text:
Mental health issues in students are a growing concern, especially in Florida, where around 13% of the youth face some mental health challenges. Moreover, the stigma surrounding mental health also limits the availability of proper help, especially for students from low-income backgrounds who are already affected by the growing economic crisis. Here are some tips from mental health experts to help to lead a better life.
Maintain Your Sleep Cycle
Proper sleep is the single most important thing in your life. Without sleep, you can’t function properly or think properly, and your brain gets tired easily. This leads to a plethora of issues, including several mental health issues. Try to add at least 8 hours of sleep to your routine and take naps at least once a day if you can. After a few weeks, you’ll notice a drastic change in your mental and physical health.
You are what you eat! This also applies to your brain. If you eat healthy and nutritional foods, your brain will stay healthy and won’t get exhausted so quickly. Moreover, eating well automatically improves a lot of your body functions, which in turn can reduce the signs of stress, depression, and anxiety. Also, try to have more meals in a day. You can decrease the portion size to prevent any stomach issues.
Do Mental Exercises
This may not seem like an important enough tip, but a lot of your mental health issues stem from poor living standards. If you’re sleeping, eating, or working out, it’ll definitely affect your brain. Doing a few mental exercises improves blood flow and helps strengthen the connection between your brain cells. This prevents your brain from getting exhausted, which helps you stay active. You can learn a few easy brain exercises through your therapist, or you can play a few brain games to achieve more or less the same effect.
Find Support Groups
Support groups are a great way to find like-minded individuals for support. Joining a support group not only helps you open up your feelings, but you also get the chance to connect with therapists or other professionals for help. You can find support groups at your university by talking to your student counselor. They usually have a list of such groups and organizations offering help.
Take Small Steps
Doing too much at once can overwhelm anyone. On top of that, if things don’t work out according to your plan, you end up feeling like a failure. If this continues, it doesn’t take long for your to develop some sort of mental health issues. The best way to prevent feeling overwhelmed and burned out is by taking small steps and giving ourselves credit whenever we achieve a milestone, no matter how big or small it is.
Reach Out To People You Trust
The biggest cause of a lot of mental health issues is bottling up your feelings and having no outlet to channel them. This can isolate you pretty quickly and worsen your state. Always keep a line of communication open when you feel overwhelmed. Moreover, people around you can help you find proper support and help you need to maintain a healthy mental state.
Find Mental Health Resources with Helping Miami
At helping Miami’s website, you can find several resources on many issues related to mental health, along with programs run by the government and private organizations to help people with mental health issues. Besides that, we also have resources related to educational assistance in Miami, non-profit fundraising, traffic and moving violations, and autism. You can read all of our blogs by visiting our resource page.

domain: psychology


text:
Last month, Justin Timberlake released the audio for his new song Mirrors for our listening pleasure. Last week, he performed that song live on Late Night with Jimmy Fallon. Today, we get to see the just-released music video for the song. As you can see above, the video tells a supercute love story … and then JT breaks it down and gets his boogie on in a hallway of mirrors. Symbolic. Check out the video and see what you think. Thus far, Mirrors is my favorite jam from his just-released new album The 20/20 Experience … do you think the music video represent the song well?
|Charlie Sheen Rejects Farrah Abraham Publicly – FOX News|
|More Troubles For Miley Cyrus – TMZ|
|Kim Kardashian Stuns In Maxi Dress – Huffington Post|
|Drug Outbreak Behind Lindsay Lohan Rehab Switch? – FOX News|
|What Does Amanda Bynes Have To Say To Drake Now – Huffington Post|
|Ouch... Justin Bieber Gets Snubbed – Huffington Post|

domain: celebrity


- "Blasters charged!"
- ―Gunship Fighter pilot
The gunship fighter was the direct predecessor of the LAAT/i and advanced gunship fighter. It was used in the Clone Wars by the Clone Army.
The gunship fighter was most effective at destroying enemy starfighters as well as ground units. The fighters were armed with a single laser cannon and were used in space or in atmospheric conditions.
Ad blocker interference detected!
Wikia is a free-to-use site that makes money from advertising. We have a modified experience for viewers using ad blockers
Wikia is not accessible if you’ve made further modifications. Remove the custom ad blocker rule(s) and the page will load as expected.


2008 IMPALA FUSE BOX LOCATION
Interior Fuse Box Location: 2006-2016 Chevrolet Impala
The video above shows how to replace blown fuses in the interior fuse box of your 2008 Chevrolet Impala in addition to the fuse panel diagram location. Electrical components such as your map light, radio, heated seats, high beams, power windows all have fuses and if they suddenly stop working, chances are you have a fuse that has blown out.
SOLVED: Where is fuse box located impala 2008 at? - Fixya
Needing to know where the fuse box is that houses the windshield wipers for a 2008 chevy impala fuses are located on the passenger side of the vehicle. You will find an access panel near the floor.
Fuse box Chevrolet Impala - fusesdiagram
Fuse box in passenger compartment Chevrolet Impala. fuse box location. The fuse block is on the passenger side of the vehicle in the carpet molding.
Chevrolet Impala - 2008 Fuse Box Diagram
Impala - 2008 Fuse Box; PASS-Key The underhood fuse block is located in the engine. compartment. See. Engine Compartment Overview on. page 5-14. for more information on location. Notice: Spilling liquid on any electrical components. on your vehicle may damage it. Always keep the.
2008 Chevrolet Impala Fuse Box Location - Best Place to
2008 chevrolet impala fuse box location The chevy cruze is a compact car that has been manufactured since 2008. Read more chevrolet reviews to learn you turn ignition on then save yourself time and change the main fuse box under the Remedy dealers will replace the fuse 2004 2008 chevrolet aveo vehicles equipped with daytime running lights drl.
2006-2016 Chevrolet Impala Interior Fuse Check - 2008
The video above shows how to check for blown fuses in the interior fuse box of your 2008 Chevrolet Impala and where the fuse panel diagram is located. If your map light, stereo, heated seats, headlights, power windows or other electronic components suddenly stop working, chances are you have a fuse
2008 Impala Fuse Box Location - Wiring Diagram Fuse Box
This is a post titled 2008 Impala Fuse Box Location, we will share many pictures for you that relate to "2008 Impala Fuse Box Location". Hopefully the picture gallery below will be useful for you. Relax, if the image is not enough to help you, you can type what you are looking for on the search form.
Location of the fuse for a 2008 chevy impala windshield
Jan 27, 2012location of the fuse for a 2008 chevy impala windshield washer pump - Chevrolet 2008 Impala question. It is located in the under the hood fuse box fuse is near the bottom marked WSW. Look in the manual for fuse box (under hood) I read that Chevy recommended repacing the 10 amp fuse with a 15amp. Looking for the fuse for the wiper
Where is the fuse box for 2008 impala - answers
Answers is the place to go to get the answers you need and to ask the questions you want
2006 to 2013 CHEVY IMPALA FUSE BOX AND RELAYS - YouTube
Jan 18, 20172006 to 2013 CHEVY IMPALA FUSE BOX AND RELAYS Chelby anjos. Driver Side Low Beam Works Intermittently 2008-2012 Chevy Malibu 2004 Chevy Impala Engine Won`t Start 60 amp open fuse


text:
from Pasta & Patchwork
A globetrotting mama cooking, making and writing her way through life
from Smiles and Trials
Family life with two small children; I blog about outdoor living, crafts and messy play, and travel.
from Craft with Cartwright
sewing Patterns, craft tutorials and recipes.
from craft with cartwright
Recipes, tutorials and patterns
from Em And The Wilds
UK lifestyle blog - exploring, growing goodness, feathering our nest & cooking up feasts. With whatever else takes our fancy in between.
Mummy, Talipes Awareness, Lifestyle, craft, reviewing blog

domain: artistic


text:
God bless the local library, am I right? I mean, I almost never ever step into mine because I don’t have many reasons to, but in a situation where I do, I am the type of person to leave with five books that I plan to have read within two weeks. Yes, two weeks. And the nice thing about going to the library is you can find all sorts of books, including ones that are out of print 100% that you can barely find even on Amazon, at least not in a presentable condition. So, since I knew that Fortune and Glory was due to be re-printed by Marvel this year, I figured I’d take the opportunity to get in a review of the book before everyone re-discovers what I’m sure will be referred to as “classic Bendis” one day.
For those interested, the re-release of the book will feature new scenes and be in full color as opposed to the black and white that the original version is. It should be entertaining! You can pre-order the book set to come out March 24, 2010 right now on Amazon.com. And believe me, you may want to get that pre-order in before you even read my review.
Believe it or not, back in the day Bendis used to write everything but superhero comics. The man was big into crime and true crime, and when he wrote a little book called Goldfish, it turned some heads. That’s what this book is about. Fortune and Glory is the true Hollywood story of how Bendis caught some fame with his graphic novel Goldfish, and how it was optioned to become a feature film. Bendis takes us through every step of the story – from the awkward phone calls, to getting an agent, through going to various meetings with studio execs all the way to the picture being optioned, him writing a draft, and everything being put in place for the movie to be made. Of course, as well all know, SPOILER ALERT the movie wasn’t made. You probably would have heard about it as it was. As much as Bendis was getting the “next Kevin Smith” treatment from Hollywood (to the point where Miramax adores him), his movie never happened, despite his script being adored by all those that read it. And through it all, we get to see a wide-eyed Bendis learn just what really happens in the magic land of Hollywood that we’ve all dreamed of joining at one time or another.
What makes Bendis such a catchy, fun, and fresh writer to a lot of us is the way that Bendis handles dialogue. This is quite possibly my favorite thing about him, and is what is so great about a book like Ultimate Spider-Man. While Bendis might be most at home writing a crime story (as was proven by the optioning of Goldish and all the work done on Daredevil), he is always best when it comes to dialogue. And guess what? That’s all this book is. The book is a reinterpretation of conversations that Bendis had during his experience, and all with his trademark spin and humor on the entire situation. Some of the scenes are so downright hysterical that you have to wonder how close is his writing to what actually happened. While I had no luck finding images of the book online, there is a great scene in the book when he first finishes his draft of the Goldfish script and his friend (who reads scripts for a living and tells people how to improve them) remarks that it was 280 pages versus the average 120. Bendis’ reaction is absolutely hysterical.
Of course, if the book didn’t have good a good flow in general, it’d obviously be no good. You’re in luck though, because this is done thanks to highly entertaining cartoons drawn by Bendis himself. Bendis has obviously concocted a pretty crazy script that is done with many interesting forms of visual storytelling, and we can really see a lot of the groundwork for how Bendis would go on to instruct Oeming in Powers. While this is obviously nothing like what we had seen Bendis do before in his crime books, the cartoonish nature of the book makes the story that much more affable. Seeing Bendis in a brand new light, as a character in his own book that he quite often makes jokes at (especially in an opening scene in which Bendis describes himself as an artist and is portrayed as a stereotypical guy in a black shirt and pants smoking next to a comic rack), makes the book that much more personable.
I’d also be remiss if I didn’t at least mention that, towards the end, the story becomes quite literally just as much Marc Andreyko’s story as it is Bendis’ (to which Bendis makes note). Despite this being one of two collaborations between Bendis and Andreyko (the other being Torso, which actually becomes a part of the book in and of itself (which is also in the process of being adapted to film…)), it definitely shows the great friendship and understand that the two have with one another.
If you’re able to find a copy of Fortune and Glory, I’d snag it up right away. It’s an insanely entertaining read, and for those of us that will never make it to Hollywood, it is definitely an interesting and eye opening look at what exactly happens over there. An interesting point to note is that Bendis states near the midway point of the book that he could be lying and we’d never know, but in this situation, due to the complete candid nature of the tale, I’d imagine that most of this happened. It’s a very believable tale, especially in regards to the reactions of the studios to the work that he is doing. While I’m only somewhat familiar with Bendis’ pre-Marvel work (something that I definitely want to correct the more stuff like this I read (and, as a side note, I’m going to try a lot harder to find things besides Jinx and Goldfish)), I can easily say that this is by far the Bendis book – it’s full of his rich dialogue, good humor, and personal approach to storytelling.

domain: library


text:
Pit boss wood pellets are available in a competition blend hickory and apple. At traeger we have mastered the craft of flavor.
How To Season And Maintain A Wooden Cutting Board Serious Eats
How To Finish A Live Edge Slab Tips Tricks Youtube
Woodworking With Mesquite
By manufacturing our natural hardwood pellets in our own traeger mills across the usa we can guarantee that from mill to grill only 100 natural virgin hardwood makes up our delicious wood fired flavors.
Best oil for mesquite wood. You can used larger wood chunks or smaller smoking chips. The tung oil offered by real milk paint is a pure oil meaning it comes without any additives or distillates. 100 hand painted by talented artists.
Our end grain wood flooring products are the most durable and resilient flooring surfaces available. Museum quality unbeatable price and high customer satisfaction is our motive. The traditional ingredients for wr finishes are a solvent paraffin wax and a.
Water repellents wrs note not water proof and water repellent preservatives wrps leave the wood with a natural look it may not be apparent the wood has a finish especially a few weeks after its appliedthey are cleartransparent and help to reduce warping and cracking by limiting water absorption. Beer can chicken how to make beer can chicken. End grain is made by continuously cross cutting or slicing logs or cants into blocks or rounds with the annual growth rings exposed on every piece.
Product description enhance the character of wood with our great water resistant finish. Here is the deal i have an a maze n smoker in the mail right now and i bought hickory cherry maple and mesquite dust out of those 4 whats the general consensus on which has the better flavor for cheese. I plan on a 3 4 hour smoke for the duration not sure if that has any affect on wood type.
Different types of wood will produce different flavors and different types of trees have unique compositions and burning points. Thrift yourself a small cast iron pan to use in your bbq i see them all the time at value village and you will have it made in the shade. Real brick and mortar art store located in carrollton texas.
Most other oils can be anything from a thinned down varnish to polymerized tung and most have petroleum distillates added. Affordable elegant framed oil paintings in dallas texas. Pit boss is another big name in the smoker world and these are some of their very best wood pellets for smoking.
Sharp cheddar gouda mozzarella colby jack. I like to take a rear leg of javelina depending on the size of the animal and put in a slow cooker overnight. I add two cans of cambells cream of mushroom soup salt and pepper to taste.
All you need is a gas grill and a small pan to put those wood chips in. Now dont get all scared about using wood chips you dont need a smoker to use them.
Interior Stain Colors Sherwin Williams
Live Edge Slab Table Part 2 Sanding And Applying A Finish
Waterlox Coatings Corporation

domain: hobby


text:
Hymns For Hearing God Kampala — IT is not likely that you will attend Mass and not hear one of his hymns. With more than 700 of them to his. "The 15 years I spent in the seminary helped me understand what serving God me. Lyrics: 1. I am a child of God, And he has sent me here, Has given me an earthly home With parents kind and dear. (Chorus) Lead me, guide me, walk beside me, Unmatched talent, a passion for ministry
"Ragtime" is rated PG-13 for some offensive language. director of Lafayette College Art Galleries and collections curator. Live music and dance performances throughout the day will be held in the n.
The Bollywood dance group performed three songs and unveiled a preview. It can be watched on FIOS Channel 837 (GoLocal Liv.
J azz gained popularity in America and worldwide by the 1920s. Nothing quite like it had ever happened before in America. New exuberant dances were devised to take advantage of the upbeat tempo’s of Jazz and Ragtime music.
Classic Ragtime Piano by Ted Tjaden. Jéan-Baptiste Lafrenière appears to have published 44 compositions (all of which are below), includings rags, marches and two-steps, mazurkas, gavottes, songs and waltzes.
Santa Cruz Ragtime Festival When: Saturday and Sunday. Venues include Calvary Episcopal Church, the Calvary Parish Hall, the Santa Cruz Vet’s Hall (for dance show), the Museum of Art & History (for.
names of dances : allemande beguine bergamask bolero bossa-nova boston bourrÉe bransle buck-and-wing cabriole cakewalk canary cancan carioca cha-cha
Features ragtime, march and waltz midi files. This is the largest ragime midi collection on the internet by one sequencer. All files have been edited by Greenfield Bowie.
The worlds largest and longest summer-long Vernacular Jazz, Swing Dance festival. The camp’s historical root system is closely linked to the presence and legacy of legendary swing dancer Frank Manning.
Also, Fallon appeared in a Barbershop Quintet Ragtime Gals skit, along with Sting. The song was about a pugilist, or boxer, who “finally learned how to dance.” It sounded INCREDIBLE! After another.
Dance has been around for a really long time and it’s popular all around the world! Many dance styles have influenced modern dance. Read on to learn more about it!
Read Customer Reviews & Find Best Sellers. Free 2-Day Shipping w/Amazon Prime.
Vintage ballroom and social dance classes, workshops, and balls for dancers of all levels. Covering dances from the Victorian era through the Ragtime, 1920s, and swing periods.
Birger’s Ragtime Band performs at jazz festivals, jazz clubs, church concerts, conferences, cocktail parties, Gatsby parties, birthday parties, dance events, weddings etc.
Are you ready to revel in some ragtime? William Pemberton. and head of performance at the University of Michigan School of Music, Theatre & Dance. Pemberton said the idea for the concert came about.
An American Ballroom Companion presents this social dance demonstration from the Library of Congress. Watch this demonstration traditional dance video on the Ragtime Dance: Castle Walk.
Welcome This section of my site contains MIDI files, which I have sequenced, of ragtime piano pieces by : Scott Joplin, James Scott, Joseph Lamb, Jelly-Roll Morton, Eubie Blake, and others.
Replete with the embroidery of epithets and metaphors, their accounts suggest the E.L. Doctorow of “Ragtime” channeling the i.
Get the DVD: http://dancetimepublications.com/products/how-to-dance-through-time-volume-ii/ A glimpse of the some of the dances that were popular during Worl.
AUDITIONS FOR: “Ragtime the Musical” presented by the Charleston Light. READING Auditions are Wednesday, August 15 at 7 p.
Swing dance is a group of dances that developed with the swing style of jazz music in the 1920s–1940s, with the origins of each dance predating the popular "swing era". ". During the swing era, there were hundreds of styles of swing dancing, but those that have survived beyond that era include: Lindy Hop, Balboa, Collegiate Shag, and Charl
The Commonwealth Vintage Dancers reconstruct, teach, and perform American ballroom dance of the 19th and early 20th centuries. In addition to performing dances of the Federal, Civil War, Gilded Age, Ragtime and Roaring Twenties Eras, we host public balls, dance classes and multi-day workshops for historical dancers.
Ragtime – also spelled rag-time or rag time – is a musical style that enjoyed its peak popularity between 1895 and 1918. Its cardinal trait is its syncopated, or.
Scott Joplin, the "King of Ragtime" music, was born near Linden, Texas on November 24, 1868. He moved with his family to Texarkana at the age of about seven.
Cliquez ici pour le français Social Dances of the Ragtime Era. Music: During the 19th century, most of America's music, dances and fashions were imported from Europe, as composers and dance masters emulated the latest styles from Paris and London.
Hymns Of Eden Christian Band Download and stream Hymns Of Eden songs and albums, watch videos, see pictures, find tour dates, and keep up with all the news on PureVolume.com/hymnsofeden Of the many brass bands that have flourished in Britain over the last 200 years very few have documented records covering their history. This page is an attempt to collect together information about such bands and make it available to all. Full Index of The Hymns and Carols of Christmas. Notes 1. All links open
The dawn of the 20th Century brought a new, modern dance to match the wild new rhythms of Ragtime. It had none of the complicated footwork of a Victorian ballroom.
She danced in major productions, including the original “Follies,” and has choreographed for numerous Broadway shows, including “Ragtime,” “Annie Get Your Gun” and “Pal Joey.” Though she grew up in th.
The dance that epitomizes the 1920’s is the Charleston. The Charleston was introduced to the public in the Ziegfield Follies of 1923 by the all black cast Afro-American Broadway musical "Running Wild", and became so popular that even today, it is still a symbol for the 1920s Jazz Age.
Once a year, Greenfield Village in Dearborn, Michigan puts on this great, fun and family friendly event called Ragtime Street Fair. Employees are taught to d.
MIDI & Tablature collection of Fingerstyle Guitar. Music and Tablature from Steve McWillam, Mike Martin, and others.
His mission in "How the Beatles Destroyed Rock ‘n’ Roll" is to help you forget all that you’ve learned. Can you dance to it? Starting with ragtime, Wald recounts a nascent industry that was built a.
This webpage will serve as a tribute to a man who loved to share his gifts! He had many but one of his greatest God-given talents was the gift of music.
Amazon.com: HOW TO DANCE THROUGH TIME Vol. II. Dances of the Ragtime Era 1910-1920: S. Fava, L. Ewing, Carol Teten: Movies & TV
Still working on finding a distinctive musical voice to call their own, ”Joseph” is chockfull of well-established styles, inc.
Ragtime bands! A Banjo Jamboree! Dueling Pianos! And a "Dance Your Feet Off" swing dance marathon. Whew! With all the new and exciting action going on for the 31st annual Jazz Bash by the Bay presente.
VINTAGE VOLTAGE, 7:30 p.m., Moose Lodge, 567 25 1/2 Road, dance to country favorites and rock ‘n’ roll. performing Bach to the Beatles and Renaissance to ragtime, in the summer concert series, take.
George Gershwin, Robert Griffith, Charles Hunter, Scott Joplin, Karl L. King, William H. Krell, Joseph Lamb, Jelly Roll Morton, Spiritual Traditional, Eastern Brass Quintet – Ragtime – Amazon.com Music
There also are chapters on baseball, on dance and on when people used to gather around the. has long been considered one of the finest ragtime pianists in the country. Inspired by the musical atmos.
Vintage Voltage, 7:30 p.m. Friday, July 13, Moose Lodge, 567 25 1/2 Road, dance to country favorites and rock ‘n’ roll. pe.
Virtual lines are the latest evolution in theme parks. They can dance or take photos with an actor in the costume of Hashtag the Panda, a staple character from Jimmy Fallon’s show or listen to a pe.
Throughout the history of dance, many interesting facts marked this activity as one of the most useful tools for visual expression, exercise, social interaction, celebration and religion.
June 5, 2018 Just over a year after their debut with the original musical RAGTIME WOMEN, featuring little known Ragtime. B.
Cakewalks in the Ragtime Era by Ted Tjaden. As can be seen in the video below, cakewalking involved exaggerated steps with high kicks involved believed in part to be a "send up" by black slaves of the elitist mannerisms of their slave owners:
In southern New Jersey, a high school production of Ragtime, the musical adaptation of E.L. Doctorow’s 1975 historical novel, is facing the possibility of censorship. The story, set in New York from t.
DANVILLE – Auditions will be opened up to the community in an attempt to fill 35 roles in the spring production of the musical, "Ragtime." The Danville Area. of his or her choice and also learn a s.


text:
Instability in Washington causes uncertainty on Wall Street
Senate Minority Leader Mitch McConnell (R-KY) (C) arrives at the U.S. Capitol October 15, 2013 in Washington, DC. The U.S. government shutdown is entering its fifteenth day as the U.S. Senate and House of Representatives remain gridlocked on funding the federal government.
Senate leaders say they're close to a deal that would re-open the government and raise the debt ceiling. But there's still some horse trading going on. Some of it does involve nips and tucks to President Obama's health care reform law. And there's a developing scenario where the Senate would agree to lift the federal limit on borrowing until early next year. One Republican lawmaker says the House would accept the Senate time frame.
But if revisiting the debt ceiling soon is a component of the resolution, what does that mean to the economy? Juli Niemann, analyst at Smith Moore and Company in St. Louis, says all the squabling on Capitol Hill is making consumers and business owners hesistant to spend -- and that could cause recession-like circumstances.

domain: economics


text:
- Last Activity:
- 18th Mar, 2018 at 8:59 PM
- 18th Jun, 2015
- Likes Received:
Structuring Lawyer and Finance Broker - all states, Male, from Remote
- Terry_w was last seen:
- Managing account details, 18th Mar, 2018 at 8:59 PM
- Home Page:
I have been a finance broker for about 16 years and a lawyer for 7 years now.
- Finance broker, Lawyer, Tax Advisor
I hold a specialist qualification in Tax - Chartered Tax Adviser and am probably the only person in Australia that can advise on structuring from a taxation, legal and lending aspects.
Although I am not currently licensed I am also qualified as a financial planner.
There are no messages on Terry_w's profile yet.

domain: law


An inside candidate has landed the gig as the newest showrunner on AMC’s “The Walking Dead.”
According to a variety of media reports, Scott M. Gimple will be the new “Walking Dead” showrunner, replacing Glen Mazzara, who will be departing after completing work on the show’s third season.
AMC is not confirming the hire.
Mazzara, who replaced original series developer Frank Darabont as showrunner early in the second season, left his showrunning gig simultaneous with the fourth season renewal of “The Walking Dead” last month.
Gimple currently serves as supervising producer on “The Walking Dead” and has written several episodes including the recent “Hounded.” Gimple previously worked with David Goyer on “FlashForward,” as well as on the upcoming “Da Vinci’s Demons” for Starz. His feature credits include “Ghost Rider: Spirit of Vengeance.”


text:
Members of the military need to take special steps to protect their interests before they get married. This is because marriages that last 20 years while the service member serves 20 years and 20 years of each of those overlap can lead to claims under the 20/20/20 rule if the couple gets divorced.
Because the 20/20/20 rule has to do with your military pension and other benefits, you need to get a prenuptial agreement signed before you walk down the aisle. You can include information about how the pension and other assets are handled if you get a divorce. It is understandable that you wouldn’t want to think about divorce before you get married, but doing so might save you from financial devastation down the road. Be sure you understand how to properly create and present a premarital agreement because you don’t want a mistake to have it found to be unenforceable if you need it.
You may need to make sure that you create a power of attorney that will allow your spouse to act on your behalf when you are deployed. If you do this, you can spell out what responsibilities you are turning over. You also need to add your spouse to your Page 2 so that he or she will be notified if there is an emergency, and you might need to update your Servicemembers’ Group Life Insurance policy.
Once you get married, you need to make plans to add your spouse to the Defense Enrollment Eligibility Reporting System. The DEERS enrollment can be done when your spouse gets his or her military ID made, and you can also register your spouse’s car with the installation at which you are stationed. Your spouse can also register for TRICARE and dental coverage.
Source: Military One Click, “The Essential Military Premarital Checklist,” accessed Sep. 19, 2016

domain: finance


text:
Are the leading drugs against Staphylococcus aureus really toxic to cartilage?
Şirin, Duygu Y.
MetadataShow full item record
CitationDoğan, M., İşyar, M., Yılmaz, İ., Bilir, B., Şirin, D. Y., Çakmak, S. ... Mahiroğulları, M. (2016). Are the leading drugs against Staphylococcus aureus really toxic to cartilage? Middle East Respiratory Syndrome Coronavirus (MERS-CoV) Research Initiative Workshop içinde (251-258. ss.). Riyadh, Saudi Arabia, September 09-10, 2015. https://dx.doi.org/10.1016/j.jiph.2015.10.004
Many studies have shown that the toxic effects of local antibiotics on bone and cartilage limit orthopedic surgeons. In this study, we evaluated three antibacterial agents used locally to treat highly mortal and morbid diseases in the field of orthopedics, such as septic arthritis. Are vancomycin, teicoplanin, and linezolid, which are archenemies of Staphylococcus aureus, really toxic to chondrocytes? The purpose of the study was to investigate the effects of antibiotics, which are used against S. aureus, on human chondrocytes in vitro. Primary cell cultures obtained from gonarthrosis patients were divided into two main groups. One of these groups was designated as the control chondrocyte culture. The other group was divided into three subgroups, and each group was exposedto vancomycin, teicoplanin, or linezolid. Cell culture samples were characterized by immunophenotyping following incubation with the three different antibiotics. Before and after the agents were administered, the cultures were subjected to inverted and environmental scanning electron microscopy. The number of live cells and the proliferation rate were monitored with the MTT-assay. We found that vancomycin, teicoplanin, and linezolid do not have chondrotoxic effects. Vancomycin, teicoplanin, and linezolid had no chondrotoxic activity during in vitro culture, which supports the argument that these agents can safely be used in orthopedic surgery, especially against methicillin-resistant S. aureus agents.

domain: biology


text:
Comparative analyses of vertebrate posterior HoxD clusters reveal atypical cluster architecture in the caecilian Typhlonectes natans
BMC Genomics volume 11, Article number: 658 (2010)
The posterior genes of the HoxD cluster play a crucial role in the patterning of the tetrapod limb. This region is under the control of a global, long-range enhancer that is present in all vertebrates. Variation in limb types, as is the case in amphibians, can probably not only be attributed to variation in Hox genes, but is likely to be the product of differences in gene regulation. With a collection of vertebrate genome sequences available today, we used a comparative genomics approach to study the posterior HoxD cluster of amphibians. A frog and a caecilian were included in the study to compare coding sequences as well as to determine the gain and loss of putative regulatory sequences.
We sequenced the posterior end of the HoxD cluster of a caecilian and performed comparative analyses of this region using HoxD clusters of other vertebrates. We determined the presence of conserved non-coding sequences and traced gains and losses of these footprints during vertebrate evolution, with particular focus on amphibians. We found that the caecilian HoxD cluster is almost three times larger than its mammalian counterpart. This enlargement is accompanied with the loss of one gene and the accumulation of repeats in that area. A similar phenomenon was observed in the coelacanth, where a different gene was lost and expansion of the area where the gene was lost has occurred. At least one phylogenetic footprint present in all vertebrates was lost in amphibians. This conserved region is a known regulatory element and functions as a boundary element in neural tissue to prevent expression of Hoxd genes.
The posterior part of the HoxD cluster of Typhlonectes natans is among the largest known today. The loss of Hoxd-12 and the expansion of the intergenic region may exert an influence on the limb enhancer, by having to bypass a distance seven times that of regular HoxD clusters. Whether or not there is a correlation with the loss of limbs remains to be investigated. These results, together with data on other vertebrates show that the tetrapod Hox clusters are more variable than previously thought.
Perhaps the best studied gene clusters in animals are the Hox clusters, not only for their importance in the establishment of the metazoan body plan, but also for their tight genomic organization. Hox genes encode transcription factors that belong to the family of homeodomain proteins and play an essential role in the establishment of the anterior-posterior body axis during embryonic development. In addition, they are also involved in patterning of limbs and in organogenesis [1–4]. In vertebrates, the expression domains of Hox genes are collinear in space and time, and reflect their chromosomal arrangement .
Invertebrates possess one - often interrupted or disintegrated - Hox cluster, while all vertebrates have multiple clusters [6–8]. Gnathostomes typically have four Hox clusters that arose by subsequent duplications in the stem lineage of vertebrates, while most ray-finned fishes contain seven (e.g. zebrafish) to thirteen (e.g. salmon) clusters as a result of additional, teleost-specific genome duplications and subsequent cluster losses [9–11]. Due to these additional duplications, the gene content of the fish Hox clusters is variable, with different gene losses in different species examined. In contrast, the Hox complement of tetrapods is rather conserved, with the same genes present in mammalian and bird genomes . In the genome of the frog Silurana tropicalis, at least one and possibly two genes have been lost [12, 13].
Hox gene clusters in vertebrates are compact (around 100 kb in mammals and even shorter in teleost fishes), with highly conserved distances between paralogous genes, and with little or no interspersed repetitive DNA elements . The only exceptions known so far are squamate reptiles, with the lizard Anolis carolinensis as a striking example of having accumulated a substantial number of retrotransposons in its Hox clusters, resulting in considerably larger cluster sizes [15, 16]. In general, the tight clustering of the Hox genes in vertebrates may be the result of an evolutionary constraint to keep the genes in close proximity, thus maintaining the intergenic distances and prohibiting insertion of interspersed repeats . This constraint may be facilitated by the presence of cis-regulatory elements within the clusters that are shared by neighboring genes, as well as by remote enhancers producing regulatory landscapes that would be broken when the clusters split . In this context, a suite of global long-range enhancers that control the expression of six genes located 5' of the HoxD cluster of fishes and mammals has been discovered . This Global Control Region (GCR) reinforces the effect of another enhancer, Prox, that drives the expression of the genes Lnp and Evx-2 - both adjacent to the 5' end of the HoxD cluster - and the posterior Hoxd genes (Hoxd-13 to Hoxd-10) in the distal limb and genital buds (digit enhancer); it also regulates the expression of Lnp and Evx-2 in the central nervous system (neural enhancer) [19–21]. The action of the GCR in the nervous system is somehow restricted by boundary elements between Evx-2 and Hoxd-13, while in the limb bud its effect decreases progressively with distance from the 5' end of the HoxD cluster [22–24].
Defects in Hoxd genes or gene regulation often have an effect on limb development. For example, the mouse Ulnaless mutation causes reduction of the zeugopod, which is the result of the alteration of Hoxd gene expression due to an inversion of the HoxD cluster and subsequent change of cis-regulatory control [19, 25, 26]. Similar phenotypes can be observed in human mesomelic dysplasia patients, which results from microduplications in the HoxD cluster . Normal limb development can also be affected by mutations within Hoxd genes. For example, the expansion of a poly-alanine tract in HOXD13 results in the synpolydactyly syndrome, with abnormal reductions, duplications and fusions of digits .
Despite their differences in limb types, frogs and salamanders are characterized by the presence of only four fingers. Of all amphibians, caecilians (Gymnophiona) probably form the most enigmatic order, as most of them spend their life hidden under the ground. Similar to snakes, they have an elongated trunk and have undergone secondary loss of limbs. The diversity in body plan and limbs among the three amphibian orders may have been affected by changes in Hox gene sequence, or, more likely, regulation (e.g. ).
At present, the only amphibian genome sequence publicly available is from the frog Silurana tropicalis. We constructed a BAC library of the aquatic caecilian Typhlonectes natans to obtain the sequence of the posterior end of the HoxD cluster, including Evx-2, i.e. the part of the cluster that is important in limb development and which has been shown to be under control of the Global Control Region. Comparative analyses of this region with the orthologous region of other vertebrates and the subsequent identification of conserved, non-coding, putative regulatory elements may shed light on the evolution of the caecilian body plan.
Results and Discussion
BAC sequencing and annotation
The haploid genome size of Typhlonectes natans was estimated at 13.37 pg by flow cytometry with chicken erythrocyte nuclei as an internal standard (data not shown). A pooled BAC library of about 460,000 clones with an average insert size of 107 kb from Typhlonectes natans was constructed, comprising a theoretical 3.8 × coverage of the genome, and screened by PCR to isolate a clone that contained the posterior HoxD cluster. A single clone with an estimated size of approximately 115 kb containing Hoxd-13 was sequenced using 454 sequencing technology  and over 15,000 reads were assembled into two supercontigs. The orientation of the two supercontigs was determined by sequencing of both BAC ends, and they were assembled into one final contig with a small gap arbitrarily set at 100 base pairs (bp) because the total sequence length is consistent with the estimated insert size. The total caecilian sequence comprises 116,633 bp, including a 100 bp gap (GenBank HQ398255). An initial blastx analysis and the software GenomeScan  identified the genes Evx-2, Hoxd-13, Hoxd-11 and a large part of Hoxd-10. The exon-intron boundaries were refined manually by alignment with Hox sequences of other vertebrates. Hoxd-12 was not found in these analyses.
Vertebrate posterior HoxD cluster comparison
We compared the posterior HoxD cluster of the caecilian with the clusters of the horn shark, zebrafish, coelacanth, frog, chicken, anole lizard, opossum, dog, mouse and human. In the caecilian, the 5' end of the HoxD cluster, starting from the stop codon of Evx-2, situated on the reverse strand, until the stop codon of Hoxd-10 is 107 kilo base pairs (kb), which is over 2.5 times larger than the orthologous region in mammals (approximately 40 kb) (Figure 1). Apart from the caecilian, the coelacanth and the lizard sequences are also larger than average, with lengths of 82 kb and 95 kb, respectively. The expansion of the caecilian HoxD cluster is mainly due to the lengthening of the intergenic region between Hoxd-13 and Hoxd-11, which is over six times larger than in the human. In the coelacanth, the intergenic region between Evx-2 and Hoxd-12, where the Hoxd-13 gene was lost, is almost four times longer than the corresponding region in mammals. In the anole lizard, not only the HoxD cluster, but also the other three Hox clusters are significantly longer .
Global alignment with the other vertebrates uncovered the remains of Hoxd-12 in the caecilian (Figure 2). The presence of multiple frameshift-producing indels and stop codons implies that it is no longer protein coding and has become a pseudogene (ψHox-d12). As frogs and caecilians comprise the basal split within amphibians , the most parsimonious explanation for the absence of Hoxd-12 from the Silurana tropicalis genome and its pseudogenization in the caecilian is an early loss in amphibian evolution with unequal rates of evolution in T. natans and S. tropicalis, although two independent loss events cannot be excluded. We also discovered a pseudogene (ψHoxd-13) in the coelacanth posterior HoxD sequence. A blastx analysis revealed fragments of both exons that could still be aligned with other vertebrate Hoxd-13 genes.
The length of the coding sequences of Hox genes is similar in both amphibian species, except for Hoxd-13, which is over 100 bp shorter in Typhlonectes. In general, most posterior HoxD coding sequences have comparable lengths in all species used in this study, except for Hoxd-13 and Hoxd-11, which are about 25% longer in amniotes and placental mammals, respectively. The amniote or mammalian expansion of the HOXD13 and HOXD11 proteins can be attributed to sequences rich in alanine, glycine, serine and proline. In HOXD13, a homopolymeric repeat consisting of nine alanines in the chicken and 15 alanines in mammals  is not present in any other species. HOXD13 proteins are overall rich in serine and alanine in mammals, but not in non-amniotes. In addition to the conservation of both exons of each Hox gene, evolutionarily conserved regions were also detected in non-coding intergenic and intronic sequences. These conserved, non-coding sequences or phylogenetic footprints may represent regulatory elements and non-coding RNA genes and were further investigated.
Distribution of repeats
To investigate whether the caecilian HoxD cluster expansion was accompanied by an accumulation of repeats, we compared it to the Repbase database  of known vertebrate repetitive elements using RepeatMasker  and Censor . RepeatMasker identified 4.74% of the sequence as part of transposable elements, while Censor reports 7.45% interspersed repeats, mostly retrotransposons like LINES and SINES. It must be noted that the hits are not always very strong because of short alignment length. However, blastn searches of all the repeats resulted in a significant match with known repeats (e-value ≤ 1e-05) for 11 out of 54 repeats. Moreover, blastx searches of the intergenic regions recovered one additional SINE and one LINE. The majority of the repeats is located in the intergenic region between Hoxd-13 and ψHoxd-12 (Figure 1). The same analyses were performed for the other vertebrate sequences by comparison with the repeat databases for the respective species, if available. In the caecilian and the coelacanth, the repeats are almost exclusively found in the large intergenic region where the pseudogene is situated, i.e. between Hoxd-13 and ψHoxd-12 in the caecilian and between Evx-2 and ψHoxd-13 in the coelacanth. In the lizard and zebrafish, transposable elements are dispersed over the cluster, with a concentration of repeats between Evx-2 and Hoxd-13 in the latter. Additionally, a self-self blastn analysis was performed to identify direct and inverted repeats that are not part of known transposons. This revealed the presence of 30 inverted repeats (stem-loop), 15 palindromes (stem) and 12 direct repeats (minimum identity of 70%, e-value ≤1e-5) in the caecilian HoxD cluster. Again, almost all repeats are located in the region between Hoxd-13 and the ψHoxd-12. Some of these repeats are a part of transposable elements, but the majority is unknown. A similar result was obtained for the coelacanth sequence. Here, no direct repeats, 21 inverted repeats and 11 palindromes were found, all located in the region between Evx-2 and Hoxd-12, with several large stretches in the former intron of ψHoxd-13. The anole lizard sequence also contains a large amount of inverted and direct repetitive sequences, but spread across the cluster, which is in concordance with the distribution of transposable elements. Few or no additional repeats were found in the other vertebrate HoxD sequences. The presence of interspersed, repetitive DNA may undermine genomic stability . In vertebrate genomes, regions containing developmental genes, such as Hox clusters, are usually devoid of transposable elements , suggesting the presence of a constraint against the invasion of foreign elements into a region essential for development. The caecilian, coelacanth, anole lizard and zebrafish have a higher than average amount of repetitive elements in the posterior HoxD cluster, and the caecilian and coelacanth also experienced the loss of a functional Hox gene. Whether the presence of repeats attributed to the pseudogenization of Hoxd-12 in the caecilian (and of Hoxd-13 in the coelacanth), or whether the loss of Hoxd-12 weakened the constraint and allowed repeats to accumulate, remains unknown. In the human genome, pseudogenes are frequently found in the vicinity of long inverted repeats .
Despite being essential for proper embryonic development, the loss of one Hox gene does not necessarily have a negative impact on an organism's phenotype, as Hox genes can be functionally equivalent . However, expansion of the cluster may have an effect on gene regulation. Since all the genes in this region of the HoxD cluster are under the control of the long-range enhancer GCR, the loss of Hoxd-12 accompanied with the enlargement of the distance between Hoxd-13 and Hoxd-11 and the accumulation of repetitive elements in the caecilian may have an influence on the effect of the GCR on the expression of Hoxd-11 and Hoxd-10. Whether this effect, if any, is reflected in morphology, is unknown, but it is worth mentioning that a similar cluster architecture was found in the corn snake . Some transposable elements have been exapted to modulate gene regulatory networks (reviewed by ). As changes in - especially developmental - gene regulation may lead to morphological changes , the adoption of a highly derived body plan, such as in caecilians, may have been facilitated by transposable elements. It is possible that caecilians and snakes have employed a similar mechanism of limb loss, though this is probably not the case in other limbless squamates, since Hoxd-12 is present in the slowworm Anguis fragilis .
Identification of phylogenetic footprints
More and more regulatory sequences and non-coding RNA (ncRNA) genes are being discovered in the portion of the genome that does not code for proteins . Although it has been shown that not every regulatory sequence is evolutionarily conserved and that an apparent function cannot always easily be allocated to a conserved sequence [43, 44], screening genomes for evolutionarily conserved non-coding sequences is a widely used strategy to discover potential regulatory elements. Such elements are expected to be present in the vicinity of transcription factors or developmental genes, such as Hox genes . We identified 33 evolutionarily conserved non-protein coding sequences, or so-called phylogenetic footprints, with the software tracker  (Additional file 1). To distinguish putatively transcribed footprints, we blast searched all footprints against the NCBI database of ESTs, which resulted in the identification of eight footprints between 29 and 876 bp long, not located within untranslated regions (UTRs), with one or more EST matches (Additional file 2). Because EST data are absent for many organisms, including coelacanth and caecilians, we did not find ESTs corresponding to these footprints for every organism. However, given the high degree of sequence conservation and the finding of ESTs of each footprint in at least two different organisms, we believe it is possible that transcription and perhaps the function of these elements are conserved. Therefore, we consider these footprints to be putative ncRNA genes, and the other, non-transcribed footprints as potential cis-regulatory elements (Figure 1).
Non-coding RNAs are functional molecules that are not translated into a protein. Instead, they are involved in post-transcriptional modification or DNA replication or have a regulatory function, and can be found in intergenic regions, introns and even in the UTR of genes or overlapping with protein coding genes . The gene regulatory RNAs are usually small and act in trans by post-transcriptional silencing of target genes through the binding of complementary sites. Next to the large number of small ncRNAs, an increasing number of long ncRNAs is being described, which are at least 200 to over 10,000 nucleotides long (reviewed by ). Long ncRNAs can act independently of a target sequence, in cis, by interfering with the transcription of a neighboring gene, or in trans, by recruiting proteins that alter the chromatin state . One example of a regulatory ncRNA in the HoxC cluster is HOTAIR, which epigenetically represses transcription of 40 kb across the HoxD locus . Screening of all the expressed footprints against the Functional RNA Database  did not result in the identification of any classified ncRNA. However, four of them produced significant matches with putative RNAs predicted by Evofold, which is a method to identify functional RNA structures in vertebrates by using a combined probabilistic model of RNA structure and sequence evolution . In addition, in footprint fp1, also known as CR3 , a significant RNA secondary structure was predicted (p = 0.92). Seven of the expressed footprints lie in the intergenic region between Hoxd-11 and Hoxd-10 (Figure 1). What tracker considers to be multiple footprints may correspond to a single potential ncRNA gene, as indicated by alignment of the region between Hoxd-11 and Hoxd-10 with the corresponding ESTs (not shown). Two of the expressed footprints were previously identified as regulatory regions RVIII/RIX (fp26) and RX (fp20) ([52–55]. The fact that these two regulatory regions appear to be transcribed may shed new light on how they function.
Cis-regulatory elements can be anticipated in intergenic regions, in introns and in the 5' and 3' UTR of protein-coding genes, and may act as a promoter, enhancer, repressor or insulator . In total, 24 footprints (fps) are putative cis-regulatory elements. Nine of these were found in the intergenic regions, including the area downstream of Evx-2; four footprints are located in an intron, five located in the 5' UTR and six located in the 3' UTR of the Hox and Evx-2 genes. Three of the intergenic footprints were identified in previous studies and are known as RXI (fp18) and RXII (fp9 and fp10) [23, 56]. Two other intergenic footprints (fp 2 and fp3) are extremely conserved in all species used in this study and are located downstream Evx-2. The reason for this high degree of conservation however remains unclear . One footprint (fp23) that was found in the intron of Hoxd-11 contains a HB1 element, which consists of homeodomain binding sites. This element is previously described from the intron of Hoxa7, the Drosophila homolog Ubx, the introns of Hox4 genes and the intron of Hoxa-11 [57–60]. Of the five footprints in the 5'UTR, two also include the promoter region. The 5' and 3' UTRs of mRNAs play an important role in the post-transcriptional regulation of gene expression through the presence of cis-acting elements and through interaction with micro-RNAs (miRNAs) [61, 62]. Conserved sequences in 3' UTRs may contain potential target sites for miRNAs that are involved in post-transcriptional gene silencing. Micro-RNAs are short, single-stranded RNA molecules of ~22 nucleotides that show at least partial complementarities to their target mRNA. If the miRNA is only partially complementary, a perfect match between the seed (nucleotides 2 to 7) of the miRNA and the target mRNA is necessary for inhibition of translation or for promotion of deadenylation [63, 64]. Therefore all six footprints located in the 3'UTR of the genes were screened for the presence of hypothetical target sites for miRNAs. One footprint (fp15), located in the 3'UTR of Hoxd-13, contains a short motif of seven nucleotides, with perfect match to the seed of the miRNA miR-26.
Reconstruction of conserved element evolution
Gains and losses of putative ncRNAs and cis-regulatory elements in vertebrate posterior HoxD clusters were mapped on a vertebrate timescale under the Dollo parsimony criterion, i.e. assuming a single origin (Figure 3). Two putative cis-regulatory elements have been lost in the amphibian ancestor: fp9, which is one of two motifs of RXII, and fp15, a small footprint in the 3' UTR of Hoxd-13. The conserved region RXII is a boundary element located in the intergenic region between Evx-2 and Hoxd-13, whose promoters are in each other's vicinity. Unlike Hoxd genes, Evx-2 is expressed in the central nervous system, regulated by the neural enhancer of the GCR. RXII is considered to function as an insulator to prevent the ectopic expression of Hoxd genes in the nervous system, since GCR regulation is not promoter-specific . Although tracker failed to recover the entire RXII element (fp9 and fp10) in the zebrafish, short, apparently homologous sequences were found in the zebrafish cluster in the same area as in other vertebrates . Moreover, both footprints were found in the horn shark, which points to their ancestral presence in vertebrates, and therefore we conclude that fp9 was lost in amphibians. Footprint 15 is a short DNA stretch of ~22 bp with a highly conserved motif of 7 bp located in the 3' UTR of Hoxd-13. This motif is somewhat degenerated in the coelacanth, which does not have a functional Hoxd-13 gene, and it is located around 14 kb 5' to ψHoxd-13, which may indicate that this sequence is not homologous to the other vertebrate fp15 sequences. On the other hand, pseudogenization of Hoxd-13 and insertion of repetitive sequences may have promoted the relative relocation of this element. For our reconstruction, we considered all footprints as identified by the tracker software to be truly conserved sequences. Therefore, we conclude that this element originated in Sarcopterygii, and was lost in amphibians. In theory, this footprint can serve as a target for the microRNA miR-26, which is expressed in neurons and astrocytes of the developing mouse brain . The absence of both elements may indicate that restriction of Hoxd-13 expression in neural tissue is regulated differently in amphibians.
The posterior HoxD cluster of the caecilian is much larger than that of most known vertebrates due to the accumulation of interspersed and inverted repeats accompanied with Hoxd-12 gene loss. A similar event occurred in the coelacanth and in the corn snake HoxD cluster. Whether these occurrences are reflected in the morphology of these species is not clear, but it is possible that caecilians and snakes adopted a similar mechanism that resulted in body elongation and limb loss. The distance created between two subsequent genes that are under the control of the same long-range limb enhancer is likely to have some effect on the action of this enhancer on the genes after the void. A number of conserved, non-coding regions have been identified in the HoxD cluster, some of them showing extremely high conservation among all vertebrates. While no new footprints arose in the amphibian lineage, at least one, and possibly two conserved sequences were lost. These results, together with data on other vertebrate Hox clusters show that tetrapod Hox clusters show more variation than expected previously.
Genomic library and DNA sequencing
High molecular weight DNA was extracted from erythrocytes from two Typhlonectes natans specimens and was used to construct a BAC library according to Osoegawa et al.  and as described by Danke et al. . The genome size was estimated by flow cytometry analysis using chicken erythrocyte nuclei as a standard. EcoRI partial digests of the caecilian DNA were size selected and inserted in the pCC1 BAC vector (Epicentre) and the library was combined in 2304 pools containing 200 clones each. The library was screened by PCR with primers specific for Hoxd-13 (forward primer: 5'-GCAATGAAGGCGCCTCCAG-3', reverse primer: 5'-GGAGATATAGGTGTCGTGCCTCGG-3') to isolate the posterior end of the HoxD cluster. The positive clone was 454 GS FLX-sequenced and assembled by Eurofins MWG Operon (Ebersberg, Germany). The clone was also end sequenced using BigDye 3.1 chemistry on an ABI 3100 Genetic Analyzer. Several smaller contigs were assembled manually and contig overlaps were confirmed by PCR, except in one case where a gap was present. Because the length of the two contigs is consistent with the size of the insert, we inferred this gap to be small and therefore arbitrarily set it at 100 bp.
Caecilian HoxD cluster annotation
The genes in the caecilian posterior HoxD cluster were annotated by initial blastx searches of the entire cluster sequence and with the program GenomeScan  using mouse HOXD and EVX2 proteins as a training set. Exon - intron boundaries were determined manually by alignment with Hox sequences of other vertebrates.
Global alignments of the caecilian posterior HoxD cluster with other vertebrate HoxD clusters were performed with MultiPipMaker  and VISTA . The HoxD clusters of the following species were used: Heterodontus francisci (horn shark, AF224263), Danio rerio (zebrafish, UCSC Genome Browser) Latimeria menadoensis (Indonesian coelacanth, FJ497008), Silurana tropicalis (tropical clawed frog, JGI), Gallus gallus (chicken, ENSEMBL), Anolis carolinensis (green anole, UCSC Genome Browser), Monodelphis domestica (grey short-tailed opossum, ENSEMBL), Canis familiaris (domestic dog, ENSEMBL), Homo sapiens (human, NT_005403) and Mus musculus (mouse, AC_015584). The MultiPipMaker alignments were performed with mouse and T. natans as reference sequence respectively.
Interspersed repeats and low complexity regions in all clusters were masked by screening against a library of repetitive elements if available for the organism by RepeatMasker . If no such library was available, as for the horn shark, coelacanth, caecilian, lizard and opossum, the sequences were compared to a database of transposable element encoded proteins.
The repeat content of the HoxD clusters was determined with Censor  and RepeatMasker, using the Repbase library of the species-specific or vertebrate repeats and a database of transposable element encoded proteins. All reported interspersed repeats were taken into account. In addition, self-self blastn analyses were performed to identify direct and inverted repeats within each cluster. Only repeats with maximum 30% mismatch and an e-value ≤ 1e-05 were retained.
Analyses of phylogenetic footprints
The program tracker  was used to detect evolutionarily conserved non-coding sequences or phylogenetic footprints. This program is based on blastz  to produce initial local pairwise alignments of all pairs of the input sequences. Only the intergenic regions between two homologous genes are compared. After several filtering steps, these alignments, which contain a window of 12 nucleotides with minimum identity of 75%, are assembled into groups of partially overlapping regions, resulting in local sequence alignments, or footprint cliques. To be able to detect conserved sequences in all non-coding regions, introns were treated as intergenic regions. This analysis was repeated using the same, but repeat masked sequences and all footprints from both analyses were combined. Not every footprint is necessarily conserved in each taxon, as its presence or absence may be indicative of the loss of a cis-regulatory element and a subsequent change in gene regulation. Phylogenetic footprints were treated as two classes: potential (cis-) regulatory elements and putative non-coding RNA genes. Conserved sequences found in the 3' UTR of genes were screened for the presence of hypothetical target sites of miRNAs using TargetScan Release 5.1 [63, 71]. To detect whether footprints are expressed and thus may be ncRNA genes, a blastn search of all footprints against the NCBI database of ESTs was performed. Footprints with one or more EST matches were screened against the Functional RNA Database  for similarity with known ncRNAs. In addition, the RNAz server (Vienna RNA server, http://rna.tbi.univie.ac.at) was used to identify thermodynamically stable and evolutionarily conserved RNA secondary structures in the footprint alignments.
A Dollo parsimony reconstruction of the genes and footprints was done with MacClade v4.06  to assess whether putative cis-regulatory elements and ncRNAs were lost or gained during vertebrate evolution.
McGinnis W, Krumlauf R: Homeobox genes and axial patterning. Cell. 1992, 68 (2): 283-302. 10.1016/0092-8674(92)90471-N.
Zakany J, Duboule D: The role of Hox genes during vertebrate limb development. Curr Opin Genet Dev. 2007, 17 (4): 359-366. 10.1016/j.gde.2007.05.011.
Kondo T, Dollé P, Zákány J, Duboule D: Function of posterior HoxD genes in the morphogenesis of the anal sphincter. Development. 1996, 122: 2651-2659.
Di-Poï N, Zákány J, Duboule D: Distinct roles and regulations for Hoxd genes in metanephric kidney development. PLoS Genet. 2007, 3 (12): e232-10.1371/journal.pgen.0030232.
Kmita M, Duboule D: Organizing axes in time and space; 25 years of colinear tinkering. Science. 2003, 301 (5631): 331-333. 10.1126/science.1085753.
Lemons D, McGinnis W: Genomic evolution of Hox gene clusters. Science. 2006, 313 (5795): 1918-1922. 10.1126/science.1132040.
Powers TP, Amemiya CT: Evolutionary plasticity of vertebrate Hox genes. Current Genomics. 2004, 5 (6): 459-472. 10.2174/1389202043349048.
Seo HC, Edvardsen RB, Maeland AD, Bjordal M, Jensen MF, Hansen A, Flaat M, Weissenbach J, Lehrach H, Wincker P, et al: Hox cluster disintegration with persistent anteroposterior order of expression in Oikopleura dioica. Nature. 2004, 431 (7004): 67-71. 10.1038/nature02709.
Bailey WJ, Kim J, Wagner GP, Ruddle FH: Phylogenetic reconstruction of vertebrate Hox cluster duplications. Mol Biol Evol. 1997, 14 (8): 843-853.
Meyer A, Van de Peer Y: From 2R to 3R: evidence for a fish-specific genome duplication (FSGD). Bioessays. 2005, 27 (9): 937-945. 10.1002/bies.20293.
Mungpakdee S, Seo HC, Angotzi AR, Dong X, Akalin A, Chourrout D: Differential evolution of the 13 Atlantic salmon Hox clusters. Mol Biol Evol. 2008, 25 (7): 1333-1343. 10.1093/molbev/msn097.
Hoegg S, Meyer A: Hox clusters as models for vertebrate genome evolution. Trends Genet. 2005, 21 (8): 421-424. 10.1016/j.tig.2005.06.004.
Mannaert A, Roelants K, Bossuyt F, Leyns L: A PCR survey for posterior Hox genes in amphibians. Mol Phylogenet Evol. 2006, 38 (2): 449-458. 10.1016/j.ympev.2005.08.012.
Fried C, Prohaska SJ, Stadler PF: Exclusion of repetitive DNA elements from gnathostome Hox clusters. J Exp Zoolog B Mol Dev Evol. 2004, 302 (2): 165-173.
Di-Poï N, Montoya-Burgos JI, Duboule D: Atypical relaxation of structural constraints in Hox gene clusters of the green anole lizard. Genome Res. 2009, 19 (4): 602-610. 10.1101/gr.087932.108.
Di-Poï N, Montoya-Burgos JI, Miller H, Pourquié O, Milinkovitch MC, Duboule D: Changes in Hox genes' structure and function during the evolution of the squamate body plan. Nature. 2010, 464 (7285): 99-103. 10.1038/nature08789.
Duboule D: The rise and fall of Hox gene clusters. Development. 2007, 134 (14): 2549-2560. 10.1242/dev.001065.
van der Hoeven F, Zakany J, Duboule D: Gene transpositions in the HoxD complex reveal a hierarchy of regulatory controls. Cell. 1996, 85 (7): 1025-1035. 10.1016/S0092-8674(00)81303-3.
Spitz F, Gonzalez F, Duboule D: A global control region defines a chromosomal regulatory landscape containing the HoxD cluster. Cell. 2003, 113 (3): 405-417. 10.1016/S0092-8674(03)00310-6.
Gonzalez F, Duboule D, Spitz F: Transgenic analysis of Hoxd gene regulation during digit development. Dev Biol. 2007, 306 (2): 847-859. 10.1016/j.ydbio.2007.03.020.
Spitz F, Duboule D: Global control regions and regulatory landscapes in vertebrate development and evolution. Adv Genet. 2008, 61: 175-205. full_text.
Kmita M, Fraudeau N, Herault Y, Duboule D: Serial deletions and duplications suggest a mechanism for the collinearity of Hoxd genes in limbs. Nature. 2002, 420 (6912): 145-150. 10.1038/nature01189.
Kmita M, Tarchini B, Duboule D, Herault Y: Evolutionary conserved sequences are required for the insulation of the vertebrate Hoxd complex in neural cells. Development. 2002, 129 (23): 5521-5528. 10.1242/dev.00151.
Monge I, Kondo T, Duboule D: An enhancer-titration effect induces digit-specific regulatory alleles of the HoxD cluster. Dev Biol. 2003, 256 (2): 212-220. 10.1016/S0012-1606(02)00136-7.
Herault Y, Fraudeau N, Zakany J, Duboule D: Ulnaless (Ul), a regulatory mutation inducing both loss-of-function and gain-of-function of posterior Hoxd genes. Development. 1997, 124 (18): 3493-3500.
Peichel CL, Prabhakaran B, Vogt TF: The mouse Ulnaless mutation deregulates posterior HoxD gene expression and alters appendicular patterning. Development. 1997, 124 (18): 3481-3492.
Cho TJ, Kim OH, Choi IH, Nishimura G, Superti-Furga A, Kim KS, Lee YJ, Park WY: A dominant mesomelic dysplasia associated with a 1.0-Mb microduplication of HOXD gene cluster at 2q31.1. J Med Genet. 2010, 47 (9): 638-639. 10.1136/jmg.2009.074690.
Muragaki Y, Mundlos S, Upton J, Olsen BR: Altered growth and branching patterns in synpolydactyly caused by mutations in HOXD13. Science. 1996, 272 (5261): 548-551. 10.1126/science.272.5261.548.
Vinagre T, Moncaut N, Carapuco M, Novoa A, Bom J, Mallo M: Evidence for a myotomal Hox/Myf cascade governing nonautonomous control of rib specification within global vertebral domains. Dev Cell. 2010, 18 (4): 655-661. 10.1016/j.devcel.2010.02.011.
Margulies M, Egholm M, Altman WE, Attiya S, Bader JS, Bemben LA, Berka J, Braverman MS, Chen YJ, Chen Z, et al: Genome sequencing in microfabricated high-density picolitre reactors. Nature. 2005, 437 (7057): 376-380.
Yeh RF, Lim LP, Burge CB: Computational inference of homologous gene structures in the human genome. Genome Res. 2001, 11 (5): 803-816. 10.1101/gr.175701.
Zardoya R, Meyer A: On the origin of and phylogenetic relationships among living amphibians. Proc Natl Acad Sci USA. 2001, 98 (13): 7380-7383. 10.1073/pnas.111455498.
Jurka J, Kapitonov VV, Pavlicek A, Klonowski P, Kohany O, Walichiewicz J: Repbase Update, a database of eukaryotic repetitive elements. Cytogenet Genome Res. 2005, 110 (1-4): 462-467. 10.1159/000084979.
Smit AFA, Hubley R, Green P: Repeatmasker Open-3.0. 1996, [http://www.repeatmasker.org]
Kohany O, Gentles AJ, Hankus L, Jurka J: Annotation, submission and screening of repetitive elements in Repbase: RepbaseSubmitter and Censor. BMC Bioinformatics. 2006, 7: 474-10.1186/1471-2105-7-474.
Gordenin DA, Lobachev KS, Degtyareva NP, Malkova AL, Perkins E, Resnick MA: Inverted DNA repeats: a source of eukaryotic genomic instability. Molecular and Cellular Biology. 1993, 13 (9): 5315-5322.
Simons C, Makunin IV, Pheasant M, Mattick JS: Maintenance of transposon-free regions throughout vertebrate evolution. BMC Genomics. 2007, 8: 470-10.1186/1471-2164-8-470.
Wang Y, Leung FCC: A study on genomic distribution and sequence features of human long inverted repeats reveals species-specific intronic inverted repeats. FEBS Journal. 2009, 276 (7): 1986-1998. 10.1111/j.1742-4658.2009.06930.x.
Wellik DM, Capecchi MR: Hox10 and Hox11 genes are required to globally pattern the mammalian skeleton. Science. 2003, 301 (5631): 363-367. 10.1126/science.1085672.
Bourque G: Transposable elements in gene regulation and in the evolution of vertebrate genomes. Curr Opin Genet Dev. 2009, 19 (6): 607-612. 10.1016/j.gde.2009.10.013.
Carroll SB: Chance and necessity: the evolution of morphological complexity and diversity. Nature. 2001, 409 (6823): 1102-1109. 10.1038/35059227.
Elgar G, Vavouri T: Tuning in to the signals: noncoding sequence conservation in vertebrate genomes. Trends Genet. 2008, 24 (7): 344-352. 10.1016/j.tig.2008.04.005.
Ahituv N, Zhu Y, Visel A, Holt A, Afzal V, Pennacchio LA, Rubin EM: Deletion of ultraconserved elements yields viable mice. PLoS Biol. 2007, 5 (9): e234-10.1371/journal.pbio.0050234.
McGaughey DM, Vinton RM, Huynh J, Al-Saif A, Beer MA, McCallion AS: Metrics of sequence constraint overlook regulatory sequences in an exhaustive analysis at phox2b. Genome Res. 2008, 18 (2): 252-260. 10.1101/gr.6929408.
Prohaska SJ, Fried C, Flamm C, Wagner GP, Stadler PF: Surveying phylogenetic footprints in large gene clusters: applications to Hox cluster duplications. Mol Phylogenet Evol. 2004, 31 (2): 581-604. 10.1016/j.ympev.2003.08.009.
Mattick JS: The genetic signatures of noncoding RNAs. PLoS Genet. 2009, 5 (4): e1000459-10.1371/journal.pgen.1000459.
Mercer TR, Dinger ME, Mattick JS: Long non-coding RNAs: insights into functions. Nat Rev Genet. 2009, 10 (3): 155-159. 10.1038/nrg2521.
Rinn JL, Kertesz M, Wang JK, Squazzo SL, Xu X, Brugmann SA, Goodnough LH, Helms JA, Farnham PJ, Segal E, et al: Functional demarcation of active and silent chromatin domains in human HOX loci by noncoding RNAs. Cell. 2007, 129 (7): 1311-1323. 10.1016/j.cell.2007.05.022.
Mituyama T, Yamada K, Hattori E, Okida H, Ono Y, Terai G, Yoshizawa A, Komori T, Asai K: The Functional RNA Database 3.0: databases to support mining and annotation of functional RNAs. Nucleic Acids Res. 2009, D89-92. 10.1093/nar/gkn805. 37 Database
Pedersen JS, Bejerano G, Siepel A, Rosenbloom K, Lindblad-Toh K, Lander ES, Kent J, Miller W, Haussler D: Identification and classification of conserved RNA secondary structures in the human genome. PLoS Comput Biol. 2006, 2 (4): e33-10.1371/journal.pcbi.0020033.
Sabarinadh C, Subramanian S, Tripathi A, Mishra RK: Extreme conservation of noncoding DNA near HoxD complex of vertebrates. BMC Genomics. 2004, 5 (1): 75-10.1186/1471-2164-5-75.
Gérard M, Duboule D, Zakany J: Structure and activity of regulatory elements involved in the activation of the Hoxd-11 gene during late gastrulation. Embo J. 1993, 12 (9): 3539-3550.
Gérard M, Chen JY, Gronemeyer H, Chambon P, Duboule D, Zakany J: In vivo targeted mutagenesis of a regulatory element required for positioning the Hoxd-11 and Hoxd-10 expression boundaries. Genes Dev. 1996, 10 (18): 2326-2334. 10.1101/gad.10.18.2326.
Beckers J, Gerard M, Duboule D: Transgenic analysis of a potential Hoxd-11 limb regulatory element present in tetrapods and fish. Dev Biol. 1996, 180 (2): 543-553. 10.1006/dbio.1996.0327.
Beckers J, Duboule D: Genetic analysis of a conserved sequence in the HoxD complex: regulatory redundancy or limitations of the transgenic approach?. Dev Dyn. 1998, 213 (1): 1-11. 10.1002/(SICI)1097-0177(199809)213:1<1::AID-AJA1>3.0.CO;2-L.
Hérault Y, Beckers J, Kondo T, Fraudeau N, Duboule D: Genetic analysis of a Hoxd-12 regulatory element reveals global versus local modes of controls in the HoxD complex. Development. 1998, 125 (9): 1669-1677.
Morrison A, Chaudhuri C, Ariza-McNaughton L, Muchamore I, Kuroiwa A, Krumlauf R: Comparative analysis of chicken Hoxb-4 regulation in transgenic mice. Mech Dev. 1995, 53 (1): 47-59. 10.1016/0925-4773(95)00423-8.
Haerry TE, Gehring WJ: Intron of the mouse Hoxa-7 gene contains conserved homeodomain binding sites that can function as an enhancer element in Drosophila. Proc Natl Acad Sci USA. 1996, 93 (24): 13884-13889. 10.1073/pnas.93.24.13884.
Haerry TE, Gehring WJ: A conserved cluster of homeodomain binding sites in the mouse Hoxa-4 intron functions in Drosophila embryos as an enhancer that is directly regulated by Ultrabithorax. Dev Biol. 1997, 186 (1): 1-15.
Santini S, Boore JL, Meyer A: Evolutionary conservation of regulatory elements in vertebrate Hox gene clusters. Genome Res. 2003, 13 (6A): 1111-1122. 10.1101/gr.700503.
Mignone F, Gissi C, Liuni S, Pesole G: Untranslated regions of mRNAs. Genome Biol. 2002, 3 (3): REVIEWS0004-10.1186/gb-2002-3-3-reviews0004.
Flynt AS, Lai EC: Biological principles of microRNA-mediated regulation: shared themes amid diversity. Nat Rev Genet. 2008, 9 (11): 831-842. 10.1038/nrg2455.
Lewis BP, Shih IH, Jones-Rhoades MW, Bartel DP, Burge CB: Prediction of mammalian microRNA targets. Cell. 2003, 115 (7): 787-798. 10.1016/S0092-8674(03)01018-3.
Bartel DP: MicroRNAs: genomics, biogenesis, mechanism, and function. Cell. 2004, 116 (2): 281-297. 10.1016/S0092-8674(04)00045-5.
Smirnova L, Grafe A, Seiler A, Schumacher S, Nitsch R, Wulczyn FG: Regulation of miRNA expression during neural cell specification. Eur J Neurosci. 2005, 21 (6): 1469-1477. 10.1111/j.1460-9568.2005.03978.x.
Osoegawa K, Woon PY, Zhao B, Frengen E, Tateno M, Catanese JJ, de Jong PJ: An improved approach for construction of bacterial artificial chromosome libraries. Genomics. 1998, 52 (1): 1-8. 10.1006/geno.1998.5423.
Danke J, Miyake T, Powers T, Schein J, Shin H, Bosdet I, Erdmann M, Caldwell R, Amemiya CT: Genome resource for the Indonesian coelacanth, Latimeria menadoensis. J Exp Zoolog A Comp Exp Biol. 2004, 301 (3): 228-234.
Schwartz S, Elnitski L, Li M, Weirauch M, Riemer C, Smit A, Green ED, Hardison RC, Miller W: MultiPipMaker and supporting tools: Alignments and analysis of multiple genomic DNA sequences. Nucleic Acids Res. 2003, 31 (13): 3518-3524. 10.1093/nar/gkg579.
Mayor C, Brudno M, Schwartz JR, Poliakov A, Rubin EM, Frazer KA, Pachter LS, Dubchak I: VISTA: visualizing global DNA sequence alignments of arbitrary length. Bioinformatics. 2000, 16 (11): 1046-1047. 10.1093/bioinformatics/16.11.1046.
Schwartz S, Zhang Z, Frazer KA, Smit A, Riemer C, Bouck J, Gibbs R, Hardison R, Miller W: PipMaker--a web server for aligning two genomic DNA sequences. Genome Res. 2000, 10 (4): 577-586. 10.1101/gr.10.4.577.
Friedman RC, Farh KK, Burge CB, Bartel DP: Most mammalian mRNAs are conserved targets of microRNAs. Genome Res. 2009, 19 (1): 92-105. 10.1101/gr.082701.108.
Maddison WP, Maddison DR: MacClade, analysis of phylogeny and character evolution, Version 4.0. 2000
Alfaro ME, Santini F, Brock C, Alamillo H, Dornburg A, Rabosky DL, Carnevale G, Harmon LJ: Nine exceptional radiations plus high turnover explain species diversity in jawed vertebrates. Proc Natl Acad Sci USA. 2009, 106 (32): 13410-13414. 10.1073/pnas.0811087106.
We thank Joel Sohn for help with the caecilians, Andrew Stuart and the other members of the Amemiya Lab for technical assistance and Aru for genome size measurements. We are grateful to Kim Roelants for the drawings and to Wolfgang Otto for help with the tracker software. AM was supported by a Travel Grant of the Research Foundation - Flanders (FWO) for her stay in Seattle. FB and AM received financial support from the Vrije Universiteit Brussel (Concerted Research Action 29).
The research was designed by AM and FB. AM conducted the laboratory work, except the 454 sequencing; CTA provided assistance with the BAC library construction. AM performed the analyses. All authors wrote the manuscript.
Electronic supplementary material
Additional file 1: Summary of the conserved sequences in vertebrate Hox clusters. This table gives an overview of the positions of the coding regions in the vertebrate sequences as well as all footprint positions identified by the tracker software. The footprints are named fp1 to fp33, and footprints that are expressed in at least two species are indicated in bold. (XLS 26 KB)
Additional file 2: EST blast hits of expressed footprints. All ESTs that correspond to the expressed footprints are given with Genbank accession number, species name and tissue source. (XLS 22 KB)
About this article
Cite this article
Mannaert, A., Amemiya, C.T. & Bossuyt, F. Comparative analyses of vertebrate posterior HoxD clusters reveal atypical cluster architecture in the caecilian Typhlonectes natans. BMC Genomics 11, 658 (2010). https://doi.org/10.1186/1471-2164-11-658


Shah Rukh Khan says 'dara mat' to a Twitter user who threatens to be Gaurav from Fan, latter gets his reply framed
From his upcoming movie Pathaan, his favorite team at the FIFA World Cup to his personal life, Shah Rukh Khan recently answered all the questions asked by his fans on Twitter.
Shah Rukh Khan recently interacted with his fans on Twitter in a freewheeling Q&A chat session. The actor answered several questions about his upcoming movie Pathaan , his favorite team at the FIFA World Cup, and a lot more about his personal life. Amid this, one of the Twitter users asked him to reply to his question warning him that he will turn Gaurav, a character from his 2016 movie Fan who was obsessed with him in the movie.
The user tweeted, “Fan ka Gaurav banne ka chance mat do sir, reply dedo jaldi.” Replying to this Shah Rukh Khan said, “Dara Mat!” Well, the superstar’s reply to his tweet made his dream come true and he went on to save his tweet in a glass frame. Sharing the picture, he wrote, “Ab khan saab reply diye or use hum frame na kawaye.. yeh kabhi ni ho sakta Framing done.. will keep it till the end” Check the post here:
During his chat session, Shah Rukh revealed his favorite Salman Khan movie to be Bajrangi Bhaijaan. Someone also asked him why he doesn't make films like 'Chak De! India' and 'Swades' anymore. The actor had an epic reply, "Bana toh do kitni baar banaoon (Have made two such films, how many times should I do it)." Well, winding up the session, he wrote, “Now my team calling me to work. Will talk with you all some other day. Those who missed out please don't feel bad....abhi picture baaki hai. Thanks for the love & ur time. See u soon in a theatre now...," he tweeted.
Meanwhile, Shah Rukh will be making his comeback in movies after four years with Pathaan. The film also stars Deepika Padukone and John Abraham. Recently, the first song Besharam Rang from the movie hit headlines for being too bold. Directed by Siddharth Anand, the film is set to hit theatre on January 25, 2023.


text:
Health groups are demanding more policy action to reduce the glaring health inequities shown by the latest New Zealand Health Survey. The harms created by tobacco, alcohol and unhealthy food continue to disproportionally burden Maori, Pacific and low income communities, with no signs of improvement.
"Communities are tired of hearing about these terrible statistics year after year while successive governments offer platitudes instead of policies," said Professor Boyd Swinburn, Chairperson of the Health Coalition Aotearoa. "If we aspire to be the best country in the world to be a child, then why are we so afraid to control the harmful products that are creating one third of our health problems? Children need healthy environments to grow up in and thrive."
Overall, smoking is now down to 14% but at the current rate of slow decline, the Government’s Smokefree 2025 goal of less than 5% smokers will not be reached until 2036 for the general population. Maori smoking prevalence, at 34%, will not reach the 2025 goal until 2044. Pacific smoking prevalence has remained unchanged over 8 years at 24%. Implementing the comprehensive tobacco control strategies recommended by the World Health Organisation and the Health Coalition experts will accelerate progress towards the Smokefree 2025 goal.
One in five New Zealanders over 14 is classified as a hazardous drinker, a figure unchanged over the past 5 years. David (Rawiri) Ratu, who co-chairs the Health Coalition’s Alcohol Expert Panel, points out "These figures show hazardous drinking continues to be experienced more by Maori than non-Maori. The government has so far failed to protect Maori from the over-supply of alcohol from the numerous outlets in poorer neighbourhoods and the marketing of alcohol using sports celebrities who are heroes to rangatahi".
Two in three adults are overweight or obese and this has not changed since 2011. For children, it also remains static with one in three being overweight or obese. The large disparities are also unchanged, exposing Maori, Pacific and low income New Zealanders to much greater risks of diabetes, heart disease and cancer.
Dr Lisa Te Morenga, who chairs the Health Coalition’s Food Policy Expert Panel, said "Efforts to reduce health inequities in Aotearoa New Zealand will be futile if serious policy action is not taken to support people to eat well. We urgently need healthy food policies in schools, limits on marketing of junk foods to children and their families, and health levies on the manufacturers of unhealthy food and drinks."
The Health Coalition Aotearoa is an alliance of over 50 health groups with extensive expertise on how to improve the well-being of New Zealanders and they have identified the most effective solutions to these enormously costly health problems. They are calling for the Government to apply its ‘COVID-19 approach’ to the damage caused by tobacco, alcohol and unhealthy food - an approach they describe as: look at the evidence, listen to the experts, and act boldly and decisively in the health interests of all New Zealanders, but especially for those communities which are suffering the most.

domain: sociology


text:
The European Union reached agreement early on Wednesday on new rules designed to share out the cost and work of hosting migrants more evenly and to limit the numbers of people coming in, Reuters has reported.
Representatives of the European Parliament and EU national governments reached an accord after all-night talks on EU laws collectively called the New Pact on Migration and Asylum that should start taking effect next year. The laws cover screening irregular migrants when they arrive in the European Union, procedures for handling asylum applications, rules on determining which EU country is responsible for handling applications and ways to handle crises.
Migrant arrivals in the EU are way down from the 2015 peak of more than 1 million, but have crept up steadily from a 2020 low to 255,000 in the year to November, with more than half crossing the Mediterranean from Africa, mainly to Italy.
Italian Interior Minister Matteo Piantedosi called the pact a “great success” for Europe and Italy, which meant that EU border countries most exposed to migration would no longer feel alone. German Chancellor Olaf Scholz said agreement on the pact was very important, relieving states affected, including Germany.
Previous efforts to share out the responsibility of hosting migrants have foundered because eastern EU members in particular were unwilling to take in people who had arrived in Greece, Italy and other countries. Under the new system, countries not at the border of the bloc will have to choose between accepting their share of 30,000 asylum applicants or paying at least €20,000 ($21,870) per person into an EU fund.
The screening system envisaged will seek to distinguish between those in need of international protection and others who are not. People whose asylum applications have a low chance of success, such as those from India, Tunisia or Turkey, can be prevented from entering the EU and detained at the border, as can people believed to be a threat to security. The processing of applications would also be speeded up.
Amnesty International said the pact would set EU asylum law back decades and lead to greater suffering for people seeking asylum. The human rights organisation described the system as being designed to make it harder for people to access safety. “The pact will almost certainly cause more people to be put into de facto detention at EU borders, including families with children and people in vulnerable situations,” Amnesty insisted.


With the fast pace changes in technology business owners will want to keep up with the newest versions of the products. Buying new equipment means you keep up with changing technology; it also means you are more productive and, therefore, more competitive. You will have the warranty of a new purchase, but as you have just bought a large machine, your finances may have taken a hit. In the heavy equipment business, this may not be a viable option. What can be done with the old machinery? You can put it up for sale, but how quickly can you get it off the books? As the owner of a business, small or large, you must assess your needs to figure out which is the better option for your company. You can buy new, or you can consider buying used or refurbished construction equipment in Prichard, Alabama. However, if you plan on buying used or refurbished, you must buy from a reputable company like NLEQ. Today many in the construction business come to National Lift Equipment when considering purchasing a used excavators than any other used construction equipment broker in Prichard, Alabama. Whether you are in search of a Shooting Boom Forklifts for sale in Prichard, Alabama or straight boom lifts we will do our best to get you the right deal.
Buying used equipment may not be even considered by some, but for many small businesses, it certainly is , for some, the only option. For one thing, it’s far less expensive than buying new. Much is required of you, the buyer before you make this type of purchase. You must do your research and be sure to buy from a reputable used construction equipment company in Prichard, Alabama. Note that usually, used equipment doesn’t carry any warranty. This is why you must do research and if possible you might want to hire a mechanic. But most importantly, buy from a reputable company like NLEQ. When thinking about to acquire used pieces of construction equipment, you should let us help you. We have been in business for over 33 years, and proudly continue to serve communities like Prichard, Alabama by providing the best selection of used towable boom lifts for sale in Prichard, Alabama.
Since the economic slowdown of 2006, there has been a lot of used construction equipment lying around. However, if you take a second look, you will realized that the majority is nothing but the leftovers other construction contractors have left behind. That is not the case at National Lift Equipment. We are proud the quality of equipment we sell, and that is because every Articulating Boom Lifts that we take as trade-in goes through a 40 point inspection more rigid than our competition to insures it is being in out lot. So, keep in mind if you are in search of a Cranes or any heavy or used construction equipment for sale in Prichard, Alabama give National Lift Equipment a call. With that said, if you are just looking around we encourage you to take a look at our blog. Where you will find usefull information about and how to aquire any heavy and used construction equipment. If you are seeking the best price on excavators for sale you need to call National Lift Equipment…
It appears that a lot of contractors seeking used construction equipment in Prichard, Alabama. But the selection of used construction equipment in Prichard, Alabama is not the best. If you are in the construction industry, you may have been out looking for used construction equipment for sale in Prichard, Alabama, you may have realized that the quality of the existing inventory is not the best. That is not something you need to worry about when dealing with National Lift Equipment. If you’re searching for Skytrak Telehandler For Sale in Prichard, Alabama, you are at the right place! Every piece of used construction equipment ranging from Backhoes to Push Arounds at out lot has gone through an intensive inspection process to insure it performs as intended on the job site. Quality is what makes National Lift Equipment different. So, if you are looking to acquire used construction equipment in Prichard, Alabama give a call!
Even under the current economic land escape it may be a good idea to buy second-hand construction equipment There are lot of reasons why people chose to buy new construction equipment. Nevertheless, there are a lot of good reasons why you should consider used over new. One thing is for sure! New construction equipment comes with a huge price tag and it is not only the upfront cost and taxes but also the hidden cost like depreciation and required maintenance. Since 1990 National Lift Equipment has been providing contruction companies high quality heavy equipment from some the top names in the business. What can you expect at National Lift Equipment LLC? How about Shooting Boom Forklifts and Aerial Work Platforms from top names like Caterpillar, Kubota, Terex, and more. Before you buy take a look at our lift equipment articles above!


text:
Description: The Plat
Date Listed: 28 November 1980
English Heritage Building ID: 299510
OS Grid Reference: TQ1196222723
OS Grid Coordinates: 511962, 122723
Latitude/Longitude: 50.9930, -0.4060
Explore more of the area around Shipley, West Sussex at Explore Britain.
5404 The Plat
TQ 12 SW 15/455
Formerly 2 cottages. C17 or earlier timber-framed building with red brick
infilling, partly refaced with red brick and grey headers. Hipped tiled roof.
Casement windows. Gabled porch. Two storeys. Four windows.
Listing NGR: TQ1196222723
Source: English Heritage
Listed building text is © Crown Copyright. Reproduced under licence.

domain: geography


text:
CHARLESTON -- A married couple is suing after the husband was injured while on a job site.
Glenn Fultz and Constance Fultz filed a lawsuit May 6 in Kanawha Circuit Court against B&F Contracting Inc., Northern Management Services Inc. and B. Armstrong Electrical Services Inc.
According to the complaint on May 14, 2013, Glenn Fultz, while employed as a laborer by B&F Contracting, was delivering concrete to a work site when metal grating at the site collapsed.
The resulting collapse caused Glenn Fultz to fall 14 feet onto the ground before the grating collapsed on top of him, according to the complaint. The plaintiff suffered a spinal fracture as a result of the accident, according to the complaint.
As a result of the accident, Glenn was forced to undergo serious and extensive major surgery and medical care, suffered permanent loss of earning capacity, severe emotional distress and permanent physical and vocational impairment, among other claims.
Glenn Fultz is accusing B&F Contracting of not maintaining a safe work environment. He is naming Northern Management Services and B. Armstrong Electrical Services in the suit for failing to properly inspect the site, which led to the accident.
The plaintiffs are seeking compensatory damages and costs.
They are being represented in the case by attorneys Guy R. Bucci and Mark A. Barney of Bucci, Bailey & Javins LC, Aaron L. Harrah of Hill, Peterson, Carper, Bee & Deitzler PLLC and Christopher J. Heavens of Heavens Law Firm PLLC. The case has been assigned to Circuit Judge Tod J. Kaufman.
Kanawha Circuit Court Case No. 14-C-844


A new rumor has surfaced that suggests GSC Game World's cancelled PC-exclusive title STALKER 2 has been picked up by Bethesda, although other rumors conflict and say this isn't true
Is Windows Phone about to go big?
Digital Storm Vanquish II gaming PC
Which Android handsets would you like to see as Windows Phones?
Just three days after the launch of stalker app Breakup Notifier, Facebook promptly banned it after over 100,000 plus curious users logged into the service; but from today that relationship appears to be "on" again. The...
In another disturbing turn of events that have left Facebook CEO, Mark Zuckerberg this time fearing for his life, Zuckerberg has been forced to file legal papers claiming Pradeep Manukonda, 31, tried to "follow, surveil...
Remember back in November 2001, when S.T.A.L.K.E.R. was announced, and quickly became one of the most anticipated PC games at the time? You may then recall several delays thanks to numerous setbacks, including a pre-alpha...


Porbandar: It's a big catch for India as a Pakistani boat has been caught!
Amid heightened vigil, a Pakistan boat was seized off Gujarat coast on Sunday.
Also, nine sailors have been caught by the authorities.
Indian Coast Guard ship 'Samudra Pavak' had apprehended the Pakistani boat off Gujarat coast.
Coast Guard officials apprehended the crew members travelling in the boat as soon as it entered in Indian waters around 10.15 AM.
"In the prevailing scenario, Coast Guard ship Samudra Pavak apprehended a Pakistani boat with nine crew members," an official statement said.
Further investigation is on and the apprehended Pakistanis will be quizzed at Porbandar.
Security agencies have been extra cautious in view of the prevailing security scenario following the surgical strike by Indian army on terror camps across the LoC.


Below are the available bulk discount rates for each individual item when you purchase a certain amount
Christmas is Carll's favorite holiday...so why not celebrate the soft Christmas feelings with the softest shirt on the planet??
Super soft 50% ring-spun cotton/50% polyester (100% comfortable)
No feel water-based print
30/1 fine knit jersey, tubular construction
Ribbed crewneck, taped neck and shoulders with double-needle sleeve and bottom hem