gene names like four lips(FLP), too many mouthes(TMM), speechless(SPCH), mute(MUTE), moustaches(MUS), etc. We also have YODA. Apparently somewhere in the Arabidopsis genome lies a regulatory gene SUPERMAN and its suppressor KRYPTONITE. We like to keep ourselves entertained...[/derail]

Let me know if you want more 'microforays'!

Sunday Protist - Streblomastix: intestinal torpedo-bearing sub

Taking a bit of a break from Rhizaria... haven't done Excavates in a while. From one obscure 'kingdom' to another... (although now I have this nagging feeling that I'm really neglecting unikonts - I haven't done amoebozoa or opisthokonts in ages...)

And the creature behind (or rather, containing) Mystery Micrograph #01 is...

(Leander & Keeling 2004 J Euk Microbiol; scalebar=5μm)
Streblomastix strix, an oxymonad. 2 - cross-section

Rosie more or less got this one ^.^ (I guess one can't really expect a sane person non-protistophile to randomly pass by and yell out OMG STREBLOMASTIX!!1!) It's a thing with several extremely long episymbiotic bacteria riding along it, kinda like torpedoes, so the 'eukaryote with bacterial symbionts' was right!

I won't go into detail about oxymonads, since Opisthokont knows a few orders of magnitude more about them than I can pretend to. Perhaps he'd like to contribute some. [/hint] Essentially they are an anaerobic, amitochondriate lineage of mostly termite/cockroach gut endosymbionts. For some reason, most of those gut endosymbiont protists tend to gather entire bacterial worlds around (and inside!) them. Perhaps due to the closed, small nature of the ecosystem, where there is time for various prokaryotes to coevolve with the protists and become closely associated without too much outside disturbance. These protists have been living with termites and cockroaches at least since the early Cretaceous (Poinar 2009 Parasit Vectors), so there's been plenty of time for entire phyla to be borne of this relationship.

Oh, apparently there's such thing as hunting around for termite endosymbionts in amber! Couldn't find any of Streblomastix, but here's a ~100 x 10⁶ year old relative, Dinenymphites:

(Poinar 2009 Parasites & Vectors; open access)

That paper is rather fascinating! And this is from someone who's not much of a paleontology nut. Anyway, back to Streblomastix.

An interesting question is, what are those bacteria doing? The cell structure seems adapted for episymbiosis, complete with specialised attachment sites. Sure enough, killing off the bacteria with antibiotics results in the eukaryote's imminent death (Leander & Keeling 2004 JEM). The specifics of the relationship still remain poorly understood, it seems. There's also even more elusive intracellular bacteria. Basically if you look at an animal or plant carefully enough, you'll find entire ecosystems of protists and prokaryotes. If you look at a protist carefully enough, chances are there'll be an entire bacterial ecosystem in its own right...but I digress.

Leander & Keeling 2004 J Euk Microbiol; m1-3 - three distinct morphotypes of bacteria, scalebar=0.5um; 16 - TEM cross section of a 'vane' tip with arrows showing the glycocalyx-like connections between bacteria and the host. 17 - posterior tip of Streblomastix showing the 'cupping' of the final bacterium.

Actually, a winged termite (Zootermopsis!) had the great misfortune of appearing before my face yesterday. This is a sample of the result:

Yes, the shittiest picture of Streblomastix in the history of termite gut microscopy. I tried to get the flagella in focus, but the whole cell is pretty mangled. The Trichonymphas turned out better, but barely... Sigh, I really need Trager medium... and access to a camera that is not too slow even for plants. I guess that means no colour CCD, since those tend to need a lot of light, and therefore a much longer exposure. Although there must be fast colour cameras too...

(still must redeem self:

Trichonympha. Happy? More termite gut imagery to follow later...)

Ok, it's getting late, I think I'm getting a cold of sorts (I wish it'd make up its mind already, whether it feels like showing up or not. And please, let it be finished before I visit the parents. Mothers + cold = overreaction x 1000!!!) and I'm just gonna have to leave this post in a half-sucky state. Did any of it make sense?

As for the next major topic, it'll be the origins of Eukarya and the Neomuran Hypothesis, with plenty of famously clear and comprehensible diagrams by the God of Protistology. Also, there's a recent Science article by Carl Zimmer that awaits a bit of gentle shredding. Actually, I'll wait until Rosie's reply to his sex article is published next week =D But prior to Neomura, there is a massive rant in store. Evolutionary directionality is GOING DOWN! Mwahaha...

And as for ye who foolishly voted 'cheese!' in the poll - watch out! It's coming...
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LEANDER, B., & KEELING, P. (2004). Symbiotic Innovation in the Oxymonad Streblomastix strix The Journal of Eukaryotic Microbiology, 51 (3), 291-300 DOI: 10.1111/j.1550-7408.2004.tb00569.x

Poinar GO (2009). Description of an early Cretaceous termite (Isoptera: Kalotermitidae) and its associated intestinal protozoa, with comments on their co-evolution Parasites and Vectors, 2 : 10.1186/1756-3305-2-12

Leaves on drugs

I accidentally hit Shift+I in ImageJ, which inverts colours. The result was very trippy:

(originally an Arabidopsis leaf surface; yes, drugs have been involved - for once, on plants rather than from plants...)
Seriously, doesn't this almost look like some modern abstract painting thing?

Going back to working on the carpal tunnel syndrome quantifying microscopy data...

Btw, I proudly use ImageJ for most of the cropping/editing work in this blog. I am not a nerd. To prove the latter point, I should perhaps someday have a post dedicated to revealing the inexhaustible Awesome hidden in the ImageJ universe. We microscopists have a freaking fetish for it... >_>

Mystery Micrograph #01

Prelude to Sunday's topic:
(to be referenced later)
Anyone wanna take a stab at what's going on here? This image stumped many during a lab exam...

You have until Sunday, say 8pm Pacific Time.

Reward: Ehhh... feelings of triumph and superiority? A free beer if we ever meet in real life? 50 IKUs (Internet Kudos Units)?

Good luck!

Note: People involved with course in question forbidden from participating. As are certain other members and ex-members of the department. You know who you are.

Approvals are always nice =D

YAY! Application to coordinate a student directed seminar on 'evolutionary theory in the humanities' got approved! (plan is: 1/3 biological evolution (emphasis on molecular and microbial stuff that usually gets thoroughly ignored), 1/3 evolutionary linguistics, 1/3 critical analysis of memetics and applying prev two sections to other humanities fields; entire thing is participant-led, so no teaching; anxiously anticipating what happens to the final project, which will be literature-research based)

As excited as I am right now, there is A LOT, A freaking LOT of work to do now. This is going to be a nightmare, especially since we're trying to span two faculties and countless departments therein. But it's exciting! So lucky our university has this opportunity...

Anyway, still got a few hours of microscopy aherad of me... waaah!

Oh, and ha!: "Memetics is dead" my sorry ass! =P

I plan to post more details later, but any suggestions/criticisms/discussions are welcome and encouraged! If you know of any potentially useful sources/references, please let me know too!

Back to lab slaving... can't wait to finish so I can go home and devour my protist porn Research in Protozoology volumes...

PCR Flu

After a week of my PCR failure, I finally got it to work. However, a masters student now can't get his alive in any way, which prompts one to contemplate the possible existence of some infectious PCR disease, a flu if you will, that spreads around labspace plaguing one unfortunate victim after another.

A new journal title is in order: J. PCR Flu Epidemiology. I think I finally found a place to publish my gels!

Said journal should also feature PCR horoscopes:

eg.

Aquarius: The -something- is in the way of the path of -some star- (I don't really read these, can you tell? o_O) Check your dNTPs. Your entire stock may have gone bad.

Pisces: While at first you may feel confident following a long line of successful gels, you should be extremely cautious in a couple of weeks - treat your Taq with care and be sure to put all the reagents back in the -20.

Cancer: You're fucked. Seriously, go wash some glassware and clean the lab windows instead for the next few weeks... nothing you do will save your product from instant degradation by the wrath of the PCR gods.

Etc.
Anyway, I have a long night of microscopy ahead of me... good bye life, it was nice having you!

PS: I got catnip: 3 volumes of 1969 Research in Protozoology (ed.: Chen) from a nice ciliatologist. It's simply incredible. I'm dehydrated from drooling over it so much. Protistologists are very evil people. Worse than drug dealers... =P

Sunday Protist - Ebria

Someone here wanted Ebriids and Ellobiopsids. Since I'm still in a bit of Rhizarian mood, let's do Ebriids first. Just as you thought we couldn't get any more obscure than Phaeodaria...

Ebriids are biflagellate Cercozoans (see the Pawlowski & Burki 2009 Rhizaria tree in the Coelodiceras post for their phylogenetic neighbourhood) with a characteristic silica endoskeleton, permanently condensed nuclear chromatin and lack of cell wall or scales. They are rare and still unculturable, so very little is known about them. The taxon name derives from ebrius ('drunken' lat.) as they apparently swim in a strange fashion. (Hoppenrath & Leander 2006 Protist)

They feed on diatoms and dinos, likely using pseudopodia in their feeding (an interesting feature of Cercozoans is their affinity for pseudopodia despite being flagellate...) Interestingly, they too, like some dinos, can devour diatom chains in a process that seems similar to palium feeding. Due to their silicoflagelate nature, they have been caught up in the taxonomic chaos and have been considered to be dinos or radiolarians; they were thrown around between the two for decades, until the eerie Enlightenment through molecular biology placed them in Cercozoa. (all from Hoppenrath & Leander 2006)

(Hoppenrath & Leander 2006; arrow in A points to nucleus; scalebar: 10μm)

The silica endoskeleton leaves behind a nice fossil record stretching back to the Cretaceous, although very few modern species have been found (Hargraves 2002 Plankton Biol).

(Korhola & Grönlund 1999 J Paleolimnol. Silica Ebriid skeletons from Baltic sediment)

I wonder if their genomes too conceal some quirky stuff. Permanently condensed chromosomes tend to do that (see dinos; Euglenids have them too but nothing seems to be known about their genomes yet).

A bit tired and sleepy... apologies for low energy post today.
---
As for "just why is everything about dinoflagellates just so *%&$ing weird?"* - Simple. On the 7th day of creation, the intelligent designer took a break, got really high off some awesome shit he'd just finished creating, and made the protists. Dinos were created near the climax of his hallucinations, while ciliates were definitely made right AT the climax. Thus, ciliates are the Higher Eukaryotes, with dinos and the rest of Protista close behind. Now for a reading from the Book of Tom... oh, has anyone tried that while 'under influence'? Does it help?

*In case some are as clueless about dinos as I was less than a year ago, the following is some of the oddities:
Nucleus
- permanently condensed chromatin
- massive genomes (>10x that of humans)
- chromosome structure on crack
- mandatory trans-splicing of 5'-cap-bearing splice leaders onto mRNA transcripts. Ie each mRNA transcript must have another specific mRNA sequence attached to its beginning, otherwise it gets degraded. Interestingly, Trypanosomes and some genome in C.elegans do that too, and have polycistronic genes, which is highly unusual for eukaryotes. Ie. several genes following one promotor... more on that later sometime!

Mitochondria
- tiny genomes
- rRNA assembled from fragments, some of which are imported from the nucleus
- some species: single gene linear chromosomes; some have long scrambled repeating permutations of three conserved mitochondrial genes (COX1,3; COB)
- post-transcriptional RNA editing; ie a transcript is made, and then some bases are modified prior to translation
- trans-splicing (of exons located on different loci) (usually splicing happens in cis, by remvoing a region between two exons; trans-splicing involves taking two exons from different regions and gluing them together.

Plastids
- single-gene minicircles - genome consists of tiny circles usually containing a single gene and a replication origin sequence.

Misc
- closed mitosis (Nu envelope doesn't break apart; although not that unusual)
- prone to tertiary endosymbiosis (eg. Karlodinium, Karenia, Kryptoperidinium, Lepidodinium, Dinophysis)
- diversity of lifestyles, from endo- and ectoparasitism and endosymbiosis to myzocytosis (sucking prey through a 'straw') to palium-feeding; all while roughly half of them are photosynthetic
- Warnowiid dinos with... camera eyes!

Since it's kinda late and I'm sleepy and don't have time to reference all that... let's just pretend this footnote never happened, k? ^_~ I should blog about this properly later...
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Hoppenrath, M., & Leander, B. (2006). Ebriid Phylogeny and the Expansion of the Cercozoa Protist, 157 (3), 279-290 DOI: 10.1016/j.protis.2006.03.002

Hargraves, P.E. (2002). The ebridian flagellates Ebria and Hermesinum Plankton Biology and Ecology, 49 (1), 9-16

Korhola, A., & Grönlund, T. (1999). Observations of Ebria tripartita (Schumann) Lemmermann in Baltic sediments Journal of Paleolimnology, 21 (1), 1-8 DOI: 10.1023/A:1008019504122

"Two things are infinite..."

I generally refrain from bashing creotards here as it's done much better in other blogs, but the sheer authentic pure idiocy here is simply astounding. I mean, just, wow.

So naïve. Dawkins, I mean. Does he seriously think it's possible to have a rational reasonable any conversation that...[censored]...silly person?

Added to the So Stupid One Cannot Come up with Valid Counter-arguments list.


PS: Vote against the cheese! --->

(The cheese option comes from a joke on a forum, where they refuse to vote in a poll devoid of cheese option. However, they weren't supposed to pick the cheese option itself, those cheeky bastards! Hey, to all you voting cheese: Don't make me go into the biochemistry or microbiology thereof; it doesn't mean you'd just get away with appetising pretty pictures!)

Help me pick the next topic!

Since I'm a disorganised indecisive scatterbrain, how about you help me pick the next topic? Otherwise I start writing 10 posts in parallel and never finish any... (my drafts folder is massive...) I've set up a poll to the right --->

Also, additional suggestions definitely welcome and encouraged! As are random questions/comments/musings/criticisms/rants/etc. You guys need to comment more proliferously. Especially those whose blogs I plague with mine ^.^

Blue isopod time:

Found it in California, never seen a bluish-purple one before. I wonder if it's an allelic thing, or actually a separate species altogether...

Sunday Protist - Obscure Phaeodarian: Coelodiceras

Choreocolax and Ecomonymopha not obscure enough? Let's go for Phaeodaria then! I've been neglecting Rhizarians, just like everyone else. When I first saw a eukaryotic tree, I could recognise a thing or two in most of the 'kingdoms'. Except one: Rhizaria. All those names were absolutely meaningless to me. Those wonderful earthly aliens desperately need an introduction to the world beyond dusty 1970's oceonography journals!

Rhizarian taxonomy (nitpicky details alert)
Rhizaria is a very morphologically diverse group held together mainly by molecular data. It has everything from amoeboid Chlorarachniophytes, 'famous' for secondary endosymbiosis of a green algal plastid; seashell-like Foraminifera; flagellates like Heteromita, scaly-tested Euglyphids; to spiny Radiolaria, so wondefully illustrated in Haeckel's Kunstformen der Natur (pdf of original available on that page). It's a tremendously understudied group...

So where do Rhizaria fit in the tree of life? From the same year: Moreira et al. 2007 Mol Phylogenet & Evol: basal to Chromalveolates and Archaeplastids; Hackett et al. 2007 Mol Biol & Evol: basal to stramenopiles+alveolates. Shit. Conundrum. Knowing nothing about trees, I could resort to Proof by Impact Factor:

MPE: 3.871
MBE: 7.280
Branching with stramenopiles+alveolates it is. QED.

Alternatively, we could check later sources: Baldauf 2008 J Systemat & Evol and Keeling 2009 J Euk Microbiol place the Rhizarians basal to stramenopiles+alveolates, although not fully certain, whereas TC-S (sen. author of Moreira et al. paper) insists on sticking it basal to chromalv+archaeplastids in his latest megaphylogenetic megatree of megaeukaryotic megaevolution (TC-S 2009 J Euk Microbiol). Time for Proof by Democracy...

And suddenly I realise probably no one here (except the taxonomist =P) really cares where Rhizaria roots, so let's move on.

Another taxonomic troublespot is Radiolaria. Originally they included Acantharia, Spumellaria, Nasselaria and Phaeodaria, but it turns out Acantharians are quite unique due to their strontium(!) sulfate skeletons (as oposed to silica in Nasselaria and Spumellaria), while Phaeodarians are a whole other thing altogether, quite distant from the rest of the 'radiolaria'. Nowadays, Nasselaria and Spumellaria are clumped into Polycystea, with Acantharians forming their sister clade. Like this:

(Pawlowki & Burki 2009 J Euk Microbiol)
(Phaeodaria are in Cercozoa)

Ah, so many more new obscure organisms to read about! *drools*

Coelodiceras spinosum
Now for some Coelodiceras spinosum from Paterson et al. 2007 Deep Sea Research Part II: Topical Studies in Oceanography:
(Scalebar: 200μm)
Phaeodarians are unicellular organisms composed of a fine cytoplasmic mesh surrounding a delicate silica skeleton. They contain a central capsule with a large opening leading to a phaeodium, a congregation of food and waste vacuoles. They are fierce predators devouring anything from bacteria to dinoflagellates and diatoms by catching them with rhizopodia, or fine thread-like extensions of the cytoplasm. Some have a single large polyploid nucleus (Dogiel' 1965 General Protozoology). Would be pretty interesting if this one also has only a single large nucleus, since cells this big tend to be multinucleate.

The most is known about their skeletal structure, since that is what is easiest to preserve and deal with. They fossilise well, and are used as indicators in the oil industry. Sadly, almost nothing is known about the genetics and developmental biology of these organisms. Many of them inhabit the deep sea (eg. Coelogiceras), and are difficult or simply impossible to culture. As a rule of thumb, predators are complicated to deal with, especially if their lifestyle is unknown - sometimes they may require auxiliary organisms to be present to process their prey in such a way that they can make use of it, for example. It's a pain.

Some more details of the skeleton: (again, from Paterson et al. 2007)



It would be fascinating to know the function and development of these intricate structures, but the paper is mostly descriptive (the specimen was already dead before collection), and finding further information on their cell structure is incredibly difficult. This is probably the most difficult post I've yet written, with about two hours of literature research going into each paragraph on average. But I hope this inspired some appreciation of how complex a unicellular organism can get!

References
Dogiel, V. A., (1965) General Protozoology. Revised by J. L. Poljanskij and E. M. Chejsin (II. Auflage) VII und 747 S., 326 Abb. Oxford: Clarendon Press

Hackett, J., Yoon, H., Li, S., Reyes-Prieto, A., Rummele, S., & Bhattacharya, D. (2007). Phylogenomic Analysis Supports the Monophyly of Cryptophytes and Haptophytes and the Association of Rhizaria with Chromalveolates Molecular Biology and Evolution, 24 (8), 1702-1713 DOI: 10.1093/molbev/msm089

KEELING, P. (2009). Chromalveolates and the Evolution of Plastids by Secondary Endosymbiosis Journal of Eukaryotic Microbiology, 56 (1), 1-8 DOI: 10.1111/j.1550-7408.2008.00371.x

Moreira, D., von der Heyden, S., Bass, D., López-García, P., Chao, E., & Cavalier-Smith, T. (2007). Global eukaryote phylogeny: Combined small- and large-subunit ribosomal DNA trees support monophyly of Rhizaria, Retaria and Excavata Molecular Phylogenetics and Evolution, 44 (1), 255-266 DOI: 10.1016/j.ympev.2006.11.001

PATERSON, H., PESANT, S., CLODE, P., KNOTT, B., & WAITE, A. (2007). Systematics of a rare radiolarian—Coelodiceras spinosum Haecker (Sarcodina: Actinopoda: Phaeodaria: Coelodendridae) Deep Sea Research Part II: Topical Studies in Oceanography, 54 (8-10), 1094-1102 DOI: 10.1016/j.dsr2.2006.05.046

PAWLOWSKI, J., & BURKI, F. (2009). Untangling the Phylogeny of Amoeboid Protists Journal of Eukaryotic Microbiology, 56 (1), 16-25 DOI: 10.1111/j.1550-7408.2008.00379.x