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Lika Joenathan
Realtor, Small Business Owner, and Social Media Manager in San Diego, California
Lika Joenathan
Realtor, Small Business Owner, and Social Media Manager in San Diego, California
I like to see the world through the eyes of others: enhance the power of communication by its unique perspective to a multi-ethnic public.
A result driven professional with over 15 years of Management experience. Background: Sales, Office and Property Management, Entrepreneur, Real Estate, Mortgage, Marketing, Legal training.
Strong Eastern European education topped off with valuable multi-cultural work ethics. Personable, cooperative, multi-dimensional, and motivated self-starter and entrepreneur. Communication skills in English, Ukrainian & Russian languages. Adaptable to changing needs and times. Hands-on individual ready to assist with questions about technology devices, computer troubleshooting, business development, marketing and real estate consulting.
Born and raised in the area called "The Pearl of the Sea" aka Odesa, Ukraine – a multinational city, I learned several languages. After the break of The Soviet Union, rising economic inequality and hardship made it impossible for the citizens to pursue an education. I had applied for student's programs in Germany but ended up accepting a program offer in Indiana, USA. I enjoyed living in the Midwest and have gotten to know so many dear people. The Midwest is a quaint and friendly place to live and many of my clients became close friends. Because of the success I experienced in the Real Estate industry along with the majestic changes I enjoyed throughout the four seasons, I made the decision to stay in The United States. Such magnificence I had never seen before. However, I missed the Black Sea very much and couldn't travel as often as I used to before my business picked up. That said, after visiting San Diego for a conference back in 2008 I was drawn to move closer to the ocean. In November of 2011 I relocated to San Diego and began to explore the ocean and outdoor activities including hiking, swimming, boating, camping, playing tennis & volleyball. Although, restarting the Real Estate business has its challenges, I enjoy building my sphere of influence and clientele. Meanwhile, I am currently seeking a challenging Office Management position full- or part-time.
Education: Legal Foundations of Entrepreneurship Security at Law University of Kyiv in Ukraine.
I enjoy volunteering and helping others. My favorite places are Braille Institute of San Diego and the Balboa International Villages. I have 12+ years of combined volunteer work at a few organizations including American Cancer Society, United Way,


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Mr. YARMUTH. Madam Speaker, it seems like Groundhog Day all over again.
Earlier this week, GOP leaders laid down a new gambit on the old debate over whether to acknowledge our Nation's financial obligations. Those leaders have already abandoned the deal we made on the last debt ceiling package and are shifting all the cuts to education, infrastructure, and other vital domestic programs. Now they want another round of unsustainable cuts to these programs which will again bring us back to the brink of default.
We know the possible consequences: Market collapse, jobs lost, more than $1 trillion added to the deficit every year, interest rates will rise. Just getting close to this cliff threatens the U.S. credit rating. We know that from recent experience.
The Speaker has said, no, he doesn't want to abandon the debt ceiling, he doesn't want to violate the debt ceiling, he doesn't want to let the country go into default. But isn't this the same kind of uncertainty that our Republican friends say they are most concerned about? One day it's, Well, we're not going to raise the debt ceiling. The next day, No, I didn't mean that.
We need certainty; we need stability, and we need to recognize this Nation's obligations.
Source: http://thomas.loc.gov

subset: commoncrawl


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Play ox games online
Play 7 free ox games online.
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- OX Simple catching game
- The Ox Man The ox man is the warrior of the planet. His duty is to kill enemies coming from the other planet. Use Following key to play the game. Left Arrow - Mo...
- Noughts and Crosses Extreme The most extreme Noughts and Crosses ever created! With explosive gameplay, a spinning board and new squares that appear from thin-air, play off again...
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- Tic Tac Toe 3D An original classic. Here's a new dimension to an old favorite.
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- Gothic Angel Girl Dress Up This goth girl angel has some dark styles and fashoins to choose from. You can do her makeup and also pick her clothes and even some dark wings to com...
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- Fashion Office Lady's Wardrobe Chris is a office lady and be fashion and course,her clothes are so colorful and her to select suitable go to her wardrobe


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The following Taylorsville Elementary students were named to the All A honor roll for the 2010-2011 school year:
Connor Foster, Madison Goodlett, Jacob Goodwin, Joshua Hall, Jaynie Heckert, Joshua Richards, Allie Roberts, Zachary Tipton and Hunter Wethington.
The following Taylorsville Elementary Students were named to the A/B honor roll for the 2010-2011 school year:
Third grade:
Have you ever wondered what would happen if we in America would leave our wants and wishes to Jesus. Jesus tells us in Matthew 6:33 what to do before we start wishing and wanting.
"But seek ye first the kingdom of God, and his righteousness and these things will be added unto you." Matthew: 6:33
"Ask and it shall be given you, seek and ye shall find, knock and it shall be opened unto you. For every one that asketh, receiveth; and he that seeketh, findeth, and to him that knocketh it shall be opened." Matthew 7:7-8
Every day, whether we realize it or not, we meet people who are struggling with pain or fear or heartache. They may smile at you in the grocery store or frown at you as you wait at a stop light, but they are hurting. The teller at the bank or the guy working on your car may be hanging on by a thread, but most of the time we never see it, most of the time they never show it.
One local reader submitted these old receipts for subscriptions to The Spencer Courier and to The Spencer Magnet. The Spencer Courier receipt, pictured above, dates back 100 years.
In 1983, this scribe wrote a series of columns on one of the men of Confederate guerrilla leader William Clarke Quantrill and his infatuation with a young Spencer County girl.
A healthy diet is often mistaken for one that is devoid of desirable foods. But just because a diet is healthy doesn't mean it can't be delicious as well. A few easy modifications to your favorite recipes can make a diet healthier without sacrificing taste.
Marty Allan Harp recently returned from vacationing in Los Angeles. While there he went to the annual Days of Our Lives fanfare weekend. He is pictured here with Suzanne Rogers who plays Maggie Horton on Days of our Lives. She's been on the show since 1974. Marty has been a fan of the show since then, and has attended the event since 1986 and has met most of the cast members.
Mr. and Mrs. Phillip Benningfield, of Taylorsville, announce the engagement and upcoming marriage of their daughter, Maria Anka Dawn Benningfield, to Erin L. Thompson, son of Mr. and Mrs. Stan Thompson, also of Taylorsville.
Brittany Nicole Ross of Louisville and Jacob Michael Lancaster of Louisville announce their engagement and upcoming marriage at noon on July 1 at the Spencer County Courthouse.
The bride-to-be is a daughter of Terry and Tammy Ross of Taylorsville.
She is a 2009 graduate of Spencer County High School.
The groom-to-be is a son of Mike Lancaster of Louisville and Dottie Lancaster of Cecilia.
He is a 2001 graduate of Carroll County High School.
Father's Day is the perfect time to try out a new dessert. Gooey butter cake can be made the day before, and is actually better on the second day. That makes it perfect for a Father's Day picnic or family gathering. It is also adaptable to flavors that dads enjoy by adding extracts or other embellishments.
Gooey butter cake was invented in St. Louis, Missouri in the 1930s. It has been popular ever since.
Copyright © 2017
The Spencer Magnet is your source for local news, sports, events, and information in Spencer County, KY and the surrounding area.


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Disney's One Saturday Morning | 1997
Jump to
Wraparound segments for ABC's Saturday-morning children's series. Elements of "One Saturday Morning" include: "Great Minds Think For Themselves" (Robin Williams, in a reprise of his Genie role in Disney's animated film "Aladdin," offers his take on famous historical figures); "Manny the Uncanny" (an inquisitive character who goes out into the world to see what people are doing); "The Monkey Boys" (Hodge-Podge and Buddy combine physical humor and toony logic to produce a live-action cartoon); "Mrs. Munger's Class" (animating the mouths of typical sixth-grade class pictures, students interact with each other and their teacher, carrying on outlandish conversations); "What's Up With That?" (a young professor offers quick, informative facts); and "Tube Dweller's" (follows the antics of maintenance men Bob and Bob as they dawdle behind your screen).
New cast members for the 1999-2000 season include: Miss Moose, the mascot; Millie, the message lady; and the musical group, Roller Toasters. New segments include: "Cafeteria," a musical take on conversations that take place on a school cafeteria line; "What's the Diff?," an animation segment that challenges kids to tell two very similar-looking "snapshots" apart; and "Find Out Why," an animated segment hosted by "The Lion King's" Timon and Pumbaa, which explains a variety of scientific occurrences and commonplace technologies, including rain, sneezing, static electricity, wind, airplane flight, etc.


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Dismiss NoticeYour support makes a difference...
Industry changes have drastically reduced orthopedic partner contributions, which for 13+ years supported the operation and development of BoneSmart.org.
Our patient-focused articles and world-class joint replacement community forum have helped millions of people. Your gift today provides the important funds necessary to keep this valuable resource online.
Medicare Changes for 2018 - USA Patients
Discussion in 'Knee Replacement Pre-Op Area' started by Jamie, May 4, 2015.
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Teen Cream FunFavorites [+]
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Old West Brown Childrens Boys Man Made Material Ostrich Cowboy Boots
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PowerBar 24g Protein Plus Bar - PROTEIN PLUS
View larger image Roll over to zoom
Protein is a key macronutrient and the building block of human tissue. Because protein is an essential part of a healthy diet, POWERBAR® PROTEINPLUS® bars were developed according to the needs of world-class athletes and others to be a great-tasting and excellent source of protein. For satisfying nutrition, all POWERBAR® PROTEINPLUS bars contain 24 grams of our POWERBAR® TRISOURCE™ protein blend, a combination of high-quality soy protein isolate, whey protein isolate, and calcium caseinate.
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Items listed in the ' Sale ' section of the Running Room Shop are already at a clearance or Special price. The sale of this product cannot be combined with any other promotional offer, coupon or discount.
All Clearance and Special Priced items are final sale for online orders. We will exchange for your correct size pending inventory availability at a convenient location within 30 days, please contact Customer Service to arrange the exchange or store credit


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10' Straight Trade Show Pop Up with Fabric Panels Kit
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01 - 4 x 3 Straight Frame21 - Magnetic Channel Bar04 - Fabric Front Panel02 - Fabric CURVED End Panel02 - Lumina 200 Low Voltage 120 watt Halogen Light01 - OCP Hard Shipping Case
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2000 race well underway in Iowa
DES MOINES, Iowa - House Budget Committee Chairman John Kasich usually makes news balancing federal budgets or pontificating on Sunday network talk shows.
But a few weeks ago, the Ohio Republican found time to spend an afternoon knocking on doors in nearby Norwalk with John Parker, a GOP candidate for the Iowa House.
"I'm pretty stiff, but he's real loose at the door," Parker said. "At one house, we even shot baskets for a while."
They're spending dozens of days in Iowa, raising money for candidates, giving money to candidates, putting together organizations and helping generate media attention as they build good will with political leaders in the state that every four years hosts the nation's first presidential caucuses.
"Where else in the country do you get the chairman of the Budget Committee going door-to-door with a legislative challenger?" asked state GOP Chairman Steve Grubbs. "It's a tremendous benefit to us."
It's a 1998 political wrinkle unseen in any state except probably New Hampshire with its leading presidential primary.
That activity is amounting to an unprecedented level of attention by presidential hopefuls. GOP leaders believe Republicans are profiting more than Democrats because their presidential candidates are visiting Iowa four times more frequently.
"The caucus system has been great for the parties in Iowa," said John Norris, the state Democratic chairman.
For a long time, he might have added.
Many anti-Vietnam War activists of the early 1970s remained involved with the Democratic Party into the 1980s, when hotly contested presidential battles stirred interest that helped the party take control of the Legislature.
In 1984, Democrat Tom Harkin wrested a Senate seat from Republican Roger Jepsen "by building an organization out of everything the presidential candidates like Walter Mondale left behind," former state GOP Chairman Brian Kennedy said.
In recent years, activity has shifted to the GOP. Christian broadcasting executive Pat Robertson's insurgent 1988 presidential campaign activated many evangelicals - some of whom are running for office as Republicans today.
In 1992, Harkin's bid for president gave him an advantage here that dried up the Democratic caucuses as other candidates bypassed Iowa. That didn't change in 1996, with President Clinton seeking re-election. But GOP presidential activity has intensified, and the party has made gains at the state level.
Caucus campaigns, said Kennedy, "bring in new people. A lot of people get involved in a presidential campaign and then stay involved with the party."
Spillover from those campaigns is touching all levels of Iowa elections this year.
Iowa Republican Sen. Charles Grassley is heavily favored to win re-election, yet GOP Sens. Bob Smith of New Hampshire and John Ashcroft of Missouri have shown up at house parties to help him raise money. So have former Vice President Dan Quayle and former Tennessee Gov. Lamar Alexander.
On the Democratic side, Minnesota Sen. Paul Wellstone has helped David Osterberg in his long-shot bid to oust Grassley.
Presidential luminaries constantly line up to help Republican Jim Ross Lightfoot and Democrat Tom Vilsack raise money in the first gubernatorial race without an incumbent in 16 years. While Lightfoot leads in polls, Vilsack is pushing hard.
House Minority Leader Richard Gephardt of Missouri was in Iowa Saturday. Vice President Al Gore is scheduled in Tuesday.
With Iowa's farm economy suffering from low prices, Nebraska Sen. Bob Kerrey has campaigned in the state, blasting the 1996 GOP Freedom to Farm bill. Vilsack has echoed similar themes.
Three congressional incumbents face strong challengers - First District Republican Jim Leach in a scrappy challenge from Democrat Bob Rush; Second District Republican Jim Nussle facing Democrat Rob Tully, and Third District Democrat Leonard Boswell fighting Republican Larry McKibben. Each has had a wealth of help from presidential hopefuls with money and at media events.
The battle to control the Iowa House is being waged on about two-dozen fronts, with Democrats needing five seats to wrest control. On both sides, presidential contenders have pitched in to help the locals.
Steve Forbes has introduced candidates to business contributors. Quayle has donated an amount "approaching six figures" to GOP candidates, said Gary Geipel, his full-time Iowa staffer.
Pro-family activist Gary Bauer has hired veteran evangelical organizer Marlys Popma to help lay his foundation for a presidential bid. And as she travels Iowa, "she's out there helping candidates understand what they're supposed to be doing," said Keith Fortman, state GOP executive director.
Alexander, meanwhile, is building his 2000 organization by having his 1996 supporters turn out 1998 voters. Some 50,000 Republicans who don't vote in nonpresidential years have been targeted.
Forbes, Bauer, Alexander and Missouri's Ashcroft have run television ads in Iowa touting such GOP issues as tax cuts and family values. "It helps the environment by making them high-profile issues," Grubbs said.
Similarly, John Del Cecato, press secretary for the Iowa Democratic Party, points to presidential candidates and staffers running get out the vote clinics for party activists.
Political leaders see long-term benefits from such help.
"It helps build the party," said Kennedy, who now works for Alexander. "It lifts the level of politics in Iowa ... We get the best spokespeople and the best operatives in here, all because of our never-ending presidential campaigns."
By DAVID YEPSEN, The Des Moines Register
© Copyright 2002 USA TODAY, a division of Gannett Co. Inc.


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How to Look Great In a Couple of Simple Steps
The next article has some useful tips such as the following are walking gemstones on the road to beauty expertise.
Sun block is regarded as the healthiness of beauty items. When choosing which sun block to make use of, try items that appear at first sight great for the skin. You can preserve the skin youthful and nourished with the proper wealthy protecting ingredients.
You'll find beauty everywhere. You may take a look at trees as beautiful, art or perhaps your spouse. When to consider beauty, you will find you are feeling more beautiful yourself.
Make certain that you are not allergic to eye lash glues before you apply them. Place a protective strip within the glue for your arm and canopy it.
Sodium bicarbonate can give hair be shiny again. Mix just a little sodium bicarbonate with shampoo inside your regular quantity of shampoo. Go to wash your normal hair washing routine. Hair is going to be restored to some healthy and healthy.
You are able to adjust hair to thin the feel of the face. A haircut with lengthy streaks can have the desired effect. You may also use lowlights and highlights. They are especially flattering and draw the characteristics of the face.
Always wash off your makeup before going to sleep. Lightly wash off all makeup having a warm clean cloth. Following this, simply cleanse the face while you would. Make-up that is not removed will block your pores and acne problems.
You need to consider carrying this out throughout the summertime. By continuing to keep cosmetics for example lotions and oils inside your refrigerator, toner and oils within the fridge you'll be able for their services even when there's a heatwave. The skin will understand the cooling effect.
Replace your pricy toners, exfoliates, toners and moisturizers with pure castile soap, a gentle clean cloth, aloe, along with other natural materials. These natural products are ideal for all skin tones. Tea tree oil can be used a medicated moisturizer.
You can make your own mouthwash with peppermint oil and purified water. Use a small amount of oil per every ounce water you utilize. Boil water, then drop the oil right into a ceramic container. Add boiling water for that container. Cover the mix having a flannel and then leave to awesome. Pour right into a clean bottle and tightly screw around the lid.
Try to avoid caffeine if you are searching to enhance your beauty. Limit your consumption of tea or coffee to 1 cup each day. Consuming decaffeinated and eco-friendly teas are smarter than consuming a few of the popular beverages.
Use eyeshadow that shimmers with discretion on your beauty routine. The shiny effect you receive whenever you put on glittery shadow can make your vision appear better and better. Choose shimmery shades that compares to the skin tone to help keep flattering and mature. Try new things and experiment to obtain the right look.
An inexpensive kitchen sponge. Kitchen sponges clean bathroom surfaces along with other types and therefore are frequently significantly less pricey.
The coarseness from the paper works great to consider oil out of your oily skin. Only one square of paper and press it for your skin!
Apply lotion immediately after shaving to find the best effect. Ingrown hairs can result from dried-out skin, and taking advantage of lotion every single day works wonders. Jet Ski from formation from the ingrown hairs while treating existing trouble spots.
There's one beauty tip that whenever not adopted greatly decreases the look of both women and men. You to proper care of your eyebrows there ought to always be a couple of them.
Should you suffer dried-out skin in your face, try visiting a beautician to obtain an effective moisturizing treatment. This kind of treatment targets dry skin because it helps smooth and refresh it by eliminating dead skin cells evidently and refreshes and smooths the skin's appearance by eliminating the dead skin cells.
Skincare is essential if you wish to maintain confidence and searching more beautiful. You may even take with you lotion and employ whenever your skin feels dry.
Studying these beauty advice should already begin to make you are feeling better. You need to now have a very good idea on where one can begin when you wish to construct an individual beauty plan. You could return and browse it can.


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October 2016
February 2016
May 2017
July 2015
|June 2015
South Africa is a country built for road trips. Endless stretches of road through some of the world�s most breath-taking scenery are just a highway away for most of us. Whether you like beaches or the Berg, deserts or tropical rain forests, there is a road waiting to be travelled and years of memories to be made. Find out how to plan your next road trip right here.
What is a road trip?
Perhaps we should start with a definition of the Great South African Road Trip. Technically a road trip is defined as a car journey. However, leaping into your car at 4am on Good Friday morning and dashing to Durbs or driving up to the Kruger in a day are not considered road trips. Any true-blue road tripper knows that driving through South Africa can be a holiday in itself. This is the essence of a road trip: two or more days of easy travelling, a scenic route, top-notch stay-overs along the way and kilometres of some of the best relaxation a holiday has to offer.
Where do I start?
Planning your road trip means some planning and lots of research. Work out how many days you have to spend driving and what distance you can cover in that time (plan on doing no more than 300km � 400km each day), then plot your route and find suitable accommodation. Or, if you already have a holiday destination in mind, find the quietest roads that will take you there and build your travel time into your holiday.
Be adventurous and plan to see parts of South Africa that you have never seen before. Perhaps you would like to drive down to Cape Town through the Karoo or up the West Coast of South Africa. You could explore the Natal Midlands and the South Coast, or even head for Swaziland or Lesotho. Be sure to take note of must-see attractions along the way and build them into your itinerary.
The AA has all the tools you need to plan your trip. Try the AA Distance Calculator to calculate the distance between popular South African destinations and the Road Conditions Report to check your route. Use the AA�s online or print maps. And if it�s accommodation you are after, try AA Travel Quality AssuredAccommodation.
The 10 golden rules of every road trip
And last but not least, relax and take it easy. Remember, it is the journey and not the destination that counts! And don�t forget to take lots of photos when you stop!


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Adopt BIA Chip fat analysis technology to acquire the body fat information.
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|Brand Name
|Platform Material
|Toughened Glass
|Bluetooth Version
|Bluetooth 4.0
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|Standby Time
|323 Days
|Product Size
|320L x 320W x 23H mm
|G Sensor
|Weighing range
|Display Screen
|Large LED Screen
|iOS 7.0 above
|Wechat 6.0 above
|Android Phone
|Android 4.3 above
|Product Weight
|2Kg(without battery)
|Package Content
|1 x Lifesense Melody Fat Weight Scale
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Zapals.com is an internationally renowned, global shopping website operated by the Zapals Corporation Limited Company.
Our offices are located in Australia, Hong Kong and Mainland China and our aim is to deliver one of the largest, most innovative online department stores in the world.
We've got an extensive range of high quality products at the lowest prices for our customers. Our shopping range is designed by experienced professionals in compliance with international standards, quality control, trends, demands, and policies.
Our ultimate goal is to be an innovative, one-stop, online destination for anything you can find in the home and anything that you'll need in your life!


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Ask yourself: just how many times have you heard the phrase "the future is now"?
Believe it or not, yes, it is true. Future has never been this close and in our range, within the grasp of our hand. In this video we have eight different and innovative trucks and buses concepts that are bound to impress you and make you think.
Technology is rapidly developing, faster than ever before in history of man and is a core, vital part of our modern existence. Everyone wants to keep up and stay in touch with all the advancements, updates, improvements and innovations on the horizon, especially when it comes to techno titans such as vehicles. Indeed, these are only concepts, for now, but what can stop us from thinking they can't be achieved and realized?
Primarily, we have Mercedes Future Truck 2025, which is supposed let us take a sneak peek at the concept of a driverless truck! Pretty cool, don't you think? Another striking concept and model is the Iveco Z Truck, which looks very futuristic and promising indeed. If you are into aesthetics and you love eye-candy and appreciate beauty, then the Willie Transparent LCD Bus is bound to satisfy your criteria and impress you with its looks.
We are not going to list all the different, breathtaking models of trucks and buses from the video in this text, instead, we strongly advise that you watch this interesting video and see it with your own eyes, trust us, it is worth it!VIDEO on PAGE 2 !!


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| "Barnacles! What could be worse than a giant paint bubble!?"
It has been suggested that List of SpongeBob SquarePants marathons be merged into the List of events article.
Nick's First SpongeBob Marathon
Features 9 episodes.
Season 2 Begins
Features 8 episodes.
Season 2 Ending
Features 16 episodes.
Party Pooper Pants (Marathon)
Features 4 episodes.
Full Season 4 Marathon
Features 38 episodes.
Best Day Ever marathon
Features 100 episodes.
Valentine's Day (Marathon)
Features 7 episodes.
Atlantis SquarePantis (Marathon)
Features episodes from Z-A.
Sponge in Space
Features 12 episodes.
Season 6 First 4 Episodes
Features 8 episodes.
Pineapple Fever (Marathon)
Features 4 episodes.
Chum Caverns (Marathon)
Features 5 episodes.
Season 7 First 4 Episodes
Features 8 episodes.
Truth or Square and SpongeBob's Last Stand
Features 6 Special Episodes.
1. Procrastination
2. Sailor Mouth
3. Tea at the Treedome
4. Slide Whistler Stooges
5. Clams
6. Squidward the Unfriendly Ghost
The Ultimate SpongeBob SpongeBash
Features 10 Episodes
10. Suds
9. Patty Hype
8. Valentine's Day
7. The Paper
6. Help Wanted
4. The Camping Episode
3. Ripped Pants
1. Pizza Delivery
This list is incomplete, but you can help Encyclopedia SpongeBobia by expanding it.


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Home > Eyeglasses > Polo > PH2065 (5563)
PH 2065 is available in 5 additional attractive styles
New Releases
Size and availability
Free outbound and return shipping
30-day money-back guarantee
Low prices
Free outbound and return shipping
30-day money-back guarantee
Low prices
PH2065/5563 Eyewear
The term "visual aid" is unsuitable for PH2065. This is an accessory that ushers in a new you, emphasizing your clear view on fashion. The PH2065 is available in the Edel-Optics online shop in other styles from Polo collections 2016 and 2017.
With no frills and state-of-the-art materials and craftsmanship, these men's glasses stand for stylish design and self-confidence. Wearing full rim eyewear, you make it unmistakably clear that you are going the whole hog. Rectangular models are particularly suitable for round and oval faces. The rectangular shape also builds a bridge to corners and edges from an image aspect. Plastic is a very lightweight and flexible material. This ensures a long life and a high level of comfort. Transparent eyewear is the right choice, if you like the extraordinary but nothing too gaudy. It lends your facial features a certain lightness and gives your appearance a sense of style.
Moreover, we can also provide you with the correct prescription lenses at an incredibly low price. The only thing you need to provide us with are your current dioptre values determined by your ophthalmologist. Together with our digital dispensing optician, you can choose between reasonably priced brand lenses from Germany or premium brands such as Seiko, Hoya and Rodenstock. Edel-Optics' plastic lenses have many features such as anti-reflection coating, lotus effect, cleaning coating, antistatic and hard coating at no extra charge.
The model has already been ordered and will shortly be back in stock. If you order now, you will get the reduced price and once the goods arrive, we will send you your new Polo on the same day. And because Edel-Optics is a paradise for bargain hunters, you can get this top model at an incredibly low price. What is labelled as "in the sale" at other online shops, is for us "all-day-everyday" prices.
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By: CP Williams and SH Clearwater
Contrasts the advantages and tradeoffs of classical computing with quantum computing and point to remarkable new directions for computing.
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About the Book
"Sky Wires" creates a different way of admiring the skies on cloudy somber days after rainstorms or preceding precipitations of meteorological occurrences.
Features & Details
- Category Fine Art Photography
Small Square, 7×7 in, 18×18 cm
64 Pages
- Publish Date Aug 17, 2013
- Language English
- Tags Sky, Skies, Clouds, Weather, Electrical, Storms
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Each month, Rated People connects over 50,000 homeowners with quality tradesmen - simply enter your job details in the form above to get started.
We'll match your job to the relevant tradesmen in your area - those interested will pay to see your contact details and call you to quote.
As each tradesman expresses interest in quoting on your job, we'll send you their profile and ratings to help you choose the right one for you.
After the work is complete, return to the site and rate your tradesman based on their quality of work, reliability and value for money.
Added 09/06/2014
Build a wall approx 20ft length by 7ft tall to replace existing fence. Extend patio base 15ft...
Added 22/04/2014
Unsold lead, updated details: I really have no idea how much this job should cost but £1000...
Added 17/05/2013
WOULD LIKE A GOOD BASE ABOUT 6ft.x3ft.,FOR A GARDEN SEAT. Timing: Within 2 weeks
Added 05/02/2013
My house used to have a wall downstairs separating the living and dining room. I would like...
Added 18/06/2011
We are fixing all of the jobs that need to be done before probably selling the house. - We...
Added 12/10/2010
This is the details of the job for the expert review.
The results speak for themselves...really good job done
Very pleased with all the work carried out. The quality of all the work is excellent. Thanks so much!
Very pleased with the job. Substantial internal wall removed, steal beam installed, boarded and plastered all in one day. Would recommend.
I was very pleased work, they were always quick to respond to any emails and they showed up on time as well!
Highly recommend using this tradesman. Very reliable and did a superb job!
Managing your job is easier with the Rated People Homes App, download it today.


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My friend and I just returned from our vacation and I'm already trying to plan a trip back. The hotel has the best views of any resort I have ever stayed at and it's very clean. Two wonderful pools, and great food. But the best part of the hotel is there amazing activities director Miguel Tapia, he will make sure you have a blast around the pool and on the beach. Look forward to seeing him entertain again soon. Thank Miguel!!
- Official Description (provided by the hotel):
- The most exclusive Cancun All-Inclusive experience is waiting for you! Sunny days, white-sand beaches and spectacular views of the turquoise Caribbean Sea are just a little of the many things you'll enjoy by staying at Grand Park Royal Cancun Caribe. Boasting beachfront location, right in the heart of the Hotel Zone, our All-Inclusive family resort in Cancun promises an unforgettable stay in the most popular and exciting destination of the mexican Caribbean. ... more less
- Reservation Options:
- TripAdvisor is proud to partner with Expedia, Orbitz, BookIt.com, Booking.com, Hotels.com, Travelocity, HotelsforEveryone, Cheap Tickets, TripOnline SA, HotelQuickly, Hotwire, Cancelon, getaroom.com and Evoline ltd so you can book your Grand Park Royal Cancun Caribe reservations with confidence. We help millions of travelers each month to find the perfect hotel for both vacation and business trips, always with the best discounts and special offers.
- Also Known As:
- Grand Park Royal Cancun Caribe Hotel Cancun
- Caribe Park Royal Grand Cancun


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Join our Student-Centered College!Do you seek out opportunities to share your expertise and love of physical therapy to the next generation? Do you find yourself discussing the intricacies of kinesthetic joint movement with friends and family? Are you inspired by collaboration and community engagement? Would you enjoy the opportunity to develop a new PTA program? If these questions resonate with you, then we invite you to consider joining our team. Pierce College is an innovative and equity-focused community college. We are dedicated to helping all students achieve their academic and professional goals. About the Program:Pierce College is developing a physical therapist assistant program to serve the South Puget Sound area. Currently we partner with Whatcom Community College to provide a cohort of students on the Pierce College--Ft. Steilacoom Campus. Curriculum development and accreditation approval are scheduled to be done over the next year.About Pierce College:Pierce College is an innovative community college committed to student engagement and learning. We value student learning experiences that foster equity and diversity in the classroom and expand into our local and global communities. From closing achievement gaps for all students to building directed career pathways, faculty, staff, and administrators work collaboratively to lead initiatives that advance student success. The college serves over 15,000 students in basic skills, transfer, and professional technical programs. Through continuous efforts, we have improved retention rates by 16% and graduation rates by 25%. This year, Pierce was named a top community college in several national polls. Pierce College employees and students live and work in various communities that offer the diverse cultural and environmental benefits of rural, suburban, and urban lifestyles in the Pacific Northwest. As part of the community and technical college system in Washington State, the District includes Pierce College Fort Steilacoom, Pierce College Puyallup, Pierce College at Joint Base Lewis-McChord, as well as other extended learning sites.Responsibilities of the successful candidate include:
Application Requirements:To be considered for this position, applicants must include the following items in their application package:
Terms of Employment:This position, which is contingent upon funding, is a full-time, tenure-track assignment of 173 contracted days per year with the tenure process starting in September 2017, and with the opportunity to start working an earlier quarter. Assignment may include district discipline/program coordination, as well as evening, hybrid, and/or online teaching. This is a District position with a "home" campus assignment. Initial placement of this position will be at the Fort Steilacoom campus. Candidates will be working for a District Dean and will be required to attend district meetings as well as collaborate with colleagues across the district and across departments. Initial salary placement is based on the current full-time faculty salary schedule and is dependent upon education and experience. Salary placement for newly hired full-time faculty will be at one of five levels based upon qualifications including teaching experience and credentials. The five levels range from $50,305 to $54,498 per 173-day academic year. Opportunities for summer employment and other assignments are in addition to the base salary. Reasonable Accommodation:Applicants with disabilities who require assistance with the application and employment process will be accommodated to the extent reasonably possible. Requests should be made to the District Human Resources Office by calling 253-964-7342. Persons who are deaf or hard of hearing may dial 711 to place a call through Washington Relay, the state's free telecommunication access service Screening & Interview Process:Only those candidates who have complied with the application process and have met the required qualifications will be considered. The most desirable candidates will be contacted for an interview. Application materials submitted to Pierce College are the property of the College and will not be returned.
Conditions of Employment:
The Pierce College District is an equal opportunity employer. The Board of Trustees and Pierce College District are committed to providing equal opportunity and will not discriminate in terms and conditions of employment and personnel practices for all persons without regard to race, color, national origin, age, perceived or actual physical or mental disability, pregnancy, genetic information, sex, sexual orientation, gender identity, marital status, creed, religion, honorably discharged veteran or military status, or use of a trained guide dog or service animal.For more information or assistance regarding your application, contact Human Resources at: firstname.lastname@example.com regarding equal employment opportunities or discrimination may be directed to the following: Holly Gorski, Vice President of Human Resources/Title IX Coordinator9401 Farwest Drive SWLakewood, Washington 98498(253) 964-6519Paula Henson-Williams, District Affirmative Action Officer (253) 864-3229
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text:
The Health Care Institution/patient Relationship
Logue, Gerald
Wear, Stephen
Bibliographic Citation
In: Iltis, Ana Smith, ed. Institutional Integrity in Health Care. Boston: Kluwer Academic Publishers; 2003: 99-110.
Permanent LinkFull Text from Publisher
MetadataShow full item record
Related items
Showing items related by title, author, creator and subject.
A Desperate Solution: Individual Autonomy and the Double- Blind Controlled Experiment Logue, Gerald; Wear, Stephen (1995-02)The randomization ingredient in double-blind controlled experiments may be objectionable to patients who, in their desperation, come to such trials seeking a last chance of cure. Minogue
The Problem of Medically Futile Treatment: Falling Back on a Preventive Ethics Approach Wear, Stephen; Logue, Gerald (1995)
Toleration of Moral Diversity and the Conscientious Refusal by Physicians to Withdraw Life-Sustaining Treatment Wear, Stephen; Lagaipa, Susan; Logue, Gerald (1994-04)The removal of life-sustaining treatment often brings physicians into conflict with patients. Because of their moral beliefs physicians often respond slowly to the request of patients or their families. People in bioethics ...


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Ford Ranger Diesel Pick Up Super XLT 2.2 TDCi Lease
This deal is based on a non-maintained contract. Maintenaince is availble upon request
Share this deal!
|Insurance Group
|Engine Size (cc)
|1248 cc
|Number Of Seats
|Standard Manufacturers Warranty
|3 years
|MPG Urban
|45.6 mpg
|MPG Extra Urban
|70.6 mpg
|MPG Combined
|58.9 mpg
|3999 mm
|1488 mm
- Body Glass
- Electric windows, one touch lowering + anti trap device (Driver only)
- Heated front window
- Heated rear cab window
- Rain sensitive wipers and auto headlights
- Rear flip open windows
- Tinted glass
- Brakes
- Emergency brake assist
- ESP including traction control
- Communication
- Bluetooth hands free kit with voice control
- Driver Aids
- Driver Convenience
- Cruise control and variable speed limiter
- Easy fuel capless refuelling system
- Driver Information
- Digital tachograph
- Multifunction colour display
- Outside air temperature display with ice warning
- Seatbelt warning indicator
- Trip computer
- Tyre pressure monitor
- Driving Mirrors
- Auto dimming rear view mirror
- Chrome electrically adjustable and heated door mirrors with memory
- Electric folding door mirrors
- Electrochromic interior mirror
- Electrical
- 2 x 12V power outlets
- Embelishment Trims
- Centre console bezel with silver finish
- Entertainment
- 2 USB port
- 2 USB ports
- 6 speakers
- Aux input socket
- DAB with CD and MP3 player
- Steering wheel mounted audio controls
- USB connection
- Exterior Body Features
- Body colour bumpers
- Body colour wheel arch mouldings
- Chrome effect door and tailgate handles
- Chrome effect scuff plates
- Chrome front grille
- Rear mudflaps
- Steel double skinned pick up box with 2 position tailgate and inner fenders
- Under hood engine cover
- Exterior Lights
- Daytime running lights
- Front fog lights
- Headlamp levelling
- Heating/Cooling/Ventilation
- 4 speed air recirculation fan + heater
- Interior Features
- 2 sunvisors with ticket holders and passengers with mirror
- Carpet covered floor
- Carpet mats
- Driver and passenger sunvisors with illuminated vanity mirrors
- Front centre armrest with storage bin
- Front centre console with dual cup holders and armrest with double lidded storage box
- Front door storage pockets
- Front passenger grab handle
- High series finish instrument panel
- Illuminated glovebox
- Leather gearknob
- Leather wrapped steering wheel
- Overhead stowage with mood lighting
- Overhead sunglasses holder
- Penta cloth upholstery
- Reach and rake adjustable steering column
- Interior Lights
- Door operated dome light with courtesy switches on all doors
- Load area light
- Map reading lights
- Load Area
- High performance bedliner with protective side wall
- Interior tie down hooks and load rest
- Outer tie down hooks and load rest
- Safety
- 2x3 point rear seatbelts
- Driver and passenger airbag
- Front passenger airbag deactivation
- Front seatbelt pre-tensioners with load limiters
- Knee airbag
- Side curtain airbags
- Side Thorax Airbags
- Speed sensing door locks
- Seats
- 4 way manually adjustable driver seat with lumbar support
- Front seat back map pockets
- Rear jump seats
- Security
- Immobiliser
- Lockable tailgate
- Locking wheel nuts
- Remote central locking
- Thatcham category 1 alarm
With over 13 Years experience in the Contract Hire and Vehicle Leasing sector, Vehicle Savers have built long-lasting relationships and have partnered with some of the UK's finest organisation's and finance companies, allowing us to showcase some of the most competitive Car Leasing Deals in the UK.
At Vehicle Savers HQ we are all car fanatics, so knowlege comes second nature.
We take pride in our customer service and are always available to answer any questions you have. Even outside of working hours feel free to get in contact via our Facebook page for Vehicle Leasing related enquiries or Tweet us @VehicleSavers if you have anything fun to share.
Leasing a Ford Ranger Diesel Pick Up Super XLT 2.2 TDCi has never been easier on Business Contract Hire.
Business contract hire also known as (BCH) allows businesses to finance their vehicles with an agreed mileage and a minimum initial rental for the contract.
First choose your initial rental. The minimum initial rental on business contract hire is 3 months in advance or as high as 12 rentals upfront depending on the deal you choose.
Once you have decided you upfront initial rental it's time to choose your mileage. Mileages will range from as low as 5000 miles per annum all the way through to 50,000 miles per annum.
Then choose your term. The majority of deals advertised will range from 2-4 years.
The advantage of business contract hire is that it allows businesses to reclaim the VAT on their vehicle lease deal.
If you have any further queries please see our business contract hire and leasing guide.
Leasing a Ford Ranger Diesel Pick Up Super XLT 2.2 TDCi has never been easier on Personal Contract Hire.
Personal Contract Hire also known as (PCH) is very similar to business contract hire, with exception of reclaiming the VAT. All Personal Contract Hire deals are marked with 'Inc VAT'.
Personal Contract Hire is not a Personal Contract Purchase (PCP). However please feel free to get in touch if this is what you are seeking.
If you have any further queries please see our personal contract hire and leasing guide.


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Best Of :: Shopping & Services
Yeah, the tourists remain annoying, and the Mouse has turned into a real-life welfare queen gracias to massive subsidies by the Anaheim City Council in the form of a no-gate-tax pledge and a promise to hand over tax revenue in exchange for a new hotel. But remember when the Disneyland Hotel was run-down and grungy? Probably not because y'all are young, and we're old fuckers. The Mouse continues to do a magnificent job of tweaking its oldest hotel to keep up with the ever-growing Disnerd hordes, with themed rooms, better eats and the eternal party at Trader Sam's Enchanted Tiki Bar. And with the way CEO Bob Iger keeps swallowing up pop-culture icons under the Disney brands, expect Iron Man and Darth Vader to empty your chamber pot in a couple of years.
Readers' Choice: Montage Laguna Beach
It's really the only mall that showcases everything Orange County has to offer: ocean views, sunny skies, expensive restaurants and lots of dogs. On top of four major department stores—Bloomingdales, Macy's, Neiman Marcus and Nordstrom—Fashion Island is home to more than 100 other retailers. From affordable wear, designer goods and body-care products to cars, furniture and electronics, this shopping haven has you covered. Not in the mood for shopping? Take a ride at SoulCycle, relax at Big Newport, or kick back and watch people nearly trip into the koi pond—it happens more often than you think.
Readers' Choice: South Coast Plaza
All the boards UsedSurf takes in on consignment are in excellent condition and available via an online catalog—they have around 400 at any given time and will ship nationally. Or go to the San Clemente warehouse, a.k.a. the Factory, to browse in person or order a custom design from Edit Industries. UsedSurf stocks an extensive array of board fins and offers a board-rental service, too. Ask for Mo.
Readers' Choice: The Frog House
The name says it all, kids: SAVAGE!
Readers' Choice: E-cig Emporium
Ditch the time-consuming mess that happens daily with mascara and schedule an appointment with Megan Dunning at Luscious Lashes. Relax in a big, comfy recliner with a super-soft blanket while Dunning, who is somewhat of a perfectionist, works her eyelash-extension magic. When she's finished, be ready for all the compliments, as well as the questions as to where you got them done. The long, full lashes will last three to four weeks, and the best part is it looks as if you're wearing make-up when you're not.
A thousand pounds of Epsom salts will get anyone, no matter how dense the bones, a feeling of weightlessness. You will float. In your own private room, you step naked from the shower to the tank. The water is warm, and while you float, no sound or light will distract you from a trip into a theta-wave state of mind. Claustrophobes can float with the tank door wide open and advance to it being propped open a little bit with a towel. That zero-gravity sensation isn't only for space travelers.


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I find it a little suspicious this charity has zero reported fundraising costs. They have a Chief and a Manager on staff as a part of the leadership team who has DEVELOPMENT in their titles. Aren't their purpose within the organization is to raise funds and to manage the $6.2 million in contributions (non program service related)? I am pretty sure they are paid to do said job too. Even if a good percentage of their time can be allocated to programmatic or administrative purposes, their entire salary should not be allocated as solely either a programmatic or administrative expense. How are auditors letting them get away with that? Finally, when you go to their website, their donation page is powered by Fundly, which I know for fact charges money to provide its service. Isn't that alone a fundraising expense? So please tell me how you can conscionably report your fundraising costs as zero, Goodwill Industries of San Francisco.
Review from CharityNavigator
I've been with Goodwill for almost 3 years. I started working with Goodwill through the Community Jobs Program. Now I am a Assistant Manager and I work with and have met some great people. Goodwill has great programs to help Adults get back into the work force. I am so lucky I found Goodwill.
I wouldn't trade my experience working with Goodwill for anything. Goodwill is an amazing organization that brings together the talents and passion of professionals with those who need a helping hand. What a powerful thing it can be to give a hand-up rather than a hand-out.
I have been with Goodwill for 2 years. In that time, I have come to appreciate what a well run company, doing the work of caring for people, Goodwill is. For years all I knew about Goodwill was that one donated "stuff" got a tax credit. Now, I know of no other place that does a better job of creating ways to end poverty while making money. What a incredible experience it is.
I've worked at Goodwill for more than five years as office manager and executive assistant to the Chief of Organizational Advancement (marking/communications/fund raising), the Chief Operating Officer, the Chief Financial Officer and all of their direct reports. I have a pretty good "bird's eye view" of our day-to-day operations. We are truly one of the original "green" businesses. Not only do we recycle, reuse and give new life to literally millions of items each year, but we also take the people that society forgot and give them a leg up to sustainable careers -- not just low wage jobs. It's inspirational to see people rise above their employment barriers and shine in the workplace. It's great to be here.
I've worked in the nonprofit world for over 12 years and I'm very impressed with Goodwill of San Francisco's performance. They are a leader in safe-keeping our environment by recycling electronic waste and household materials including junk shoes and clothes. The best part is that they use the money claimed through recycling to serve the community by providing job search and training to just about any one who needs help. They don't just provide on the job training in their production and stores, but most needed training areas like refurbishing computers, computer repair, office technology, truck driving, and retail management. They structured a whole division called People Services just to server people. I think they've really got the right focus and direction in helping the people in the community and set a model for the rest of the country and the wold. They focus on people's needs.
I started working with and for Goodwill Industries of San Francisco, San Mateo and Marin nearly seven years ago. What an amazing journey this has been. I never expected a 95 year old organization, that is best known for its thrift stores and easy donation centers, to be doing such tranformation work for the Bay Area's poor and vulnerable communities. The quality of the job training, support, and education programs are outstanding. Additionally, Goodwill's commitment to growing environmentally sustainable businesses and hiring some people overcoming significan barriers to employment goes well beyond the scope of running quality programs. Goodwill is a vibrant social enterprise that is continuously innovating to better meet the needs of its communities.
Tom started at Goodwill almost five years ago as a client, and now I've worked my way up to Production Supervisor of our San Mateo warehouse. My daily tasks revolve around trying to control chaos!
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Corrie: Landfill Search Ends; No Trace
Police say search of the Milton landfill site cannot continue after officers conduct fingertip search since March
A man from Suffolk found in a submerged car after disappearing from a stag night might have driven into the river without realising it was there, an inquest has heard.
25-year-old Charles Watson from Henstead, was found in his car in the New Cut waterway between the rivers Yare and Waveney in St Olaves, Norfolk on April 13th.
Mr Watson, known as Sam, was last seen on a night out in Great Yarmouth at 1.40am on April 9th.
An inquest at Norwich Coroner's Court heard that the alcohol level in his body was almost three times the legal drink-drive limit. Friends said that he seemed in good spirits and in control of himself.
Coroner William Armstrong said his alcohol consumption may have contributed to the accident and hindered any attempt to escape as the car flipped and sunk.
Crash investigator Sergeant Andy Hood said that evidence suggests Mr Watson attempted to escape but, disorientated by being upside down and by alcohol, had accidentally lowered the wrong electric window.
"The car would have filled quickly with water", he said. "There were no mechanical defects with the car. Because of the way the road is, drivers can't always see the river beyond," Sgt Hood said.
Mr Watson was out celebrating at his friend Paul Miller's stag do. At the end of the night Mr Miller's father-in-law offered the group lifts home but Mr Watson declined.
Mr Watson arranged to stay with a colleague after the night out. He sent a text message at 3.15am saying that he found somewhere else to stay. Police believe the car entered the water at 3.16am.
One theory is that he stopped to send the message from the side road and as he turned around, accidentally drove his Volkswagen Golf into the river which is up to 15 metres deep. He was found four days later by Pc Nafica White.
Mr Armstrong recorded a verdict that Mr Watson died an accidental death from drowning.
An inquest is going to be held today into the death of a man found in a car in a river after he vanished from a stag night in Great Yarmouth.
Charles 'Sam' Watson, 25, of Henstead in Suffolk, was found in his car submerged in the New Cut at St Olaves, Norfolk, on April 13.
He was last seen on a night out in Great Yarmouth at 2.30am on April 9th and was reported missing by his family the following day.
Police launched an investigation into what happened to Mr Watson between the time he left the party and the discovery of his body in the River Waveney.
Police say search of the Milton landfill site cannot continue after officers conduct fingertip search since March
A group of men have been convicted for their part in an attack on a football fan that left him with permanent brain damage.
A man's been arrested after shots were fired in Ipswich this morning.
Shots have reportedly been fired in Ipswich this morning during a fight between a group of people in Cardinal Park.
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Novel aspects of grapevine response to phytoplasma infection investigated by a proteomic and phospho-proteomic approach with data integration into functional networks
Translational and post-translational protein modifications play a key role in the response of plants to pathogen infection. Among the latter, phosphorylation is critical in modulating protein structure, localization and interaction with other partners. In this work, we used a multiplex staining approach with 2D gels to study quantitative changes in the proteome and phosphoproteome of Flavescence dorée-affected and recovered 'Barbera' grapevines, compared to healthy plants.
We identified 48 proteins that differentially changed in abundance, phosphorylation, or both in response to Flavescence dorée phytoplasma infection. Most of them did not show any significant difference in recovered plants, which, by contrast, were characterized by changes in abundance, phosphorylation, or both for 17 proteins not detected in infected plants. Some enzymes involved in the antioxidant response that were up-regulated in infected plants, such as isocitrate dehydrogenase and glutathione S-transferase, returned to healthy-state levels in recovered plants. Others belonging to the same functional category were even down-regulated in recovered plants (oxidoreductase GLYR1 and ascorbate peroxidase). Our proteomic approach thus agreed with previously published biochemical and RT-qPCR data which reported down-regulation of scavenging enzymes and accumulation of H2O2 in recovered plants, possibly suggesting a role for this molecule in remission from infection. Fifteen differentially phosphorylated proteins (| ratio | > 2, p < 0.05) were identified in infected compared to healthy plants, including proteins involved in photosynthesis, response to stress and the antioxidant system. Many were not differentially phosphorylated in recovered compared to healthy plants, pointing to their specific role in responding to infection, followed by a return to a steady-state phosphorylation level after remission of symptoms. Gene ontology (GO) enrichment and statistical analysis showed that the general main category "response to stimulus" was over-represented in both infected and recovered plants but, in the latter, the specific child category "response to biotic stimulus" was no longer found, suggesting a return to steady-state levels for those proteins specifically required for defence against pathogens.
Proteomic data were integrated into biological networks and their interactions were represented through a hypothetical model, showing the effects of protein modulation on primary metabolic ways and related secondary pathways. By following a multiplex-staining approach, we obtained new data on grapevine proteome pathways that specifically change at the phosphorylation level during phytoplasma infection and following recovery, focusing for the first time on phosphoproteome changes during pathogen infection in this host.
Flavescence dorée (FD) is an economically important grapevine disease in Europe caused by phytoplasmas, plant phloem-limited pathogens in the class Mollicutes
Over recent years, genomic and proteomic strategies have been successfully used to analyse plant-pathogen interactions. In particular, proteomic approaches have largely evolved in pursuit of the functional assignment of proteins expressed during phytopathogenic interactions
Studies on post-translational modifications (PTMs) are considered important in clarifying pathogen/host interactions, and their role in the dynamic adaptation of plants to different conditions has been demonstrated
In this work, we monitored changes in the total proteome and phosphoproteome of FDp-healthy, infected and recovered grapevines through different staining techniques. Spots showing qualitative and quantitative changes were trypsin-digested and the resulting peptides analysed by Matrix-Assisted Laser Desorption Ionization-Time Of Flight (MALDI-TOF-TOF). The differentially expressed proteins were integrated into functional networks in order to represent their biological significance in the host-pathogen interaction. Our data gave new valuable insights into specific protein modulations that occur in grapevine during progression of Flavescence dorée and after recovery from the disease, providing valuable proteomic data on this poorly characterized phenomenon.
Results and discussion
Grapevine midrib tissues from healthy, infected and recovered plants were considered for the extraction of total proteins. In order to minimize the environmental and sample variability we collected the plants in the same vineyard area, nearby each other and in the same day. Grapevines were scored for symptoms and screened for phytoplasma and virus infection by molecular assays, as described in Methods section. In all the samples, we excluded the presence of seven out of the eight viruses for which we tested, with the exception of Grapevine fleck virus (GFkV), which was present in each vine in the vineyard. Furthermore, its presence was not expected to interfere with our investigation, as GFkV is generally latent and asymptomatic in grapevines
Figure 1
2-DE proteomic maps of proteins and phosphoproteins of
2-DE proteomic maps of proteins and phosphoproteins of
For spot identification, the 2-DE electrophoresis was coupled with MALDI TOF/TOF analysis. This system is suitable for the identification of the most abundant proteins in the spot analysed. However, this technical limit did not hamper the ability to provide a description of the main changes that occur in the grapevine proteome and phosphoproteome during phytoplasma infection and recovery (Figure
Figure 2
Differentially expressed/phosphorylated proteins in phytoplasma-infected (I) and recovered (R) grapevines in comparison to healthy (H) ones.
Differentially expressed/phosphorylated proteins in phytoplasma-infected (I) and recovered (R) grapevines in comparison to healthy (H) ones. Differentially expressed proteins (S) have been revealed by Sypro® Ruby stain and differentially phosphorylated proteins (P) have been revealed by ProQ Diamond stain. For each protein spot ID, number of isoforms, GenBank code and fold change are reported. Red: up-regulated/up-phosphorylated. Blue: down-regulated/down-phosphorylated. Green shows chloroplast-localized
Additional file 1
Detailed list of the differentially expressed proteins. The following parameters are reported for each spot: sequence of the matched peptides, normalized volume values (PPM) and coefficient of variation, statistical significance. Sheet 1: healthy/infected Sypro® Ruby staining (total proteins); sheet 2: healthy/recovered Sypro® Ruby staining; sheet 3: healthy-infected ProQ Diamond staining (phosphoproteins); sheet 4: healthy-recovered ProQ Diamond staining.
Click here for file
In Figure
Figure 3
Protein distribution and expression trend in infected and recovered plants.
Protein distribution and expression trend in infected and recovered plants. A) Venn diagram showing the distribution of the differentially expressed proteins in each category. B) Induced and repressed grapevine proteins in response to infection and in recovered plants. Inf: infected. Rec: recovered. S: Sypro® Ruby staining (total proteins). P: ProQ Diamond staining (phosphorylated proteins).
Proteome and phosphoproteome changes in FDp-infected grapevines
Proteome changes
A previous study analysed the response of the 'Nebbiolo' grapevine proteome to phytoplasma infection using a Coomassie Blue staining approach
Twenty-nine out of the 37 differentially expressed proteins identified in FDp-infected samples were up-regulated, whereas 8 showed a down-regulation with respect to healthy plants (Figure
Figure 4
Grapevine proteins with predicted plastid localization changing their quantitative and/or phosphorylation levels in infected and recovered plants.
Grapevine proteins with predicted plastid localization changing their quantitative and/or phosphorylation levels in infected and recovered plants. The model gives an overview of the quantitative and qualitative (phosphorylation) changes in chloroplast-localized proteins, showing their involvement in specific metabolic pathways and predicted integration into biological networks according to the KEGG database. The expression trends are represented using a coloured legend in the upper left corner of the figure.
Multiple isoforms of enolase and aconitase enzymes were found to be differentially expressed in infected leaves. The enolase protein has been found to be responsive to several biotic
Spot 2602 was identified as being isocitrate dehydrogenase. This protein shares 89% identity with the homologous protein in
Several pathogenesis related proteins belonging to class PR-5 (thaumatins, osmotins), PR-10 and PR-17 (plant basic secretory protein) were up-regulated in infected tissues. PR-5 proteins were previously reported in grapevines infected by FDp
Spot 3304 corresponded to an aluminium-induced protein. The same protein was found to be up-regulated in the grapevine cv. Razegui under salt stress
Consistent with the importance of plant cell wall modifications in response to pathogen invasion, the up-regulation of proteins involved in cell wall re-organization was present in FDp-infected grapevines. An alpha endo-1,3;1,4-beta-D-glucanase (spot 6308), a beta 1,3-glucanase (spot 7403) and two glycosyl hydrolases (spots 5510 and 1201) were up-regulated. Our data are consistent with the results of previous mRNA analyses of grapevine cultivars
Among the responsive proteins identified in infected 'Barbera' plants, eight proteins have already been reported in FDp-infected 'Nebbiolo'
Phosphoproteome changes
Statistical analysis identified 15 differentially-phosphorylated proteins in infected vs. healthy plants (Figure
Alteration of the light-chain reaction is strictly related to alterations in electron transfer and generation of reactive oxygen species
Peroxiredoxins (Prx) are a family of peroxidases found in all organisms and represent central elements of the antioxidant defence system
A PAP-fibrillin protein was previously found to be up-regulated in grapevine tissues infected by phytoplasmas
Among the proteins involved in response to stress, an HSC70-1 protein was up-phosphorylated. These proteins are involved in protein targeting and degradation
Proteome and phosphoproteome changes in recovered plants
Proteome changes
Analysis of the whole proteome of recovered plants showed differential expression for 24 proteins when compared to healthy grapevines. Six were common to the 37 proteins found in the comparison between healthy and infected plants. Among them, the plant basic secretory protein (gi|147784683), translationally-controlled tumour protein (gi|225432248) and putative acidic endochitinase (gi|225454408) were up-regulated. As these proteins are reported to be involved in the general response to stress (Additional file
Additional file 2
GO classification and KEGG map assignments of the identified
Click here for file
Most of the remaining 17 proteins, uniquely found in recovered plants, were down-regulated compared to healthy plants. Among them, we found an ascorbate peroxidase (APX), a major enzyme involved in H2O2 scavenging. Previous studies of grapevine have suggested that recovery from phytoplasma disease is associated with down-regulation of enzymatic H2O2 scavengers, supported by direct evidence of the accumulation of H2O2 in the leaves of recovered plants but not in healthy or diseased plants
Phosphoproteome changes
Twelve proteins were differentially phosphorylated between recovered and healthy plants. Nine of these were also differentially phosphorylated in infected tissues: 8 displayed an increase in phosphorylation levels, while the auxin-induced protein PCNT115 isoform 1 was the only to show a decrease in phosphorylation levels in recovered tissues. The remaining proteins which were phosphorylated only in recovered plants, in comparison to healthy and FDp-infected plants, included: sedoheptulose-1,7-bisphosphatase, mitochondrial-processing peptidase and inorganic pyrophosphatase (Figure
Comparison with phosphorylated proteins in other plant species and prediction of Serine/Threonine/Tyrosine phosphorylation sites
The differentially phosphorylated proteins of
GO functional analysis, enrichment and integration of proteome data into biological interaction networks
In order to identify possible mechanisms of regulation or functional pathways that were specifically activated in our experimental conditions, differentially expressed proteins were associated to GO terms using the plant GOslim ontology (Additional file
Enrichment analysis applied to the GO cellular component terms showed that "plastid" (GO:0009536), a child category of "organelle", was over-represented both in infected and in recovered plants (Figure
Figure 5
GO classification and GO enrichment analysis of differentially expressed proteins in infected and recovered plants.
GO classification and GO enrichment analysis of differentially expressed proteins in infected and recovered plants. A) Cellular component term assignment and distribution. B) Biological process term assignment and distribution. The statistical significance of the Enrichment Analysis is represented by a scale of red tones whose intensity is proportional to the degree of significance starting from FDR ≤ 0.05.
Additional file 3
List of the over-represented GO categories in infected and recovered plants following Singular Enrichment Analysis. Proteins are grouped according to the GO categories to which they belong. Only the statistically significant (False discovery rate: FDR ≤0.05) GO terms are shown. "Category" indicates the main GO categories: "B", biological process; "C", cellular component. There were no statistically significant molecular function terms. "Number of sequences" indicates the number of proteins assigned to each GO term.
Click here for file
The same statistical analysis was applied to the GO biological process terms (Figure
Regarding the stress/defence responses, the GO categories "response to stimulus" (GO:0050896) and its child categories "response to abiotic stimulus" (GO:0009628), "response to biotic stimulus" (GO:0009607) and "response to stress" (GO:0006950) were over-represented in infected plants (Figure
Figure 6
Expression levels of proteins in the "response to stimulus" category in infected and recovered plants compared to healthy plants.
Expression levels of proteins in the "response to stimulus" category in infected and recovered plants compared to healthy plants. Singular Enrichment Analysis of GO categories distribution showed that the specific category "response to stimulus" (GO:0050896) was over-represented in both infected and recovered plants. The majority (71%) of the proteins belonging to this biological process was up-regulated in the infected plants, while, in contrast, 50% of the proteins was down-regulated in recovered plants compared to healthy plants. For each specific protein the fold change in expression value is reported compared to healthy plants (reference value = 1).
In this work we have successfully used a multiplex staining approach to deeply characterize the proteome and phospho-proteome of grapevine in response to phytoplasma infection by 2-DE. Using Sybro Ruby dye, we were able to describe 37 proteins which were significantly differentially expressed between healthy and infected plants. Furthermore, by using the ProQ Diamond dye we described for the first time phosphorylation changes (15 proteins) in grapevine during pathogen infection. Based on evidences from the literature, some phosphorylated proteins were previously reported to be regulated following abiotic stress in other plants
Figure 7
Mapping of the differentially regulated metabolic proteins during FDp infection according to KEGG "Metabolic pathways - Reference pathway (KO)".
Mapping of the differentially regulated metabolic proteins during FDp infection according to KEGG "Metabolic pathways - Reference pathway (KO)". The web-based server KAAS was used to display KEGG map assignments of the differentially regulated proteins. The "Metabolic pathways" map (in various colours) was the one that included the majority of the protein hits (1–15, in black). 1: anthocyanidine synthase; 2: acetyl-CoA acetyltransferase; 3: aconitate hydratase 2; 4: enolase; 5: 2,3-bisphosphoglycerate-independent phosphoglycerate mutase; 6: alcohol dehydrogenase 6; 7: isocitrate dehydrogenase; 8: succinate dehydrogenase; 9: ATP synthase CF1 alpha subunit; 10: ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit; 11: ribose-5-phosphate isomerase; 12: glutamine synthetase; 13: cystathionine beta-synthase (CBS) and cysteine synthase-like protein; 14: oxygen-evolving enhancer protein 2 23-kDa polypeptide of photosystem II; 15: ATP synthase CF1 beta subunit.
Plant material collection
Field-grown grapevine samples were collected in Cocconato, Asti province (Piedmont, north-western Italy), in summer (August) 2009. The vineyard had been monitored since 2005 for phytoplasma infection and a map of the sanitary status of the plants (healthy, infected and recovered) was available at the beginning of this study. All samples were collected based on symptom observation and previous monitoring data: healthy plants did not show any suspicious symptoms, whereas infected plants showed at least three typical symptoms of phytoplasma infection
Protein extraction and 2-DE
Total proteins were extracted using a TCA/acetone method
Multiplex staining and image analysis
After 2 -DE, the gels were treated with fixing buffer (50% (v/v) methanol, 10% (v/v) acetic acid) and then washed three times in water for 10 min. To detect phosphoproteins, the gels were stained using the Pro-Q Diamond phosphoprotein gel stain kit (Invitrogen). The manufacturer's protocol was followed, except that half (50 ml) of the recommended quantity of stain was used, in order to reduce the background, as shown by our preliminary work and other authors
Total proteome analysis of SyproRuby-stained gels was performed by creating a statistical (ANOVA test, p < 0.05) and a quantitative (volume variation by at least a factor of two) analysis set. Next, a Boolean analysis set that identified the spots common to the previous sets was created to select the differentially expressed spots. Phosphoproteome analysis was performed according to published methods
Mass spectrometry
Mass spectrometry was performed at the Proteomics Technology Facility, Department of Biology, University of York (
The FlexAnalysis software version 3.3 (Bruker) was used to perform the spectral processing and peak list generation. The mass spectra were analysed using the MASCOT peptide sequence matching software (Matrix Science Ltd., version 2.1) through a Bruker ProteinScape interface (vs. 2.1), against the NCBI
Only peptide mass maps that matched single annotated
Bioinformatic analysis
The web-based functional annotator KAAS (KEGG Automatic Annotation Server) was used to assign the differentially regulated proteins to KEGG pathways.
To identify newly reported phosphorylated proteins, differentially phosphorylated proteins were analysed by the Phosphoprotein BLAST available on the P3DB database (
The presence of potential Serine/Threonine/Tyrosine phosphorylation sites was predicted by NetPhos 2.0 software (
Competing interests
The authors declare that they have no competing interests.
Authors' contribution
PM participated in the design of the study, collected samples, carried out the proteomic analysis, participated in the bioinformatic analysis, and wrote the manuscript; SA performed the bioinformatic analysis and helped with manuscript writing; SP conceived the study and its design, coordinated the activities, and drafted and edited the manuscript. All authors read and approved the final manuscript.
The authors thank Dr. Cristina Marzachì, Dr. Massimo Turina and Dr. Marta Vallino for critical reading of the manuscript, and A. A. Dowle at the Proteomics Technology Facility, Department of Biology, University of York for mass spectrometry services. This work was supported by funding from "Servizio fitosanitario regionale", Regione Piemonte, Italy (Rep. 12851).
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