Trouble-shooting Avizo (and Amira by extension)

Some of my work involves dealing with 3D images, either CT scans, MRI scans, or z-stacks from a confocal microscope. To analyse these images there are a variety of tools available but the industry standard seems to be Amira. Amira is an incredibly powerful piece of software not just for analysing 3D images, but also for editing them and making them look pretty for figures and presentations. Unfortunately, it comes with the kind of price tag you'd expect. As an academic user, I'd be able to purchase Amira at half price, about $5000 AUD, before taxes and extensions. For my work, I'd probably ultimately need at least the quantification+ extension, and possibly the neuro and large data set extensions as well.

Fortunately, I've been able to avoid this insurmountable expense through collaborations. I collaborate with biologists, neuroscientists and computational wizzes at Monash University and the University of Queensland. Because I am a staff member at Monash, I've been able to access the MASSIVE remote desktop. Through this desktop I have access to Amira, almost. MASSIVE does not have a licence for Amira. Instead they have a licence for Avizo. Avizo, to my understanding, is the same program as Amira, with a different skin. Amira is geared towards the life sciences, while Avizo is geared towards the physical sciences and engineering, but they both ultimately do the same thing.

Since I have access to Avizo but not Amira, I've been using Avizo for things that Amira's really meant for. Furthermore, as far as I can tell (and I've done some sleuthing), I'm the only person at the Australian National University using Amira/Avizo (side note: if you're at ANU and use either of these programs, please let me know!) That means, though I have support from experts at the University of Queensland and Monash, I've largely been on my own in figuring out how to use Avizo/Amira. Usually, when I have problems figuring out how to use software, Google is my saviour, however I've found the online resources for troubleshooting Avizo/Amira woefully inadequate (again, if you know of good online resources for this, please let me know!)

As I've been learning to use Avizo/Amira, I've encountered quite a few challenges, which I've mostly solved through trial-and-error, with some desperate e-mails to experts. Since I haven't been able to find these solutions online, I thought I'd put them here in a series of posts, so that the next desperate person might solve these problems without wasting quite as much time banging their head against a keyboard as I have. I'd like to note that these problems are beginner problems and will look very simple to the advanced (or even just regular) Avizo/Amira user, but they were quite frustrating at the time!

First, I must say that the Amira training manual is very useful for learning both Amira and Avizo, and is very easy to use. I've done Chapters 2 and 7, which teach the basics (including segmentation, the tool I use most) and animations, respectively. The troubleshooting I describe will assume knowledge of Chapter 2, sections 2.1-2.5, which cover loading an image, viewing an image, and segmenting an image. The reason I use the Amira training manual rather than the Avizo training manual is that the Amira manual uses a neuroscience example (a bee brain) and neuroscience is what I do, so it's much easier to directly translate the examples to my work.

In addition to the steep learning curve involved in learning to use Amira/Avizo, there are two additional sources of confusion that I've found. First, there are some things that are different between Amira and Avizo which can make the Amira training manual difficult to follow in Avizo. I'll cover the issues I've found later. Second, I'm a Mac/Linux user, and I think that there are some bugs in Amira/Avizo that are particular to these two systems. Here's the first one:

Using the "hot keys" of the Image Segmentation Editor in OS X or Linux

First things first, and this might seem simple, but it was really confusing at the time. Any of the "hot key" commands (what Avizo/Amira calls keyboard shortcuts) that use the Control key on PCs STILL use the Control key on Macs. This is despite the fact that every other program I've ever encountered uses the Apple/Command key in place of the Control key for keyboard shortcuts on Macs. Get used to it, Mac users, it's the Control key in Amira/Avizo. Here is a PDF of the Segmentation Editor's hot keys.

Second, as far as I can tell, not all the hot keys actually work (at least for me, using OS X and Linux). I made this table as a guide for myself so I would know if a key command normally works or if I'm going nuts. Most of them do work for me and I recommend going through each of them one-by-one, just to see what they do. If I'd done that two years ago, I'd have saved a lot of time!

The issues I'm going to go through on this blog are similar to this one. They will be very basic to the regular or advanced Amira/Avizo user, but boy would I have been overjoyed for someone to have told me these things when I was learning! If you have a particular issue with Amira or Avizo, please let me know and I will help if I can.

 

Australian Pastoral Scenes

Spring is, to me, a time of idillic pastoral scenes played out in real life. One of the many fun things about rural Australia is the abundance of kangaroos where you would expect deer or domestic ungulates. Sometimes, driving around in Australia, I notice a scene that reminds me of an Alvan Fisher painting of rural nineteenth century America, except with kangaroos.

Of course, Australia is full of domestic ungulates, in addition to the roos.

Spring is here!

Spring has finally sprung in Canberra! I'm wearing t-shirts again, we're eating lunch outside in the sun, and babies seem to be popping up all over the place. Last season's baby eastern grey kangaroos are just about ready to leave the pouch. They are super cute, looking like the under-sized second head of a two-headed roo.

They look even more awkward when they dive into their mother's pouch head first.

This does not look comfortable. Tidbinbilla Nature Reserve, Australian Capital Territory.

The roos aren't the only ones with babies. There's an adorable family of musk ducks living on Yerrabi Pond in Gungahlin. Musk ducks are weird, unique animals. The males have a very strange, fleshy lobe hanging from their bills. Both sexes have spiky tail feathers that are unique amongst Australian ducks. Unfortunately we found the family just after sunset. The only pictures I have of the mother and duckings are silhouetted, and the uniqueness of the musk duck isn't really visible. However, I was able to get close enough to the father for the flash to be effective, and so both the fleshy lobe and the spiky tail are visible in the last picture.

While looking for the musk duck family I found a purple swamphen family on their massive nest of reeds. The purple swamphen is an extremely successful animal here in Australia, benefiting from human habitation and the creation of artificial waterways. They are a common sight around the creek that runs through the ANU campus and any Australian wetlands. Nonetheless they are a beautiful and charismatic bird and a pleasure to see.

Sunset may be an awkward time to photograph birds, but it is an excellent time to photograph sunsets:

Vegetation-dwellers, group 2

In his book "Biology and Evolution of Australian Lizards", Allen Greer sorted the lizards I study, the genus Ctenophorus, into different groups based on where they live: those that live in burrows, those that live in rock crevices, and those that live in vegetation. Looking at the phylogenetic relatedness of the Ctenophorus dragons, there are two groups of each: two groups of burrowers, two groups of rock-dwellers, and two groups of vegetation-dwellers. I thought it'd be fun to put up pictures and descriptions of all these groups, since I have a large pile of pictures from my fieldwork. These are my posts so far: Rock-dwellers #1, Rock-dwellers #2,  Burrowers #1, Burrowers #2, and Vegetation-dwellers #1.

This last group of Ctenophorus dragons are known as the military dragons. I don't know why, maybe because their patterns are full of stripes. They are also the largest group of Ctenophorus, with seven species. They're all quite small, weighing around 10 grams or less, and are long-legged sprinters. They don't shelter anywhere in particular, so they use a combination of their speed and any available vegetation to hide from predators. During my PhD, I deliberately went after two species, and we came across a third coincidentally while looking for other dragon species.

The two species in this group that I needed for my PhD were the mallee military dragon (Ctenophorus fordi) and the central military dragon (Ctenophorus isolepis). These two lizards are almost the exact same. They are quick, skittish lizards that are always on the flat ground. They never perch on branches, climb trees, or sit on top of bushes as other dragon lizards often do. They do everything - bask, hunt, mate, etc. - on solid ground. They also have a preferred habitat type, spinifex, and are so closely associated with spinifex that we called them the spinifex dragons. Spinifex is a famous type of grass here in Australia, famous mostly because it is basically a clump of outwards-pointing spears. I vividly remember my volunteers trying to catch mallee military dragons by hand; I had horrible visions of them poking their eyes out on the spinifex as they dove for dragons. The dragons use this to their advantage. If they're scared, they dive into a dense spinifex bush, which is basically an impenetrable fortress of spikes!

The immense spikiness of a spinifex plant. Mulga Park Station, Northern Territory, 2012. Photo by Angus Kennedy.

There were a few key differences between the mallee and central military dragons. First, the mallee military dragon was extremely abundant. We broke all records for single-day lizard catching with this species when, on the very last day of my 2012 field season, we caught 32 mallee military dragons between noon and 5 p.m. That is a lot of dragons! On the other hand, central military dragons are only moderately abundant. It took us two days to catch the same number of central military dragons as we caught mallee military dragons in half a day. This is still much more abundant than any other dragon species. That number of dragons would usually take us 4-6 days to catch, and with the rusty dragon it took three weeks!

Central military dragons are also about double the size of mallee military dragons. Mallee military dragons prefer their spinifex to be under the shade of a canopy of mallee trees, whereas central military dragons like their spinifex exposed, with no trees in sight, like in the picture above.

An additional difference is that central military dragons are sexually dimorphic, meaning that the males and females look different, while mallee military dragons are sexually monomorphic, meaning both sexes look the same. In the case of the central military dragon, this means that the male has a heck of a lot more black on him than the female, and has a more complex pattern. Male and female mallee military dragons look almost identical. There is no sexual dimorphism in this species, except for some extra black markings on the underside of the male.

A male central military dragon (Ctenophorus isolepis). Mulga Park Station, Northern Territory, 2012. Photo by Angus Kennedy.

A female central military dragon (Ctenophorus isolepis). Mulga Park Station, Northern Territory, 2012. Photo by Angus Kennedy.

A male mallee military dragon (Ctenophorus fordi). Gluepot Reserve, South Australia, 2011. Photo by Tobias Hayashi.

A female mallee military dragon (Ctenophorus fordi). Gluepot Reserve, South Australia, 2011. Photo by Tobias Hayashi.

While looking for claypan dragons in southwest Western Australia, we came across a few spotted military dragons (Ctenophorus maculatus). This species is very similar to the other two military dragons, but it doesn't live in spinifex. Instead, they seemed to like areas that were open, but with short woody plants all over the place. They also didn't seem to be nearly as common as the other two, but maybe that was just because we weren't looking.

A female spotted military dragon (Ctenophorus maculatus). near Lake Cronin, Western Australia, 2013. Photo by Angus Kennedy.

Those are the three members of this group of vegetation-dwellers we came across during my fieldwork. Of the other four, only one is widespread and well known. The lozenge-marked dragon (Ctenophorus scutulatus) is the largest member of this group and is found over a large area of Western Australia just north of Perth. The other three species, the long-tailed military dragon (Ctenophorus femoralis), the rufus military dragon (Ctenophorus rubens), and McKenzie's dragon (Ctenophorus mckenziei) are all very poorly known critters. They're all restricted to small areas of remote habitat: the former two halfway up the coast of Western Australia, and the latter on the Nullabor plain. McKenzie's dragon also has the unfortunate distinction of being endangered.

Burrowing Dragons, group 2

In his book "Biology and Evolution of Australian Lizards", Allen Greer sorted the lizards I study, the genus Ctenophorus, into different groups based on where they live: those that live in burrows, those that live in rock crevices, and those that live in vegetation. Looking at the phylogenetic relatedness of the Ctenophorus dragons, there are two groups of each: two groups of burrowers, two groups of rock-dwellers, and two groups of vegetation-dwellers. I thought it'd be fun to put up pictures and descriptions of all these groups, since I have a large pile of pictures from my fieldwork. These are my posts so far: Rock-dwellers #1, Rock-dwellers #2  Burrowers #1, Vegetation-dwellers #1.

This second group of burrowers is awesome! Two of the dragons in this group are absolutely spectacular in terms of colour and behaviour, and the other two are mysterious beasts that are little known, rarely seen dwellers of Australia's most unforgiving habitat: salt lakes. They all dig cute little burrows and duck into them if they feel threatened.

They are among my absolute favourite things to chase, which is great because this is the only group that we went chasing all three seasons of fieldwork. We started off chasing painted dragons at the very beginning of my very first field season and we ended my very last field season chasing claypan dragons over two years later.

Painted Dragon (Ctenophorus pictus)

Painted dragons live up to their name. They are gorgeous! The males have brightly coloured heads that are blue, red, orange or yellow, along with a beautifully coloured and intricately patterned backs. The females are also quite intricately patterned, but they are not brightly coloured like the males. This is one of the first species we chased back in September 2011, and we had so many problems! It was a huge challenge just to catch one, and often it took three of us all surrounding the same tree to get a lizard. The trouble we were having had me worried for the rest of my fieldwork. Would we be able to catch enough lizards for my project to work?

We needn't have worried. The problem wasn't that that painted dragons are hard to catch, it was that we sucked at catching painted dragons! By 2012 we'd had a lot of practice, and we were hoovering them up. It turns out painted dragons are among the easiest dragons to catch, owing partially to their habit of sitting conspicuously on the tops of bushes. 

Male painted dragon, Ctenophorus pictus. Turlga Station, South Australia, 2012. Photo by Angus Kennedy.

Female painted dragon, Ctenophorus pictus. Turlga Station, South Australia, 2012. Photo by Angus Kennedy.

Claypan Dragon (Ctenophorus salinarum)

Who knows why they call these things claypan dragons. They don't live in claypans, which are basically dried-up ponds. They live in salt pans, which are basically dried up salt lakes. When it's wet in the Australian scrub, like it was when we were looking for these guys in October 2013, they live on the salt crust around the edges of filled salt lakes. It's a very narrow strip of land, and makes their suitable habitat almost two-dimensional. 

Claypan dragons have a reputation for being hard to find. There are few records of them in the Atlas of Living Australia and almost nothing about them in the published literature. Maybe they have this reputation because people keep looking for them in claypans, because we had no troubles! Before my fieldwork I was worried that we'd have a lot of problems finding these guys. As it turns out, we came across them completely by accident while visiting a tourist spot before even starting our search in earnest! We did have trouble finding lakes where they lived. We visited a lot of lakes where we'd walk all the way around the lake and not find a sign of the dragons. However, when we visited a lake and found one, we'd find another, and then another, etc. They live in impressively high densities where they do live, it just seems that they're picky about which lakes they live around. Someone should use science to figure out why!

Male claypan dragon, Ctenophorus salinarum. Varley, Western Australia, 2013. Photo by Angus Kennedy.

Female claypan dragon, Ctenophorus salinarum. Varley, Western Australia, 2013. Photo by Angus Kennedy.

Lake Disappointment Dragon (Ctenophorus nguyarna)

These critters live in the middle of absolute nowhere. They was discovered by accident in 1996 when a vehicle full of scientists became stuck in the mud on the edge of Lake Disappointment. Lake Disappointment is so named because if you are wandering, lost and dehydrated, in the desert and see Lake Disappointment in the distance, you will think that you're saved. You will be wrong, disappointed, and more likely than not you will shortly be dead. Lake Disappointment is a salt lake and no one's saviour, except perhaps for the Lake Disappointment dragon's. Due to their remoteness we did not go after this dragon during my fieldwork.

Bicycle Dragon (Ctenophorus cristatus)

It's not completely clear that the bicycle lizard belongs in this group. When Greer wrote his book back in the 1980's, he included them in the burrowers, but Melville (2001) places them among the vegetation dwellers. Since the vegetation dwellers don't have a burrow or crevice close by for security, they tend to be long-legged, dainty things that are very good at taking off at high speed. The bicycle dragon certainly fits this bill. They're called bicycle dragons because, when scared, they lift up into a T-rex position, pin their front legs against their chest and rotate their back legs as if they're peddling a bike. This gets them far, fast.

But back to the burrowing. According to the most recent phylogeny (Chen et al., 2012), bicycle dragons are part of this group of burrowers. However, Chen and her colleagues can't be completely sure of this relationship, according to their statistics all they can say is that the bicycle lizard is probably part of this group (thanks to JP for clarifying the meaning of phylogenetic probabilities). Of the four phylogenies published on Ctenophorus over the past fifteen years, none have placed the bicycle dragon in the same place. So the genetics, at the moment, are inconclusive.

What about practically? Do bicycle dragons actually dig burrows? Greer, in his 1989 book "Biology and Evolution of Australian Lizards", says yes. However, others since then have said no, mostly citing personal comments and observations (eg. Meville et al., 2001; Thompson & Withers, 2005). During my fieldwork, we probably caught about forty bicycle lizards, and had another twenty or so escape on us. Not a huge sample size, but not small either. The vast majority took off as I described above. One individual ducked into a burrow. Compare this to the other burrowers we caught: the painted dragon, claypan dragon, and central netted dragon. The painted dragon would duck into a burrow probably around 80% of the time, the rest of the time taking off running or ducking under a bush. The claypan and netted dragons, which live in more exposed habitats than the painted dragon, ducked into burrows close to 100% of the time. So are bicycle dragons burrowers? Maybe they're facultative borrowers, building burrows when it suits them. This is another question that science is a very useful tool for answering.

Male bicycle dragon, Ctenophorus cristatus. Lake Hurlestone Conservation Reserve, Western Australia, 2013. Photo by Angus Kennedy.

Female bicycle dragon, Ctenophorus cristatus. Lake Hurlestone Conservation Reserve, Western Australia, 2013. Photo by Angus Kennedy.

References

Chen, I, Stuart-Fox, D., Hugall, A.F., and Symonds, M.R.E. 2012. Sexual selection and the evolution of complex colour patterns in dragon lizards. Evolution, 66-11:3605-3614.

Greer, A.E. 1989. The biology and evolution of Australian lizards. New South Wales: Surrey Beatty and Sons.

Melville, J, Schulte II, JA, and Larson, A. 2001. A molecular phylogenetic study of ecological diversification in the Australian agamid genus Ctenophorus. Mol Dev Evol., 291:339-353.

Thompson, GG and Withers, PC. 2005. The relationship between size-free body shape and choice of retreat for Western Australian Ctenophorus (Agamidae) dragon lizards. Amphibia-Reptilia, 26:65-72