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The modern world of wildlife photography is saturated with thousands of talented photographers producing a huge number of incredible photographs. As a result it is difficult to produce original shots without really pushing the boundaries and striving for new perspectives. Often, this means putting the camera into places that may at first seem impossible.
With this in mind, we were driven to embark upon an ambitious project to photograph African wildlife from a new perspective, one that would involve getting close to potentially dangerous animals and photographing them with a wide-angle lens. Traditionally, this has been achieved using camera traps – stationary cameras triggered when an animal breaks an invisible infra-red beem. The problem with this method is that it requires a great deal of time, patience and luck. We therefore decided to invent something a little more proactive! And so BeetleCam was conceived; a DSLR camera mounted on top of a four-wheel drive remote control buggy.
I booked a trip to Tanzania and set about designing, building and testing BeetleCam. The first step was to get up to speed on the necessary robotics and electronics that would be required to build such a vehicle from scratch. Having conducted some research, I sourced components from around the world. Construction then began in earnest with sawing, soldering, sewing and super gluing taking place around the clock in my garage.
BeetleCam's primary challenge would be getting over the uneven African terrain with a heavy payload of camera, lens and flashes. I therefore ordered the most powerful motors I could find and large off-road tires. BeetleCam had to be able to operate for long periods without being charged, so I stuffed the vehicle with the biggest batteries I could squeeze in.
I constructed a split ETTL off-camera flash cord that would allow the camera to control the output of two flashes depending on the light conditions (this would be important for filling in the shadows cast by the bright African sun).
After much deliberation, I figured out how to get the camera (my trusty Canon EOS 400D) to interface with the same controller used to drive the buggy. The finishing touches were to camouflage BeetleCam and seal the camera gear and internal mechanisms to protect them from the dusty African environment.
The prototype was finished with a month to spare but proved to be catastrophically unstable! An emergency redesign was undertaken to lower the centre of gravity and, a few days before our departure, BeetleCam was ready to be let loose in the wild!
Our destinations in Tanzania were Ruaha and Katavi National Parks, both in the South West of the country.
We thought that Elephants would be an easy subject for BeetleCam's first outing. We were wrong… we quickly learned that Elephants are wary of unfamiliar objects and due to their highly sensitive hearing, they are almost impossible to sneak up on! Over the course of the trip, we learnt that the best way to photograph an elephant was to position the camera well in front of it and then let the animal approach in its own time. With this technique we enjoyed great success and managed to get some incredible photos of these colossal creatures.
After obtaining our first photographs of Elephants we were buoyed with optimism and decided to make our second subjects Lions. In hindsight this was a foolish idea; BeetleCam was promptly mauled, and carried off into the bush. A long recovery mission ensued and we were extremely lucky to retrieve an intact memory card from the mangled Canon 400D body. On downloading the images, we were delighted to find that BeetleCam had performed its duty admirably, and we got a great series of images from the encounter.
Remarkably, although the 400D sustained irreparable damage, the rest of BeetleCam proved very resilient and, with a few pieces of string and bits wood, we were able to patch it up. We replaced the 400D with our only other available camera, a Canon EOS 1D MK III. This increased the stakes massively and obviously meant that lions were off the menu for the rest of the trip! Over the next few days we nervously drove our beloved 1D MK III in front of various unpredictable beasts and prayed it would come back intact!
To our surprise it was Africa's second most dangerous animal that proved to be the most cooperative subject, the African Buffalo. Adult males that are too old to compete for females collect together and form bachelor herds. Despite their reputation for being bad tempered and aggressive, these old brutes were totally unconcerned by the small robot. Some of the buffalo even showed mild curiosity and would amble up slowly to investigate.
Upon returning to the UK, we were thrilled with the photographs that we had managed to take during our two-weeks spent in the field. We have already started work on BeetleCam Mark II and plan to return to Africa this summer to take more photographs. To receive news about BeetleCam's future escapades, and to be the first to see the resulting photographs, please subscribe to our free email newsletter.
Update: Read the next instalment of this project here: BeetleCam vs the Lions of the Masai Mara
78 Comments
Lyza said: April 19, 2010 at 7:36 am
Wonderful work!!!
rushh said: April 19, 2010 at 7:38 am
woww…. nice work here.. cool gadget u've made… i like large picture @ middle…
Ruben Vicente said: April 19, 2010 at 7:55 am
Genius work! Maybe it's better to get some sort of metal case around the camera…lion proof =p
Good luck!
anonymous said: April 19, 2010 at 8:15 am
WOW! I want to do this one day..
Morkel Erasmus said: April 19, 2010 at 8:17 am
Wonderful shots of Africa's legends…thanks for sharing this, I know Hans Rack of Namibia has a similar type of gizmo with which he operates around the Etosha waterholes.
Shirley said: April 19, 2010 at 10:17 am
With great patience and marvellous enthusiasm
you have given us all some terrifc animal shots.
What about some African Sunsets next time you are over there ??
Thank you for sharing.
Sasa Huzjak said: April 19, 2010 at 11:35 am
Great idea, very cool photos, wish you even more luck on your next BeetleCam adventure!
PS: Sorry about the 400D, good news about 1D MK III 🙂
Giles Breton said: April 19, 2010 at 11:50 am
this is fantastic work, and you're absolutely right about the need for a new fresh perspective on wildlife photography. Have you contacted the guys who set up Spy in the Jungle using trunk cams and tusk cams. Their cameras were tiger proofed as much as you can
MBUGUA said: April 19, 2010 at 12:17 pm
Guys, this is great.
I have seen your pics and all can say is that they are very nice. the idea behind it was great.
i am also a photojournalist based in kenya.
i am hoping to get a better camera since what i have is a D50 nikon.
thanks guys.
post more of that we need to learn more.
mbugua kibera
Wanda Krack said: April 19, 2010 at 12:32 pm
Wow, what an adventure! Hats off to both of you for your work, enthusiasm, and new perspective! Wishing you luck with your next one!
Emilie said: April 19, 2010 at 2:49 pm
Great invention.
Livia said: April 19, 2010 at 3:56 pm
Amazing guys!
You are really doing a great job! Keep on the track, you can go even futher on this project! Becareful with the wild!
Looking forward for the next pics!
Sebastian Kennerknecht said: April 19, 2010 at 4:25 pm
Great stuff as usual. In many of the pictures it looks like you have small DOF considering the wide-angle lens. How where you focusing the camera?
Eddy Wee said: April 19, 2010 at 4:27 pm
Wow, great work guys!
Anna said: April 19, 2010 at 4:42 pm
What a great adventure you guys had. Awesome pictures great idea.
Jerad Hill said: April 19, 2010 at 6:11 pm
Awesome. I mounted my 5D MKII to my old Traxxas Emax. Maybe what you need to do is rig up a pepper spray to one of the servos that you can fire when it needs to fight back.
Marie Dunphy Harding said: April 19, 2010 at 7:27 pm
I thoroughly enjoyed 'Will & Matts' excellent adventure' story. Can't wait for BeetleCam2.
By the way, do bachelor herds only come in buffalo?
All the best with your future endeavours.
Ravindra Joisa said: April 20, 2010 at 8:34 am
wow.. Too good… great work.. even i'm into photography .. but you guys have tried something extraordinary.. .:)
Vijay Padiyar said: April 20, 2010 at 10:08 am
Whoa!!! Awesome work, folks! Maybe for BeetleCam 2.0, you could try elevating the camera and flashes with a camera rod mounted on the buggy (and a retractable one at that). That would help you take pictures from a more conventional height (similar to a sitting person), which would look even more natural.
No one would be able to figure out that the pics were taken remotely! Also, you would be able to eliminate the twigs from the pics by raising the camera above them. And by making it retractable, you would be able to lower the camera when not taking pictures.
Vijay Padiyar said: April 20, 2010 at 10:14 am
Or perhaps work on a CopterCam for next time! A camera and flashes mounted on a radio-controlled helicopter. That would allow you to take close snaps while still keeping your camera out of range of the animal!


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Diabetes Channel
Related Channels
Actos and Weight Gain
Related MultimediaClick on an icon to view.
One very important point to keep in mind is that Actos may worsen congestive heart failure or, in some cases, even cause heart failure. One of the main symptoms of heart failure is unexplained weight gain. In fact, people with symptoms of congestive heart failure can often have rapid weight gain (three to five pounds) over several days.
This weight gain is often combined with other symptoms, such as a cough and/or swelling in the ankles, feet, or legs. Make sure to contact your healthcare provider if you notice unexplained weight gain or swelling (see Symptoms of Congestive Heart Failure for more information on heart failure symptoms).
If you are noticing gradual, unexplained weight gain while taking Actos, there are some things that you can do.
Some suggestions include:
- Eating a heart-healthy diabetic diet. This diet should include:
- Lots of fruits, vegetables, grains, and fat-free or low-fat milk and milk products
- Lean meats, poultry, fish, beans, eggs, and nuts
- Limited amounts of saturated fats, trans fats, cholesterol, sodium (salt), and added sugars.
- Getting regular physical activity for at least 30 minutes a day on most days of the week.
- Limiting your intake of alcohol.
Also, if you are noticing gradual weight gain with Actos, talk to your healthcare provider. He or she can give you other suggestions for dealing with any weight gain. He or she may also look for other causes of weight gain, such as certain types of medical conditions. If the weight gain continues, he or she may also recommend other lifestyle changes or a possible switch to another diabetes medication.
(Click BMI Calculator to see if your weight is within a healthy range.)
Actos Drug Info
Written by/reviewed by: Kristi Monson, PharmD; Arthur Schoenstadt, MD
Last reviewed by: Kristi Monson, PharmD

subset: commoncrawl


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RightNow have a great looking QR Scavenger Hunt at SXSW. Players start by finding RightNow runners (black t-shirts that say #theGame_SXSW on the front and with a QR Code on the back (images below) and then prove it by submitting a photo via Twitpic. To get subsequent QR codes you will need to crack runner locations, activities and identities. Prizes include a Nikon D7000 kit and other goodies. The full game rules and the live Twitter image feed are online here . Start with a tweet "#theGame_SXSW I want to join!" and RightNow will tweet you back immediately with the first set of clues.
Tip: Sunday 12:30-1:30 at the Hyatt 🙂


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Protecting Brainpower During Radiation
July 8, 2011 | Leigh MacMillan
Cranial irradiation to treat brain cancers often has long-term negative neurocognitive effects, including lowered IQ, learning difficulties and memory loss, especially in the pediatric population.
Fen Xia, M.D., Ph.D., and colleagues have reported that these effects may be due to radiation-induced damage to the hippocampus, the brain region responsible for learning and memory. They previously demonstrated that inhibition of the protein GSK3β (glycogen synthase kinase 3β) reduces radiation-induced hippocampal neuron cell death and protects neurocognitive function in mice treated with cranial irradiation.
The team has now discovered that inhibition of GSK3β – with specific inhibitors or using genetic manipulations – accelerates the repair of radiation-induced DNA damage (double strand breaks) in normal hippocampal neurons, but not in malignant glioma cells. The findings, reported in the May issue of Neuro-Oncology, link GSK3β to DNA repair pathways and suggest novel targets for the development of neuroprotective drugs to use during whole brain irradiation. In particular, the research suggests that GSK3β inhibitors may have neuroprotective effects.


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Beyonce fans were super excited to hear that she had a new single coming out with Naughty Boy, and their excitement will definitely be compounded by the fact that the video for Runnin' (Lose It All) is absolutely incredible.
The video was created entirely underwater, and features world champion free diver Guillaume Nery in the leading role. The song begins as a piano ballad, then explodes into a high-energy song powered by Beyonce's vocals. Nery and his costar are depicted running down the ocean floor to unite with one another. It's a powerful effect, and after you see the video, you're going to wonder how they filmed it.
Watch Nery dive into the deepest hole in the world, Dean's Blue Hole.
Watch Nery dive into the deepest hole in the world, Dean's Blue Hole.


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Matt Morrison
Armwood High Class of 1992
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Visit us to learn why parents and children love Apple Montessori!
Choosing the right preschool program for your child is one of the most important decisions you'll ever make. Family owned Apple Montessori has been a trusted leader in early childhood education for over 45 years. We make the most of the critical early learning years from birth to age 6 by fostering each child's fullest potential, at his or her own pace.
The Montessori approach is unique in that it builds on the way children naturally learn, and Apple's Montessori Plus approach distinguishes Apple from other Montessori schools. Our programs are research-proven. Our schools are safety-certified. Our educators love what they do. Visit us to see our whole child approach in action, meet our nurturing educators, and explore the many ways your child will thrive at Apple.
Interested in summer camp? Learn about the Hoboken Summer Camp Top Reasons Sinatra
- Age Group:6 weeks to 6 years
- Hours:7:00 am – 7:00 pm (school day is 8:30 am – 3:30 pm)
- Address:220 Sinatra Dr
Hoboken, NJ 07030Phone:
Hoboken - Sinatra Drive Overview
We offer two schools in Hoboken for your convenience with infant, toddler, and preschool/kindergarten programs as well as before and after school daycare. The Sinatra Drive school, located on the south side of town is close to the PATH train, ferry and light rail. Expansive views of the Hudson and the magnificent Manhattan skyline beyond can be seen from the classrooms. Our nurturing educators will inspire a love of learning for your child in a warm, supportive environment. Our indoor playground is one of the highly regarded physical features of the school. The safety and security of your child is paramount. The school is equipped with a state-of-the-art security entry system with fingerprint technology and a surveillance system with live, remote access for parents. One of the ways we ensure the safety and well-being of your child is by using a germ sanitizing machine which kills 99.9% of germs and viruses.
We welcome you to call or visit to tell us about your child and learn more about our Sinatra Drive or Maxwell Lane Hoboken schools.
Google & Facebook Reviews
Hear what our Hoboken families love about Apple
Learn about our partnership with SEC, a company owned and operated by former US Secret Service Agents, to ensure the highest level of safety and security possible.


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Introducing CodeSafe
MiSAFE offers CodeSafe as a simple way for any business to deliver and capture valuable information. Capable to work offline and online, with unlimited users across all subscription levels.
Record inspections, tasks, incidents etc. Employees and contractors can even attend role specific training in the field via their smart-device rather than spending countless hours in a training room.
Worker Endorsed Risk Compliance SoftwareBridge the gap between the office and the field using CodeSafe's risk compliance software app. Whether you have non-compliance issues, language and literacy barriers, a lack of visibility down the supply chain, rework losses or other challenges, MiSAFE has the solution for you.
Smart Induction SoftwareAre you looking for a smarter way to do workplace or multi-site induction training? Ones that save you time and money, and eliminate paperwork? Ones that are less frustrating for your staff and contractors? Integrating CodeSafe's Smart Forms with your safety induction processes will provide the simplest and most assuring way for everyone to comply with their legal obligations, and get on with the job.
Easy to Use
Setup your risk compliance software in minutes with your own customised content.
Offline Capable
Users don't need to be connected to the internet to use CodeSafe's risk compliance software.
Smart Device Enabled
CodeSafe risk compliance software works on any smart device.
Improve Worker Engagement
Monitor worker compliance and improve engagement with your risk management systems.
Download the CodeSafe App for Free
Download the CodeSafe App for free today and have a play around to see what it can do. Give us a call on 07 5641 2101 or send us an email email@example.com if you wish to know more about how to use CodeSafe App.
Hazard & Incident Reporting
Report hazards and incidents anywhere in the field. Online and offline. Geolocation tracking..
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Record plant and equipment inspections (pre-starts) using QR codes or drop-down menu selections. Simply scan the QR code, or choose the item from the drop down list, and complete/submit the form. Done!
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Workers can attend their induction training before turning up on site. Reduce the time spent on inductions and training, by allowing the workers to attend at their own pace. Offline or online.
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Record journey management plans, and remote worker actions. Scheduled task management, and geolocation logging.
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Conduct customised Audits & Inspections offline or online.
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Allow contractors to upload licenses and certificates (i.e. VOC information) prior to turning up on site.
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Upload valuable documents such as policies and procedures, allowing all users to access the company's core Management Systems Documents.
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Don't just take it from us, let CodeSafe customers do the talking!
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Can the software be used remotely where there is limited internet access?
Yes, CodeSafe is fully functional in remote locations. Simply use the app as usual and it will synchronize with your system when the user gains internet access again.
Does the software come with content?
The software is feature rich, but requires the user to upload their specific content and configure the system that suits their business best. If you don't have content or don't know where to start, simply contact us on 07 5641 2101 or firstname.lastname@example.com and we can provide a quote to develop the content for your business.
How many users does each subscription allow?
The beauty of CodeSafe is that you can have an unlimited amount of users at no extra cost. The sky's the limit.


text: Hello, How would this do as a mass RFP :) http://sal.kachinatech.com/ SAL (Scientific Applications on Linux) is a collection of information and links to software that will be of interest to scientists and engineers. The broad coverage of Linux applications will also benefit the whole Linux/Unix community. There are currently 3,070 entries in SAL Most of the software is not GPL, but about a half is and can be packaged. I never thougt that Linux has THAT many scientific progs that are not in Debian jet. :) -- Best regards, Aigars Mahinovs mailto:firstname.lastname@example.com #--------------------------------------------------------------# | .''`. | | : :' : Debian GNU/Linux & LAKA | | `. `' http://www.debian.org http://www.laka.lv | | `- | #--------------------------------------------------------------#


text: November 9, 2012 at 1:51 PM 220 marijuana cases dismissed in King, Pierce counties Posted by Jonathan Martin UPDATE 3:03 p.m. King and Pierce County prosecutors are dismissing more than 220 misdemeanor marijuana cases in response to Tuesday's vote to decriminalize small amounts of pot. In King County, 175 cases are being dismissed involving people 21 and older and possession of one ounce or less. I-502 makes one ounce of marijuana legal on Dec…. More Comments | More in homepage, Marijuana, Politics Northwest | Topics: dan satterberg, initiative 502, King county prosecuting attorney


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ABF The Soldiers' Charity
ABF The Soldiers' Charity are for soldiers, for life; giving support to serving and retired soldiers and their immediate families.
They risk so much for us, but for them, it's "just doing their job". In return, The Soldiers' Charity give them down-to-earth help and friendship, with no hassle or bureaucracy.
We help in ways that make a big practical difference to individuals. For example a level access shower for an amputee from a recent conflict or care home fees for an elderly veteran.
That's what we've been here to offer since 1944 and will be here for as long as there is a need.


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I've always dreamed that one day I will have my perfect kitchen. One with white/off white painted cabinets, light colored counter tops, an island painted in a fun color, wood floors, with splashes of different colors all over.
I have already started my colorful accessory collection. I received this awesome 'lemongrass' green kettle for Christmas:
And this is a close-up of the rug I got from Target the other day. It has the green from my accessories, plus the aqua/turquoise of the antique blue glass Ball jars I love so much.
So when I came across these gorgeous images of this kitchen, it made me wish that it was mine. I would sit in there all day just admiring my kitchen.
I love the little pennant garlands hanging on the windows - such a fun alternative to traditional window treatments. Pillows make the kitchen cozier, it's usually all hard surfaces. The white food floor is beautiful, though I think I would stick with natural wood tones so it didn't get too white.
That kitchen reminded me of Nicole's kitchen...
Love the turquoise island. Love the exposed brick, and the hidden spice shelves. Love the chalkboard on the pantry doors. I could settle right into that kitchen too!
This kitchen just popped up on Design*Sponge... Love it too!
One day...


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One day when I was doing research on baby food, I came across a simple baby biscuit recipe that stuck in my brain. Sometimes it's handy to have easy-to-hold finger food around. I've found that sticks of banana, cooked carrots, or avocado are easy to grab, but fall to pieces when baby squishes them or bites into them. I have yet to find store-bought baby cookies that don't include some form of sugar, and since when do babies need sugar? (Well, for that matter, since when do any of us need sugar? Too bad we're all addicted…)
I made the cookies for Halia last week and just had to share the recipe because it is so damned simple. There are two ingredients: ripe bananas and oats.
I bet you already have those in the house, don't you?
Oats are great for babies (unless you're avoiding all source of gluten, that is) and did you know that they're always 100% whole grain? The fast-cooking ones are just rolled out thin, but none of the grain is removed.
For the cookies, all you have to do is stick a couple of handfuls of oats (I used quick-cooking ones — not instant, mind you) into a food processor and whirl them until they're fine. Then mash up your banana and add enough oats to make a sticky dough.
Spread out the batter onto a cookie sheet (I put mine on parchment paper) in whatever shape you like (I made mine into simple rectangles). Bake at 350 °F for 10-15 minutes until golden. The bars will be chewy when cool. Try one! The natural sweetness of the banana is so yummy.
Halia was so pleased to be able to hold and munch on her own food. Sadly, I normally don't give her a lot of freedom with her food because I'm terrified of the carbs getting everywhere and contaminating something Jade will touch. I bet no baby in the history of the world has ever stayed so clean while being fed. So getting to have her own cookie was a huge privilege for her. Just look how much she enjoyed it:
I also tried freezing the bars because I figured they'd make good teething biscuits that way. Halia enjoyed a frozen one on our way to Fox Lake yesterday.
Here's the recipe in it's entirety, copied directly from this page on the Homemade Baby Food Recipes site:
Baby Oatmeal and Banana Bars
This easy recipe, sent in by a visitor to our site, produces chewy oatmeal bars for babies coping well with finger foods.
1 ripe banana
handful of oats
- Preheat the oven to 350 °F (180 °C).
- Mash the banana thoroughly.
- Grind the oats in a food processor.
- Combine the banana with the enough ground oats to form a sticky dough.
- Divide the mixture into small pieces and press it into the shapes of your choice (we make simple bars).
- Bake for around 10-15 minutes until golden brown in colour – they become harder the longer they are cooked, so keep the baking time as brief as possible if you wish to retain a chewy texture.


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You are beginning treatment with ribavirin pills, and peginterferon -- which must be injected once a week.
Be sure to store your peginterferon in the refrigerator, and out of the reach of children. Also make sure that your ribavirin pills are kept in a place where children can't find them and take them.
It's important to give your injection exactly the way your healthcare provider shows you. This will ensure that you get the right amount of peginterferon. If you forget to take an injection, be sure to call your healthcare provider right away for further instructions.
By following your treatment plan, and taking your medicine as directed, you will be more likely to have a successful treatment. If you have any questions about your medications, or how you should take them, be sure to talk with your healthcare provider. Even if your question seems simple, remember, any question you have is an important one.


text: Here's a reminder now that's it is spring that you can still look like the man at any event if you forego the tie and choose, instead, a nice linen pocket-square that has tipping in a color that's the same tone and, while not exactly matching, coordinates with the rest of your look. You might even consider one from Ermenegildo Zegna, like Chris Pine did at last night's CinemaCon gala. But, no matter which brand you choose, it's a trick worth trying the next time you need to wear a suit at night.


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Rapamune Solution (Sirolimus)
Sorry, we currently do not carry this product.
Sorry, we currently do not carry this product.
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Multiple Dates
Sketching the interiors of Assembly Rooms Edinburgh - 3 hour drawing tour
Capture the beauty and detail of this unique and iconic Edinburgh venue in it's 230th year with this one day only experience.
Explore the timeless elegance and beauty, the enormous crystal chandeliers, gleaming gold leaf and giant gilt mirrors with a sketch book in hand, using ink and watercolour to bring your visit to life.
With helpful instruction from Mark Kirkham, a local artist known as the Edinburgh Sketcher, you will quickly pick up sketching skills while increasing confidence in your own artistic ability as you tour the various rooms of this Georgian venue.
Once you've filled your pages with beautiful ink drawings, we'll end our tour with tea or coffee and time to add watercolour to your sketches and compare our doodles.
No previous experience is required for this relaxed and informal drawing tour and all materials are provided.


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Trypanosomatid parasites are protozoa that are agents of human diseases, such as Chagas disease, African trypanosomiasis and leishmaniasis. Many plants also harbour trypanosomatids, which primarily reside in the xylem, phloem tubes (França 1914, Holmes 1925, Harvey & Lee 1943, Dollet 2001), fruits and/or seeds (Jankevicius et al. 1987, 1989, 1993, Conchon et al. 1989, Sánchez-Moreno et al. 1995) of infected plants. The presence of the pathogens in the fruits or seeds may yield local pathological symptoms, but never systemic symptoms. Trypanosomatids are agents of plant diseases, such as Hartrot of the coconut (Parthasarathy et al. 1976), Marchitez of oil palm (Dollet et al. 1977), wilt of the ornamental ginger Alpinia purpurata (Gargani et al. 1992) and coffee tree phloem necrosis. The commercial importance of these crops emphasises the need for more research on these devastating diseases. Such diseases are likely transmitted through the bite of phytophagous insects (Camargo et al. 1990, Camargo & Wallace 1994, Redman et al. 1995). Tomatoes can be infected by Phytomonas serpens without apparent pathological effect, but the fruits present yellow spots that are consequently of low commercial value.
Plants produce compounds that provide chemical defences against pests, pathogens and invasion by neighbouring plants. These compounds include sterols and triterpenes that are formed by cyclisation of 2,3-oxidosqualene. Sterols and triterpenes accumulate as glycoside conjugates and include glycoalkaloids, which are commonly referred to as saponins. Saponins possess antimicrobial and antifungal activities that act as plant defences. Saponins act by increasing membrane permeability because of their ability to form complexes with cholesterol. Tomato plants (Solanum lycopersicum L.) produce tomatine, which is a tetrasaccharide linked to the 3-OH group of the aglycone tomatidine (Fig. 1). Immature green tomatoes contain up to 500 mg of tomatine/kg of fresh fruit weight (Friedman 2002). The compound is largely degraded as the tomato ripens until it reaches levels in mature red tomatoes of 5 mg/kg of fresh fruit weight. Glycoalkaloids from potatoes (α-chaconine, α-solanine), eggplant (α-solamargine, solasonine) and tomatoes (tomatine) are growth inhibitors of Trypanosoma cruzi, strain EP, in liver infusion tryptose medium (Chataing et al. 1998).
Fig. 1: structure of tomatine (A) and tomatidine (B).
The major sterols of the trypanosomatids are C28-ergostane (24-methyl group) and C29-stigmastane (24-ethyl) types, in which an extra methyl or ethyl group is added to carbon-24 of the sterol side chain. In contrast, animals synthesise C27 cholestane-based members of the steroid family. The alkylations of the sterol side chain are performed by two distinct 24-sterol methyltransferases (SMT), SMT type 1 (SMT1) and type 2 (SMT2). Various azasterols with a nitrogen substitution in the side chain inhibit C-24 transmethylation reactions involving S-adenosyl methionine as the methyl donor and a ∆24(25)-sterol or ∆24(24`)-sterol substrate. 24-SMT is present in fungi, plants and trypanosomatids, but it is absent in mammals, which makes this enzyme an attractive target for chemotherapeutic agents against pathogens (Haughan et al. 1995, Rodrigues et al. 2002, Roberts et al. 2003, de Souza & Rodrigues 2009).
Here, we investigated the toxicity of tomatine and its aglycone, tomatidine, against P. serpens. Both compounds inhibit the growth of Phytomonas, but they utilise different mechanisms. The glycoalkaloid tomatine disrupts the plasma membrane, which kills the parasite and tomatidine induces notable morphological changes in the parasite, apparently due to the inhibition of alkylated sterols synthesis.
Materials - Tomatine and tomatidine were acquired from Sigma Chemical Co Stock solutions of both alkaloids were prepared in dimethyl sulfoxide (DMSO). DMSO alone had no effect on cell proliferation. BODIPY FL C16 was obtained from Molecular Probes (USA).
Cell cultures - P. serpens (CT.IOC 189) were grown at 27ºC in a medium containing 20 g/L sucrose, 20 g/L KCl, 3 g/L peptone and 1 mg/L folic acid, supplemented with 10 mg/L haemin and 10% (v/v) foetal bovine serum.
Effects on growth - Cultures were initiated at a cell density of 1 × 106 cells/mL and drug was added 24 h later, when the number of parasites in the cultures reached approximately 5 × 106 cells/mL. Cell densities were evaluated daily in a Neubauer chamber after 96 h of growth. The half maximum inhibitory concentration (IC50) values were calculated using GraphPad 5.0 software.
Cell permeabilisation assay - Promastigotes of P. serpens were harvested after two days of growth, washed in phosphate buffered saline (PBS), pH 7.2 and counted in a Neubauer chamber. Cells were incubated with tomatine, tomatidine or digitonin at the concentrations indicated in the Figure legends for 10 min. Cell suspensions were centrifuged at 12,000 rpm in an Eppendorf centrifuge for 5 min. The amount of pyruvate kinase (PK) (cytosolic enzyme) in solution was measured in 0.5-mL aliquots of supernatant using a coupled assay in media containing 50 mM Tris HCl pH 8.0, 5 mM MgCl2, 1 mM ADP, 1 mM phosphoenolpyruvate, 100 mM KCl, 0.25 mM β-NADH and 0.1 U lactate dehydrogenase in final volume of 2 mL. Changes in absorbance at 340 nm were monitored at 2-min intervals to estimate the rate of β-NADH consumption.
BODIPY-palmitic acid labelling - Cells were grown for 24 h and 5 µM BODIPY FL C16 was added to 2-mL aliquots of culture, in which 50 µM tomatidine was or was not included. After 48 h, the top 1.5 mL portion of the culture was withdrawn and washed twice with PBS. Cells were fixed using 4% formaldehyde and observed under a fluorescence microscope (Axiovision; Carl Zeiss) using a 505 nm excitation filter and a 515 nm emission filter. The images were further processed using Adobe Photoshop CS2 (Adobe Systems Inc).
Transmission electron microscopy (TEM) - Growing parasites were treated with tomatidine after 24 h and cells were harvested 48 h later, washed with PBS and fixed using 2.5% glutaraldehyde grade I in 0.1 M cacodylate buffer for 2 h at room temperature (RT). Cells were washed twice with 0.1 M cacodylate buffer, pH 7.2 and post-fixed in the dark using 1% osmium tetroxide, 0.8% potassium ferrocyanide and 5 mM CaCl2 in cacodylate buffer for 30 min at RT. Cells were washed with cacodylate buffer, dehydrated in acetone solutions of increasing concentrations and embedded in epoxy resin (Polybeb 812). Ultrathin slices were obtained using a Reichert ultramicrotome, collected on 300-mesh cupper grades and contrasted with 5% uranile for 45 min and lead citrate for 5 min, according to the protocol of Reinolds. The samples were analysed using an electromicroscope Zeiss 900 and Jeol 1200, operating at 80 KV.
Sterol analyses - Parasite lipids were extracted using the method of Bligh and Dyer (1959). Briefly, Phytomonas were washed with PBS and 5 x 107 cells were intermittently agitated for 1 h with ~4 mL of a methanol:chloroform:water (2:1:0.8) mixture. The tubes were centrifuged at 2,000 g (15 min) and the supernatant, which contained the lipids, was separated. The pellets were extracted one more time and the supernatants were combined. A two-phase separation was forced by the addition of 1 mL water and 1 mL chloroform. Samples were agitated and the organic (bottom) phase was separated and dried under a dry N2 stream.
The residue was saponified using alcoholic KOH, extracted and dissolved in pure chloroform. Samples were treated with trimethylchlorosilane and analysed using gas chromatography with detection by mass spectrometry (GC-MS). A GC-MS Shimadzu gas chromatograph was used, model GP2010 Plus, with an RTX®-5 MS (5% phenyl 95% dimethylpolysiloxane) column from Restek® (30 m x 0.25 mm x 0.25 µm). The injector was maintained at 250ºC with a 1:1 split flow ratio. The column oven temperature was maintained at 170ºC for 5 min, then increased to 250ºC with a heating rate of 20ºC min-1 and finally to 280ºC, with a high rate of heating of 5ºC min-1, which was maintained for 20 min. Helium was used as a carrier gas, with a linear velocity of 41.9 cm s-1. Volumes of 1 µL were injected into the chromatograph for analysis.
The MSdetector was equipped with an electron (EI-70 eV) ionisation source and a quadrupole mass analyser, operated in SCAN mode (50-700 µm.a.). The interface was maintained at 230ºC and the ion source was maintained at 200ºC. The components were identified by comparing their mass spectra with spectra in the NIST05 library stored in the computer controlling the mass spectrometer and a critical analysis of fragmentation patterns. Retention indices were also used to confirm the identity of the peaks in chromatograms.
Statistical analysis - Data are expressed as means ± standard error from at least three independent experiments. Statistical significance was calculated using two-way ANOVA followed by Bonferroni's post-test. A difference was considered to be statistically significant when p ≤ 0.05. Analyses were performed using GraphPad 5.0 software.
Effects of tomatine and tomatidine on the growth rate of Phytomonas - Tomatine induced a dose-dependent inhibition of Phytomonas growth with an IC50 of 9.9 µM after 48 h of incubation. Fig. 2A shows that the presence of 50 µM tomatine in the culture media caused the death of all cells. Tomatidine also produced a dose-dependent inhibition of growth (IC50 = 14.2 µM). In contrast, 50 µM tomatidine stopped the growth of Phytomonas in culture (Fig. 2B), apparently without killing the cells. This behaviour could be a result of the inhibition of the cell cycle via blockade of nucleus or kinetoplast separation during the mitotic process. However, the results obtained by the labelling of DNA with 4'-6-diamidino-2-phenylindole excluded this hypothesis because no increase in the number of cells containing more than one nucleus or kinetoplast was observed (not shown). Additionally, the fluorescence intensity of these organelles was comparable in normal and treated parasites, which suggests that the amount of DNA per cell was not changed.
Fig. 2A, B: growth curve of Phytomonas serpens in the presence of tomatine and tomatidine. Promastigotes (1 × 106/mL) were grown in medium supplemented with 10% foetal bovine serum at 27ºC. After 24 h of growth, tomatine or tomatidine were added at the indicated concentrations. Each point represents the mean ± standard error of the means of three independent experiments. *: p < 0.05; **: p < 0.001 (2-way ANOVA with Bonferroni's post test). The half maximum inhibitory concentration values obtained after 48 h of treatment were 9.9 μM (for tomatine) and 14.2 μM (for tomatidine). Dimethyl sulfoxide alone had no effect on cell proliferation (data not shown).
Permeabilisation of the cell membrane - Some glycoalkaloids interact with sterols, such as cholesterol and ergosterol, to disrupt the membrane and cause a loss of cellular content (Morrissey & Osbourn 1999). The leaking of PK (which is a cytosolic enzyme) after treatment of Phytomonas with either tomatine or tomatidine was measured to test this hypothesis. Tomatine (10 µM) was very effective in permeabilising the cellular membrane (Fig. 3A). The amount of PK released was proportional to the amount of cells and the release was very similar to the extent of release induced by 200 µM digitonin. This result supports that tomatine kills cells via permeabilisation, which is similar to other glycoalkaloids.
Fig. 3: tomatine induced pyruvate kinase (PK) leakage from Phytomonas serpens. Cells were harvested after two days of growth and resuspended in phosphate buffered saline at the concentrations indicated below. The tomatine (A) or tomatidine (B) was added and after 10 min, the cells were spun down and PK was measured in 0.5 mL of the supernatant (see Materials and Methods). : control without alkaloids;
: 1 x 106 cell/mL treated with 10 μM tomatine; □: 3 x 106 cell/mL treated with 10 μM tomatine; : 3 x 106 cell/mL treated with 200 μM digitonin; ▲: 30 mU of purified PK; : 3 x 106 cell/mL treated with 10 μM tomatine and measured in the absence of lactate dehydrogenase; º : 1 x 106 cell/mL treated with 10 μM tomatidine; : 3 x 106 cell/mL treated with 100 μM tomatidine. Each set of curves was repeated three times and the Figure is representative of these experiments.
In contrast, tomatidine concentrations as high as 100 µM did not trigger any membrane leakage (Fig. 3B), which implies a different mechanism of cell poisoning.
Cell morphology alterations - Optical microscopy of Phytomonas treated with tomatidine and stained using Giemsa revealed several clear spots, like "holes", along the cell body (not shown). Treated cells observed under TEM revealed the presence of large rounded bodies, some with multiple membrane layers (Fig. 4B-D) and others with the appearance of lipid bodies (Fig. 4D-E, arrow heads). No significant changes in nuclear or kinetoplast morphologies were observed.
Fig. 4: ultrastructural alterations of Phytomonas serpens induced by tomatidine. A: ultrathin section of P. serpens without treatment which presents a normal ultrastructure of organelles such as kinetoplast (k) and nucleus (N); B-D: electron micrograph of P. serpens treated with 30 μM tomatidine for 48 h presenting many vacuoles (arrows), in some cases localised to the posterior region of the parasite; E: after treatment with 50 μM tomatidine for 48 h, it is possible to observe the presence lipid bodies (arrowheads). Bars = 1 μM (A-C, E) and 0.25 μM (D).
Fatty acid incorporation in Phytomonas - The microscopy data suggested the formation of vacuoles that contained a clear material with the appearance of lipids. Assays of fatty acids incorporation using the fluorescent analogue BODIPY were performed to confirm this hypothesis. Tomatidine-exposed cells presented a build-up of label lipids in rounded structures in the cytoplasm (Fig. 5). These spots had a similar size and distribution to the vacuoles observed in Fig. 4, which indicated that tomatidine may perturb lipid metabolism and membrane organisation.
Fig. 5: fatty acid incorporation in Phytomonas serpens vacuoles. P. serpens was grown for two days in the presence of 5 μM labelled BODIPY-palmitic acid (green) and in the absence (A) or presence (B) of 50 μM tomatidine as described in Materials and Methods, then harvested and resuspended in phosphate buffered saline for fluorescence microscopy. Right panels are images obtained by interferential contrast microscopy. The inserts represents typical cells in detail. Bars = 10 μM; inserts = 0.25 μM.
Analysis of lipid composition - Lipids extracted from Phytomonas were analysed using GC-MS (Table). The most abundant Phytomonas sterols were cholesterol (14.8 %), ∆7- ergosterol (15 %), ∆7-sitosterol (37.9 %) and 24 ethyllophenol (20 %). Phytomonas and other trypanosomatids do not have a complete pathway for cholesterol synthesis. Therefore, the presence of cholesterol must be a consequence of its uptake from the culture media.
The sterol pattern was deeply modified when the cells were grown for 48 h in the presence of tomatidine. The cells exhibited an intense decrease in 24-alkylatedsterols and a marked increase in desalkyl sterols, including zymosterol and cholesta-7,24-dien-3β-ol (Table). DMSO (vehicle) had no effect on sterol composition (not shown). The accumulation of zymosterol is consistent with an inhibition of 24-SMT. This enzyme is responsible for the incorporation of methyl groups at position 24 of sterol intermediates, which is an essential step for the production of ergosterol and other 24-alkylated sterols in protozoan parasites (Urbina et al. 1996, Nes 2000, de Souza & Rodrigues 2009).
The antifungal effect of azasteroids has been known for quite a long time, but the recognition of these agents as efficient chemotherapeutics against trypanosomatids was only recognised recently (Burbiel & Bracher 2003). Natural azasteroids are found in plants as part of the defence barrier against fungi and yeasts. Azasteroids are steroidal alkaloids that are hydroxylated in the three position, conjugated with glycosides and referred to as saponins. Tomatine is the most abundant saponin in the tomato plant and its antifungal properties are well documented (Sandrock & VanEtten 1998, Ito et al. 2007). However, the defensive properties of the aglycone tomatidine have been neglected because its concentration is nearly 100-fold lower than tomatine.
Some pathogenic fungi have acquired the ability to hydrolyse the glycoside portion of the glycoalkaloid, which produces compounds with no toxic characteristics. Removal of the terminal glucose of α-tomatine yields β-tomatine, which exhibits little or no toxicity against some fungi (Sandrock & VanEtten 1998). Removal of all four sugars yields the aglycone tomatidine, which maintains toxic properties against fungi, such as Neurospora crassa [50% effective dose (ED50) 51 µM], Aspergillus nidulans (ED50 81 µM), Cryphonectria parasitica (ED50 81 µM) and Stemphylium solani (ED50 300 µM) (Sandrock & VanEtten 1998). This study showed that tomatidine exhibited toxicity against P. serpens. However, tomatidine levels in the tomato fruit are too low to be an effective barrier against infection by this trypanosomatid. Tomatine was more effective against Phytomonas and it must be the more important defensin because of its elevated concentrations in green tomatoes. The tomatine content of tomatoes decreases during maturation. Roddick (1976) attempted to define the distribution in plant tissues and the dynamics of the synthesis and degradation of tomatine during ripening in a series of studies. Tomatine was found in the supernatant of suspensions of pericarp tissue of green tomato fruit and the expressed sap from intact tissues. This glycoalkaloid is synthesised in microsomal organelles and it accumulates in vacuoles and/or the soluble phase of the cytoplasm. The tomatine content of sap was 0.4 mM (Roddick 1976, Eltayeb & Roddick 1984). Tomato glycoalkaloids content depends on cultivar, fruit ripening stage and agricultural practices (Friedman 2002).
The action of tomatine, like other glycoalkaloids, is likely an efficient permeabilisation of the cell membrane as a consequence of the formation of complexes with steroids, such as polyene antibiotics or the induction of apoptosis mediated by reactive oxygen species (Ito et al. 2007). The leaking of PK from cells in P. serpens suggests that the primary mechanism of cell death is cell membrane disruption. Fig. 2A shows that no cells were found in the culture media after treatment with tomatine (50 µM). All cells precipitated to the bottom of the bottle. However, only 10 µM was used for the cell permeabilisation experiments. Part of tomatine likely interacts with sterols in the serum, which explains this difference in drug sensitivity.
In contrast, tomatidine inhibited the growth of P. serpens and caused important morphological and sterol level alterations instead of membrane leakage. Nakamura et al. (1999) determined that ergosterol is the main sterol component in Phytomonas. We did not detect this sterol in our cells. One likely reason for this discrepancy is the poor resolution of the thin layer chromatography used in the previous study. The complexity of sterol composition in the living organism can only be resolved using sensitive instrumentation, such as GS-MS, which was used in the present study.
Some of the remarkable effects of tomatidine on Phytomonas were a decrease in the amount 24-alkylated sterols (∆7-ergosterol, ∆7-sitosterol, ∆7-avenasterol, 24 ethyllophenol and 24 ethylidenelophenol) and an increase in proportion of nonalkylated sterols (cholesta-8,24-dien-3β-ol and its isomer cholesta-7,24-dien-3β-ol) (Table). This pattern is consistent with an inhibition of 24-SMT, which is the enzyme responsible for the addition of these alkyl groups to carbon C-24. The inhibition of 24-SMT of Leishmania amazonensis was also suggested as the primary mechanism of tomatidine toxicity on this parasite (Medina et al. 2012).
Tomatidine inhibits ergosterol biosynthesis in Saccharomyces cerevisiae and prevents cell growth (Simons et al. 2006). The decrease in ergosterols in Saccharomyces occurs in parallel with an increase in zymosterol, which was interpreted as an inhibition of 24-SMT (Simons et al. 2006). Notably, other structural analogues of tomatidine, such as solanidine and solasodine, which are naturally present in potatoes and eggplant, respectively, are growth inhibitors of the yeast-like alga Prototheca wi- ckerhamii (Mangla & Nes 2000). These authors showed an in vivo inhibition of sterol biosynthesis in this alga and inhibition of 24-SMT in a cell-free preparation.
Membrane sterols in trypanosomatids are present as a complex mixture of 24-ethyl sterols and 24-methyl sterols, with cholesterol as a minor component. Our results show an increase in cholesterol levels in parasites treated with tomatidine (Table). These parasites may attempt to counterbalance the altered sterol biosynthesis with a higher cholesterol uptake from the medium. These results are consistent with a previous study (Haughan et al. 1995, Medina et al. 2012) in which an inhibition of sterol biosynthesis increased the amount of exogenous cholesterol uptake. Exogenously acquired cholesterol in trypanosomatids may also presumably be incorporated into the membranes, but cholesterol apparently cannot totally substitute the endogenously synthesised 24-alkylated sterols (Roberts et al. 2003).
Phytomonas have a high proportion of C24-ethylated sterols, such as ∆7-sitosterol. ∆7-avenasterol and ∆7-sitosterol are present in some plants, including American ginseng (Panax quinquefolium L.) (Beveridge et al. 2002). The presence of C24-ethyl sterols in trypanosomatids was described previously (de Souza & Rodrigues 2009) and, in some cases, such as T. cruzi epimastigotes, these sterols represented up to 30% of the total sterols (Beach et al. 1986). The biosynthesis of these compounds involves the participation of a putative SMT2, which, as a rule, adds a second methyl group at C24. SMT2 exhibits distinct inhibitor sensitivities that were described previously in T. cruzi (Urbina et al. 1995). However, more studies are needed to assess the sensitivity of SMTs to tomatidine. Sterols of in the C28-ergostane (24-methyl group) and C29-stigmastane (24-ethyl) types are essential for trypanosomatids. Therefore, the introduction of the alkyl group at C24 of the sterol side chain must confer a desirable property that is needed for some essential cellular function (Haughan et al. 1995, Roberts et al. 2003). In addition to their altered sterol compositions, cells treated with tomatidine showed significant ultrastructural changes and membrane disorganisation, including the appearance of large vacuoles in the cytoplasm that localised to the posterior region of the parasite in some cases. Our results suggest that the main target of tomatidine in P. serpens is the 24-methylation reaction, which causes a depletion of the C29 and C28-sterols and an accumulation of C27-sterols.
Tomatine does not appear to be toxic when consumed orally in moderate amounts (Friedman et al. 2000a, b). The absence of a 5,6-double bond in the B-ring of tomatidine results in a much less toxic molecule in both pregnant and non-pregnant mice compared to the structurally similar solasodine (which contains such a double bond). Friedman et al. (2003) confirmed related findings by Gaffield and Keeler (1993, 1996) on the influence of the structure of glycoalkaloids and aglycones on teratogenicities in hamsters.
In conclusion, our results indicate that tomatine disrupts the plasma membrane, which causes death of the parasite and tomatidine interferes with the growth and ultrastructure of P. serpens, apparently due to the inhibition of alkylated sterols synthesis. There is a clear necessity for novel drugs against trypanosomatids and apicomplexa parasites to help control the diseases caused by these pathogens. Our previous study reported that tomatidine was toxic to the promastigote form of L. amazonensis (Medina et al. 2012). Additional studies are needed to determine whether other trypanosomatids are also sensitive to this alkaloid and whether they have synergistic effects with other antiparasitical drugs to evaluate the potential of tomatidine as an adjuvant in the treatment of parasitic diseases.
To Rosângela Ferreira, for technical assistance.
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The U.S. was under scrutiny and coach Bob Bradley under fire after several poor results, the worst a 2-1 Gold Cup group stage loss _ their first-ever _ to Panama. But after their best showing in a year, Sunday's quarterfinal rout of Jamaica, only a rematch with Panama stands between the U.S. and the final…and vindication.
They'd shown their various Achilles Heels _ disorganization, lack of imagination, a sorry start and worse finishing _ in a June 11 loss to Panama. But after a 2-0 win over a Jamaica team that'd been unscored-upon in the tourney, they earned tomorrow's semifinal date at Houston's Reliant Stadium (7 p.m., FSC, Univision).
"I think they'd certainly be excited about the opportunity to have a rematch. It'll be a good game,'' said Bradley, who wouldn't admit concern over his job status. His quickly-improving team can not only take some of heat off him, but earn a trip to Saturday's final in the Rose Bowl and a shot at the 2013 Confederations Cup.
The U.S. has largely struggled since reaching the knockout stage at the World Cup, including a 4-0 beating at the hands of a Spain team they'd upset in the 2009 Confederations Cup and that 2-1 loss to Panama. But even with starting forward Jozy Altidore likely out with a strained left hamstring, to a man they claim they're gelling at the right time with Sunday's performance their best since South Africa.
"No question. It was complete: We passed well, defended well,'' said Landon Donovan. "As the tournament goes on, you start to build team unity. It doesn't happen right away; you can't come in and expect after a week or two everything's going to be clicking. You can see guys are starting to trust each other, learning how guys want the ball in different spots, and we're starting to play as a team.''
Donovan came in a reserve Sunday for the first time in four years, following his 7 a.m. arrival from his sister's wedding. He'll need to be at his best with Altidore forced off the field in the 9th minute Sunday and undergoing an MRI today in Houston. If he can't play, Red Bull teen Juan Agudelo would likely start.
"I'd love to play. Unfortunately we lost a player that's key to our team and for this tournament, but I feel like this stuff happens for a reason,'' said Agudelo, a New Jerseyan like Altidore. "This is my chance, and I'm focused more than ever now.''
Bradley has made a number of adjustments, ditching his 4-4-2 for a 4-2-3-1 that was more effective than it's been in the past, thanks to former New Jersey natives Alejandro Bedoya and Sacha Kljestan. And he benched centerback Tim Ream, shifting Carlos Bocanegra back inside for leadership and entrusting Eric Lichaj at left back.
"We're trying to get stronger as the tournament goes on,'' said Kljestan.
Panama _ which needed luck, a controversial late goal, and PKs to escape El Salvador Sunday _ has its own issues. Luis Tejada has three goals, but F Blas Perez, who gave the U.S. fits in Tampa, is out with a red card suspension.
Mexico plays Honduras in the other semi, with many in the sold-out crowd of 70,000-plus green-clad El Tri fans likely set to root against the archrival U.S.


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THE best priced 2-bedroom/2-bath apartments at the prestigious Kaanapali Royal. Excellent location - one of the nearest apartments to the beautiful gardens & pool/recreation area. Kaanapali Royal is right in the heart of the famous Kaanapali Beach Resort close to shopping, beaches and restaurants. Kaanapali Royal is known for some of the largest interior living areas in Kaanapali for 2 bedroom apartments. Many amenities are available to you at Kaanapali Royal including a spacious and secluded pool area with B B Q\'s and an entertainment area. Lush mature landscaping are a trademark of Kaanapali Royal. This fine condominium shows pride of ownership throughout. This apartment will be sold furnished with just a few minor exclusions. Move right in or allow a professional management company the opportunity to rent the apartment out for you for part time vacation rentals.
This information is believed to be accurate.
It has been provided by sources other than the Realtors Assoc. of Maui
and should not be relied upon without independent verification.
You should conduct your own investigation and consult
with appropriate professionals to determine the accuracy of the information provided
and to answer any questions concerning the property and structures located thereon.
Featured properties may or may not be listed by the office/agent presenting this brochure.
Copyright 1995-2015, REALTORS® Association of Maui, Inc.


text:
The Joint Tool is an advanced feature that allows you to connect objects together to create all sorts of things, like spinners, wheels, dials, and many other fun things.
Some good examples of the Joint Tool can be found in our Battle For Souls DLC with the Victory Points dial, our Example Spinner Mod on the Workshop, and Roulette on the Workshop by Moneyman12167Casey.
You can learn more about Joints in our Tools Menu Tutorial video.
– To create a joint, select the Joint Tool on the side menu.
– Using the Joint Tool, draw a line connecting two objects together. Always connect objects to the base. In the example gif, two outside blocks are connected to the inside one. In the second example gif with the spinning object, the 4 marbles are connected to the base block.
– You can really tweak joints to make them move how you'd like depending on what you're creating.
– Right click on a joint and click on Joint in the contextual menu.
– The Joints menu will pop up with 3 tabs for the different joints: Fixed, Hinge, and Spring. Each have their own properties that you can tweak. Mousing over each property will give a brief description.
– There's a lot of information to absorb here so read everything carefully and ask questions on the forums as some members of the community are really good at using the joints feature.


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The best tips that I can give you are the following 25 that are basic web design tips which the majority of web designers would also agree with.
Tip 1: Ideally use a light background with a dark font, preferably white background with black text. This is so much easier on the human eye & the last thing you want is getting a visitor to your site & for them to just click off because the content was hard to read.
Tip 2: Content is very important, on every page you should have at least 100 words of copy which will include your most important keywords. Google loves content & the more up to date & relevant the content is the better.
Tip 3: Have your logo positioned top left of your website as this is the first thing the human eye will see. Along with your business or website name, e-mail address & your phone number just to the right of this.
Tip 4: If your website is quite large with many pages & products then make sure you have a Site Search option for your website. Ideally this will be placed


text:
Product ID Number: uk1582
Product Features
Mini Size Hovercraft
The Hovercraft is rechargeable
Comes with mini size RF controller
Battery is not included
This Mini Remote Control Hovercraft features with compact design. It is only 8 x 5.5 x 4cm(LxWxH). It equips with two independent motor. So, it can go forward, backward, backward left, backward right, forward left and forward right smoothly. Get it now and enjoy the fun of surfing on the water.
4 pcs of AA batteries are needed for Charging Station.
1 pcs of 9V is needed for Remote Controller.
Battery is not included
What's inside the box
Mini Remote Control Hovercraft (x1)
Remote Controller (x1)
Charging Station (x1)


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Maryland is apparently the latest target in an ongoing campaign by Texas Gov. Rick Perry (R) to lure businesses and employees to the Lone Star State.
"When you grow tired of Maryland taxes squeezing every dime out of your business, think Texas, where we've created more jobs than all the other states combined," Perry says in a new ad set to begin airing this week in the Washington region, according to WTOP radio.
Besides touting the economic advantages of his state, Perry also takes a shot at Maryland Gov. Martin O'Malley (D), who is weighing a 2016 White House bid.
"Unfortunately, your governor has made Maryland the tax-and-fee state, where businesses and families are paying some of the highest taxes in America," says Perry, who ran unsuccessfully for president last year and has not ruled out a 2016 campaign.
O'Malley told reporters on Wednesday that he had not heard the spots but was familiar with Perry's tactic, which he has already tried in a half-dozen other states.
"I know he does that tired old P.R. gimmick all around the country," O'Malley said following an event in Baltimore to promote public service projects.
Similar ads have already been aimed at California, Illinois, Missouri, New York and Connecticut, according to WTOP.
In some states, the governor has fought back.
Missouri Gov. Jay Nixon (D), for example, countered Perry's radio ads with those of his own, which promoted the economic strengths of the Show Me State.
In a statement, O'Malley's office highlighted some of Maryland's achievements during his tenure, which began in 2007.
Among them: a No. 1 ranking of the state's schools for five years in a row by Education Week, and a No. 1 ranking for "innovation and entrepreneurship" for two years in a row by the U.S. Chamber of Commerce.
"Instead of engaging in P.R. stunts, Governor Perry should come to Maryland to see firsthand the better choices that have led to these better results," said the statement from the governor's office.
Perry's office did not immediately return a phone call Wednesday.


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At first sight Rwanda is a young country – young, green and clean. Most of the time we have no idea what the hell people are saying but they seem to smile a lot. After almost four months on the road The Land of a Thousand Hills managed to climb all the way to the top of our "favorites list", seemingly without even trying.
A small landlocked country, Rwanda is most known for the genocide that brutalized its people in 1994. It is often viewed as one of the most savage genocide that mankind has ever seen but as you drive through the spotless, friendly streets of Kigali you find it so difficult to believe. Kigali seems more like a European city than an African city and it is not just because of the cleanliness. Unlike most other African countries there is no method in the madness of the traffic because there is no madness in the traffic. Even with driving on the wrong (right) side of the road for the first time, both Guillaume and I feel safer on Rwanda's road than in any one of the nine other countries we had driven through. Car doors are unlocked, people seem very honest and there is a general feeling of safety and order.
Yet just 18 years ago this country was thrown into the deepest, darkest pit of cruelty and trauma – worlds apart from where we were back then. April 1994 was for South Africa, our home, one of the happiest times and brought our long sought after democracy, for Rwanda it brought the death of a country. While we celebrated freedom, they were running for their lives.
One word keeps coming to mind to describe the atrocities that had taken place here – unimaginable. The hideousness of the genocide, all the blood spilled and bodies rotting in the streets, all the fear, hate, anger and mankind's ability to commit such ghastly acts is unimaginable, and then 18 years later your mind can hardly comprehend that all this played out in the stunning, welcoming boulevards of these cities and towns. If not for all the museums, books, mass graves and historical reminders of the genocide one would never imagine that Rwanda was once a society of "brutal, sadistic and merciless killers and innocent victims".
But at a closer glance I realised that the genocide is still everywhere – "many of the survivors are young and will carry the trauma with them through their lives". About 500 000 women were raped, intentionally by HIV-positive attackers and they will carry the scars with the disease till they die. The Genocide Museum in Kigali proclaims; "Almost every corner of Rwanda was in some way touched by the genocide. It is impossible for us to forget the past but also extremely painful to remember".
After hours spent in this museum I was emotionally drained and almost relived to leave the building to roam Kigali's peaceful streets but the genocide seems to sneak up on you from within the beauty in the city. The very local and cheap hotel we stayed at is run by three kids, no older than 22, they too were orphaned by the attacks. Saddened again by another account of the carnages it does however not come as a surprise. The genocide made orphans out of 300 000 children. Kids were also not excused from dreadful deaths during the genocide and many were tortured, hacked to death and shot. The museum commits a room to these tiny victims introduced by a heart wrenching dedication: "In memory of our beautiful children who should have been our future". In 1994 Rwanda lost its youth and the ones that remained lost their childhood – scar that are still visible today.
In the end Kigali was a whirlwind of emotions and contradictions. A scenic as well as humanitarian inspiration. A phoenix which had risen from the ashes of rampant, lawless looting and unimaginable cruelty. I left the magnificent land of a thousand beautiful hills, a million murders and a nation oozing immense forgiveness, behind, with one quote I will remember forever, a quote by Felicien Ntagengwa which I hope the whole world will remember: "If you knew me and you really knew yourself, you would not have killed me. "


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Kronos is a slot machine made by WMS Gaming. The games made by WMS, like Kronos slots, are quite new to the UK, despite having been very popular in the USA for many years.
Kronos is one of the best examples of their G+ series of slots from WMS and is still really popular in Las Vegas casinos, even though it is old. A good example of game play coming first in the design process.
Bonus Round in Kronos Slots
As one of the biggest hits in Las Vegas, the Kronos slot is famous for its bonus round. When you get a bonus win (three Kronos symbols anywhere on the screen), you get a free spin bonus. During the bonus you get a lot more wild symbols and also a lot more Kronos symbols, so the chances of a big win are much improved.
More Slot Machines like Kronos Slots
As well as Kronos slots, we also have the following free online slot machines made by WMS:
Kronos Slots Demo
Kronos is one of the most popular G+ WMS slots in Las Vegas and has inspired a generation of new games that are very similar.
The big thing about these games, are the free spin bonus rounds, which always creates a great atmosphere and anticipation. Kronos is quite a highly volatile game, meaning you can go through long periods without a win, but then suddenly get a super massive payout


text: This doesn't really remind me of pear so much as it reminds me of apple pear cheesecake. Or cream cheese frosting. It's unpleasantly tart hot so I let it chill in the fridge for a few hours. Cold, this tastes both refreshing and decadent. It's like fresh apple/pear juice but there also a creaminess that's more tangy than just cream. So all combined it's apple pear cheesecake. I enjoyed it. :)


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1. a cloud-like mass or layer of minute water droplets or ice crystals near the surface of the earth, appreciably reducing visibility.
2. any darkened state of the atmosphere, or the diffused substance that causes it.
3. a state of mental confusion or unawareness; daze; stupor.
4. Photography. a hazy effect on a developed negative or positive, caused by light other than that forming the image, by improper handling during development, or by the use of excessively old film.
5. Physical Chemistry. a mixture consisting of liquid particles dispersed in a gaseous medium.
–verb (used with object)
6. to cover or envelop with or as if with fog.
7. to confuse or obscure.
8. to bewilder or perplex.
9. Photography. to produce fog on (a negative or positive).
–verb (used without object)
10.to become enveloped or obscured with or as if with fog.
11.Photography. (of a negative or positive) to become affected by fog.
1. a second growth of grass, as after mowing.
2. long grass left standing in fields during the winter.
I seek the faintest trace of fog
On the mirror,
A sign you might still be there,
That you still thrive and sigh,
Measured in wisps of vapour,
In faint feeble heartbeats.
But the mirror does not collaborate,
Refusing to reflect life's shadow,
To bear ghosts and witness,
Pristine in its defiance,
The statement all too clear.
5th December 2019
13 hours ago


text: BUSINESS NBC revamps Digital Studio 5:00 AM PDT 4/17/2008 by Alex Woodson , AP FACEBOOK TWITTER EMAIL ME Empty Related: NBC nabs Rosario Dawson Web seriesNBC Universal relaunched its Digital Studio on Thursday with a focus on branded entertainment and two series already in the works.The studio is headed by its vp, Cameron Death, who was hired last year by Ben Silverman, co-chairman NBC Universal Entertainment and Universal Media Studios. Death and his team report to Silverman and Vivi Zigler, executive vp digital entertainment and new media at NBC.The two missions of the studio, Zigler said, are to provide an avenue for branded entertainment and to "find amazing talent." To those ends, the studio has named Omnicon Media Group Digital as its first strategic partner and has signed a deal with Electric Farm Entertainment to produce two shows."Gemini Division," a sci-fi series starring Rosario Dawson, is set to debut in the summer, while "Woke Up Dead" is a zombie comedy in preproduction. Both will be available on company Web sites, including NBC.com, and the network has the options to bring both to TV with Sony Pictures TV International holding the series' international rights.Intel, Microsoft, UPS and Acura TSX have signed on as sponsors for "Gemini."Zigler said the studio has been in the works since Silverman joined the network a little more than a year ago. One of the first conversations she had with him was about the future of digital entertainment."He really set the seeds for this," Zigler said. "He said, 'What if we put together people who understood digital? What if he started them off on this path?' "The exact plan started to coalesce in November, when Death, formerly U.S. director of branded entertainment at MSN, got his start at the network. Silverman worked with Death at the portal through Reveille's partnership with MSN.Although the NBC studio is entering a crowded online video market -- Disney and Warner Bros. already have their own digital studios -- Death said the high standard of quality combined with NBC Uni's distribution reach and an interactive approach will help the studio stand out."It's not just about the video," Death said. "It's about deep interactive experiences that gives consumers and viewers an option for what's next, after they consume the video."Zigler said the studio would look to distribute its content to partially and fully-owned NBC Uni sites, including Hulu, and mobile destinations as well as sites that the company doesn't own. FACEBOOK TWITTER EMAIL ME Alex Woodson email@example.com @thr AP email@example.com @thr


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Monthly Archives: October 2012
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text:
USTA New England Magazine
October, 2008
NEWPORT, RI--The International Tennis Hall of Fame played host to the Champions Cup last summer, a regular stop on the Outback Champions Series tour. With eight world-class players vying for the title, only one man remained standing after Sunday's final. Pat Cash proved that grass is still his domain, as the 1987 Wimbledon champ served and volleyed his way to the title, beating out Hall of Famer, Jim Courier.
But the biggest story of the tournament came from John McEnroe's default during a match against MaliVai Washington. After McEnroe took the first set 6-3, Washington was able to climb back to 4-2 in the second set. After McEnroe received four code violations for multiple obscenities and unsportsmanlike conduct, he was tossed out of the match. Jim Courier chimed in on the situation. "I think the umpire has to enforce the code. If John makes some infractions, then he should be penalized appropriately," he said. Despite the default, McEnroe remained in the tournament due to the round-robin format.
On Sunday afternoon, the pressure was on, with the Champions Cup title just one match away. Courier and Cash had previously faced off in the 2007 third place match in Newport, in which Courier took out Cash in a three-set tiebreak. But Courier was a bit uneasy going into this final. "Pat is such a natural grass-courter and a very natural serve and volleyer," says Courier. "I'm a fish out of water here –- I'm serving and volleying in a first and second serve which I probably haven't done since I was 18 or 19 years old at Wimbledon." After winning the first set 6-3, Cash lost a bit of control in the tail end of the second set. Courier saw an opportunity to break Cash, and proceeded to step up his game. The two battled it out in a seemingly never-ending 22-point game in which Courier saved five match points, bringing the second set to 5-4. Courier put in a great effort, but the birthday boy didn't get his wish to conquer Cash's big serve-and-volley game, losing 6-3, 6-4. "He clearly outplayed me today," said Courier. When Cash was asked if he felt guilty about beating Courier on his birthday, he laughed and said: "No…"
Sunday's matchup also included a battle for a third place finish between Magnus Larsson and MaliVai Washington. This was Larsson's first time playing the Hall of Fame Champions Cup, and he cinched the win 6-4, 6-4, which proved to be a very competitive match. "Larsson has one of the best serves on the tour," says Washington, a sentiment he shares with Cash. After the match, Washington was asked how he feels overall. He said jokingly, "Well, I feel like I can beat anyone in this crowd."


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A black bourgeoisie perspective on U.S. politics
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Cheryl Contee aka "Jill Tubman", Baratunde Thurston aka "Jack Turner", rikyrah, Leutisha Stills aka "The Christian Progressive Liberal", B-Serious, Casey Gane-McCalla, Jonathan Pitts-Wiley aka "Marcus Toussaint," Fredric Mitchell
Special Contributors: James Rucker, Rinku Sen, Phaedra Ellis-Lamkins, Adam Luna, Kamala Harris
Technical Contributor: Brandon Sheats


text:
ESTEEMED London-based artist Daniel Silver has collaborated with Acne on a capsule collection for spring/summer 2012 that was unveiled last night at their Dover Street flagship store alongside an exhibition of Silver's own work.
The upshot? A perfectly edited collection of Acne's signature modern silhouettes, in layered up clashing prints (set to be a huge trend for next summer). Star pieces include a collarless kid leather jacket in peach, a tribal and animal print pencil skirt, and a pair of silky tomato red trousers.
Although Silver has worked with Acne in the past (he was photographed for Acne Paper's Atelier issue at his studio last year), this was the artists first collaboration with a fashion house.
"It happened organically", explained Silver. "I met Thomas Persson, the editor of Acne Paper, at an opening of a show I was in at Lismore Castle and we became very good friends. Jonny Johansson saw the issue of the Acne paper I was photographed for and was interested to meet up with me and so we started to talk about processes of making and how it's different or similar in fashion and in art."
Silver began designing the collection in much the same way he works on fine art projects; cutting and pasting collages, without too much preconception of what the results would be, confessing, "It was then left to the design team at Acne to make sense of what I did. I was very taken with the way they translated my work into fashion."
Before moving to London to study Fine Art (at Slade, and then the RCA), Silver grew up in Israel, and with parents and grandparents hailing from Eastern Europe, South Africa and Zimbabwe. He explained how his upbringing influenced his designs for Acne, and his own art.
"The African fabrics I used are traditionally made in Holland and shipped to Africa. I came across them outside my studio in Dalston and was drawn to them more for the reason that they are around me and I like to pick from my close environment - so seeing them in Africa and seeing them here in London on people on their way to church on Sunday morning made it familiar to me. I draw from the different places I have lived in and the memories of my family's stories and places. I feel connected to these places in different ways. I draw from storytelling, myths and imagery as well as art history and archaeology of these places."
The collection will be sold exclusively at the Acne's Dover Street flagship, and a video of the collection can be viewed on the newly made over Acne website at ACNESTUDIOS.COM.
Acne, 13 Dover St, London, W1S 4LN, 0207 629 9374
YOU SHOULD SEE: The latest Acne show...
YOU SHOULD SEE: Tali Lennox as Acne's star muse...


text:
The last I remember would probably be during season 4s run
The last I remember would probably be during season 4s run
Have any Canadian s preordered these at ebgajes? Gamestop owns it here
There are no pre-orders on GameStop.
Placed my pre-orders in store today. You may need to provide the SKU for them to find it as details are still scarce. It's Digimon product though, I could never go wrong with Digimon products!
Its in store only at this moment
I think the last Digivice in the US was Xross but I didn't pay attention to it so I'm not sure. Otherwise it was the IC.
As for my 2 cents. $20 seems to much for a bit of plastic and too little for a Vpet.
I would have expected $25 for the Vpet, $30 for Digivice.
That said, my question is, Is it cheaper for Bandai to redesign the OG Vpet electronics or to just use the 20th that they likely still have on hand?
I know that sounds stupid, but from what little I know, a lot of the expense of production is in the design phase and while the 20th Vpet were more expensive, they were also Pbandai, which always has a mark up in price.
If I remember right they were $30-$35 and from the few PBandai Gundam kits I have payed attention to, a $10 premium is pretty reasonable.
Sooo, there's a chance?
Just thought I'd throw another option into the ring for everyone to ponder for the next 2 or 3 months until we get more information.
They were $20.
For $20 I'll still buy 1 or 2 of these if they are Vpet rereleases though. I'd like to see some more Digimon Merch making it's way into the US that isn't cutsy blindbag type stuff.
I am going to pre-order them but I really hope its a v-pet and not a keychain if its a keychain I might just sell it for what I pay + s/h I got to many keychains as it is.