There it is in an ad for a fancy Swiss watch: three of the top soccer players on the world's No. 1 sports franchise, wearing bright red jerseys emblazoned with the logo of AIG.
Yes, all through the financial meltdown and the revelations about American International Group Inc.'s disastrous derivatives trading, the jerseys of England's Manchester United displayed those three tall white letters across the chest.
Now that's about to change, raising the hopes of fans who want more sweeping changes as well.
Starting June 1, Chicago's Aon Corp. will take over what is arguably the most prominent walking billboard in sports, at a cost of $80 million over four years.
A new "Aon" jersey won't debut until mid-July, after all the hoopla from the impending World Cup dies down.
The deal comes amid fan discontent with Man U controlling shareholder Malcolm Glazer, the Florida billionaire who also owns the National Football League's Tampa Bay Buccaneers.
Glazer's acquisition of the "Red Devils" put the team deep in debt. Restive followers have been pushing for a new owner, and some have called on sponsors to intervene.
Into this high-stakes drama steps Aon, the insurance broker that traces its roots to W. Clement Stone, who famously put his "positive mental attitude" to work selling policies.
Uberfan Sean Bones of the Manchester United Supporters Trust warns that the value of Aon's sponsorship could fall. "It's something they have to look at very seriously," he said.
Aon has spent the past year developing its Man U marketing strategy, and it has no plans to intervene in running the team, said spokesman David Prosperi.
"It's not our place to weigh in," he said. "We didn't do it because of the owners or because of one or two great players. What they represent in the global market transcends everything else."
With AIG in no position to renew its four-year sponsorship after receiving a $180 billion federal bailout, Man U sent several possible successors a memorable sales pitch. A pair of framed prototype jerseys arrived last spring at Aon's Chicago headquarters for Chief Executive Greg Case and marketing chief Phil Clement, who jumped on a plane to England.
"He came back convinced," Prosperi said. Within a few weeks, on June 3, Aon announced the deal.
Unlike so many recession-battered companies, Aon had the money, as well as a strategic objective. Having jettisoned low-margin underwriting for more profitable reinsurance, the company wants global clients to view it as a broad-based professional-services consultancy.
Competing insurance brokers are seeking greater visibility, too, including Willis Group Holdings PLC, best known in Chicago for renaming Sears Tower.
"We think this is a lot bigger than putting your name on a building," Prosperi said. "There will be millions and millions of impressions."
In 2009, Man U sold more than triple the sales of all NFL jerseys, Prosperi said. Along with its name on the shirt and seats at the matches, Aon already is staging client events such as exhibition games and training programs.
"There was a method to the madness of spending what some would see as an exorbitant amount of money," said David Herman, who runs the team's U.S. supporters' club.
Some fans will be grateful to get rid of the tainted AIG, Herman noted. As for welcoming new corporate sponsor Aon, "a good majority generally doesn't care," he said.
Still, the deal puts a company no one would mistake for a household name front and center.
"It's not about selling shirts," Herman said. "The brand resonance is seismic."
I believe Aon's contract was $160,000 over 4 years.
With no disrespect intended, please do not refer to Manchester United as "Man U," because it not only shows your ignorance, but it also discredits your entire article.
The biggest miss in your article was that you failed to inform your readers by mentioning that the American tax payers paid almost $21 million of their hard-earned tax dollars to Manchester United this passed year.
I would highly recommend that you take time out of your busy schedule this summer, and go see the World's most valuable sports franchise in your backyard.
Hopefully after that experience, you will begin to understand the brand recognition, and the global power that Manchester United Football club has.
For your reading, I have attached an article that will hopefully shine some light on why people should never refer to Manchester United as "Man U" as you did five times in your article.
Manchester United fans should never refer to themselves, or the club as "Man U," because it is derogatory insult towards the club.
The term is only used by supporters of other clubs as a complete and utter insult to our Manchester United. The "U" is meant to be "you" by the rival fans.
An early example of its usage is this chant by West Brom fans: "Duncan Edwards is manure, rotting in his grave, man you are manure - rotting in your grave". The origin of "Man U" is a song to insult the dead Duncan Edwards.
Liverpool and Leeds fans copied this with their own man you /u versions to insult all of the lads who died at Munich: "Man U, Man U went on a plane Man U, Man U never came back again" and "Man U Never Intended Coming Home" (if you combine the first letter of each word you get the word "MUnich").
This makes it a lot clearer that saying "Man U" is an insult, particularly to the older supporters and to see United fans using it now is shameful.
If you are a true Manchester United supporter, you will have read this and repost this so all United fans can spread the word, because ….. WE ARE MANCHESTER UNITED!
GLAZER OUT! AON OUT! LUHG!
Well it is great to see Americans taking an interest in something outside their own Country. I suspect that this article is written because Aon has its HQ in the writers fair city , so its only correct that this sort of news does make the local papers.
Do not doubt the value on a global scale of this arrangement for Aon and lets see what additional "connections" they make from just the Logo on the shirt as that will be the true test of how the average supporter/ or prospect makes that judgement on "what or who is an Aon "
Great loved it, will be waiting for your future posts Thank you for sharing.


21,163 Square Feet, comprised of 1,596 Sq.Ft. of main floor showroom that can be converted to office space, 18,297 Sq.Ft. of warehouse space, 753 Sq.Ft. of second floor office space and 517 Sq.Ft. of mezzanine storage. Approximately 2 ACRES OF PRIVATE, PAVED, FENCED YARD STORAGE. Warehouse features 2 dock loading bays and 2 drive in bays and a sump. Located in Meridian, easy access to main routes of Barlow Trail, Deerfoot Trail and Trans Canada Highway.


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Hidden-Gem Restaurants at Orlando's Top Resorts
By Nate Shelton on Jan. 17, 2018
Throughout the Orlando destination, culinary delights can be found at hidden-gem restaurants inside top hotels and resorts, including Hemisphere Restaurant at Hyatt Regency Orlando International Airport (pictured; photo by Michael Stavaridis).
With more than 5,000 restaurants throughout the destination, Orlando presents a vast array of dining options. Many of our best eateries are easy to track down, but as visitors and locals have discovered, numerous hidden gems can be found inside our resorts and hotels — including these five standouts.
Please note: Depending on the timing of your visit, some experiences may be temporarily modified or closed. Learn more about healthy travel and what's open in Orlando, and check with your favorite restaurants and other businesses for their current status.
Hidden-Gem Resort Restaurants in Orlando's Tourism District
Hemisphere Restaurant at Hyatt Regency Orlando International Airport (Photo: Michael Stavaridis)
Hemisphere Restaurant at Hyatt Regency Orlando International Airport
Led by Executive Chef Jefferey Powell, Hemisphere at Hyatt Regency Orlando International Airport focuses on robust, international flavors; seasonal, local ingredients; and careful pairings of select wines and cocktails. The restaurant also features incredible, panoramic views of Orlando International Airport and the surrounding landscape.
Tabla Restaurant at Clarion Inn & Suites Orlando
Tabla at Clarion Inn & Suites Orlando presents a symphony of Indian, Chinese and Thai flavors, with dishes designed to delight neophytes and connoisseurs alike. What's more, their ambience, service and presentation are every bit as good as the food, all of which earned them a TripAdvisor Certificate of Excellence Award in 2016.
The Chef's Table at The Edgewater and The Tasting Room
Located inside The Historic Edgewater Hotel in Winter Garden, about 25 minutes from Disney, this is technically two restaurants in one. In the main dining room, enjoy an ever-changing, three-course, prix-fixe meal with optional wine pairings. Also on the premises, The Tasting Room features small plates, cocktails, beer, wine and entertainment.
Learn more: Winter Garden
Hidden-Gem Resort Restaurants in Downtown Orlando & Winter Park
The Boheme at Grand Bohemian Hotel Orlando
The Grand Bohemian Hotel is arguably downtown Orlando's most prestigious hotel, and its onsite restaurant has much to do with its refined reputation. Like the hotel itself, The Boheme is tastefully decorated with an eclectic array of art. More to the point, Chef Laurent Hollaender and his staff present deliciously inventive takes on classic dishes, such as Colorado Lamb T-Bone and Beef Tenderloin Au Poivre.
Learn more: Downtown Orlando
Hamilton's Kitchen at The Alfond Inn in Winter Park
Hamilton's Kitchen at The Alfond Inn
In addition to displaying an exquisite collection of contemporary art on loan from Rollins College's Rollins Museum of Art, The Alfond Inn in Winter Park, about 20 minutes from downtown Orlando, is also home to one of the area's best eateries. Hamilton's Kitchen specializes in traditional Southern fare fusing locally sourced ingredients with rustic, open-table charm.
Learn more: Winter Park
Stay Connected to Orlando!
You never know what we'll dream up next. Connect with Visit Orlando's Vacation Planning Services for personalized assistance, and follow us on Facebook, Twitter, Instagram, Pinterest and YouTube for the latest offers, exclusive tips, newest attractions and fun events. You can also subscribe to our eNewsletter for more information from Orlando: Theme Park Capital of the World.
Categories: Dining & Nightlife
Tags: Alfond Inn, Boheme, Chef's Table, Clarion Inn & Suites Orlando, Downtown Orlando, Edgewater Hotel, Grand Bohemian Hotel Orlando, Hamilton's Kitchen, Hemisphere Restaurant, Hyatt Regency Orlando International Airport, Neighborhoods, Places to Stay, Tabla Restaurant, Winter Garden, Winter Park
Nate Shelton
Content senior manager at Visit Orlando. He's a native Floridian and longtime Orlandoan who enjoys good food and drink, live entertainment, music, theme parks, comic books, video games, movies, and all the other things that make life worth living.

redpajama_set_name: RedPajamaCommonCrawl


This is a test post using Squarespace iOS app.
Testing Photo Uploads using a picture from my iPhone Camera Roll.
Posting a picture and moving it around works fine but I can seem to find a way to re-size it.
The app like all of Squarespace seems to works fine.


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E3 2018: Microsoft Announces Press Conference Date and New Venue
The press conference will be held on June 10 at 1pm PT / 4pm ET / 9pm UK (June 11 at 8am in Australia).
By AlexGilyadov
Updated: 14 Mar 2018 3:42 pm
Posted: 14 Mar 2018 3:33 pm
Microsoft has revealed when and where it will host its E3 press conference this year.
As detailed on a recent blog post, the Xbox E3 2018 Briefing will be held on June 10 at 1pm PT / 4pm ET / 9pm UK (June 11 at 8am in Australia) in the Microsoft Theater, located across the street from the Los Angeles Convention Center.
Along with the press conference, the Microsoft Theater will also host various Xbox FanFest activities, hands-on gameplay and demos for attendees, and more throughout E3. Microsoft promises this new venue will allow the company to "include even more fans and partners in the Xbox E3 2018 Briefing than ever before."
Microsoft will also continue to have a presence at the Los Angeles Convention Center with a new booth dedicated to Mixer. Attendees will be able to play, interact with, and stream games.
E3 2018 will be held from June 12-14. Check out what games are rumored to be at the event and check back here for all the press conference and live stream times. Also find out about EA's plans for E3 2018.
Alex Gilyadov is a freelance writer for IGN. Be sure to check out his YouTube channel and follow him on Twitter.


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Spain | Español
Welcome to your Red Folder
Is Your Marketing in the Right Place but at the Wrong Time?
Proliferation of digital channels and options for customers raises the importance of an underappreciated variable: message timing.
By Laura Beaudin and Francine Gierak
This article originally appeared on HBR.org.
Time is running out on personalized marketing as a means of continually raising the return on investment of campaigns. The use of advanced data analytics to identify the right customers, while still valuable, is reaching a plateau. Proliferation of digital channels and options for customers raises the importance of another, still underappreciated, variable: message timing.
By communicating at the most opportune times based on insights into consumer behavior, companies can generate more business with fewer or more efficient ads, or expand their audience to find unexpected wins.
Timing comes into play through signals, sequence and speed. And the technology now exists for marketers to test with high confidence when to communicate and in what order—and to do so in near real time. How exactly are they doing that, and thereby realizing further gains in ROI? Bain & Company recently surveyed nearly 1,700 marketers globally, in partnership with Google, and found three areas of importance.
Signals. By understanding consumer intent, marketers can identify the right moments in the customer journey where ads will be most relevant.
In the market for cold medicines, for example, a leading global brand mapped its customers' past search and purchase behavior, determining that people who take their cold medication in the first two days of symptoms are more likely to get relief and become greater brand advocates. Knowing this, the company worked with Google and WebMD to reach consumers whose search terms suggested they had common cold symptoms in that narrow period. As a result, the brand substantially increased conversion of searches to sales of its cold medication, building greater brand loyalty at the same time.
Sequence. Once a company spots the crucial signals, it should deliver a sequence of messages tailored to each customer.
A global sporting goods manufacturer used to mix brand-oriented, aspirational messages and price-focused promotional messages during the same time frame, in part because its e-commerce and brand teams did not coordinate with each other. Yet leading with a promotional message, while it might generate a short-term sales boost, sacrifices building long-term brand value. By contrast, leading with brand messages early in a campaign can have a halo effect on the subsequent promotional messages.
This company now runs campaigns jointly between the two teams, sending and testing messages in a carefully calibrated sequence. As the senior director of digital media told us, "We can sequence our ads to start with a brand message for our most inspirational product, then start promoting a different product with an offer attached if the customer doesn't bite. This saves money [on promotions] and ensures our positioning remains premium with those for whom our brand really resonates."
Speed. Timing also affects how quickly a company is able to react to a customer's behavior. Faster reaction times go a long way toward more effective marketing.
When companies get more proficient in timing their marketing activities, they often realize another benefit: freed-up time for brand building, creative development and data mining. The digital director of the Dutch cosmetics retailer Rituals noted that improving its customer prospecting gives the firm more time to "develop creative and ad messaging that is most true to our brand."
What Marketing Leaders Do Differently
Advanced marketers have already begun to master signals, sequence and speed. For these firms, our research identified improved understanding and selection of customers as being their single-greatest investment area over the next three years. But while all marketers share this goal, the leaders (defined as the top 20 percent of companies based on a composite score of revenue and market share growth), have a sharper focus.
Leaders own their digital destiny by taking control of their consumer data and by integrating their marketing and advertising technologies. For example, leaders in North America are 1.6 times more likely than laggards to prioritize integrating their platforms. European marketers rely more on outside agencies, but a majority still value integration, and 85 percent of Australian marketers are looking to integrate.
The leaders have built a test-for-results culture, refreshing metrics and dashboards at least weekly and using data to directly inform decisions. Leaders also make decisions faster, in part by having the in-house team responsible for customer acquisition and retention take more accountability for budgets, technology and analytics. There are regional differences, of course—slower data refreshes in Europe, more budget constraints in Australia—but the trends for leaders apply globally.
Companies that take more control of their marketing and advertising data and technology will be able to respond quickly to customer insights and send a tailored message at precisely the right moment. That raises the odds of seizing attention for the immediate purchase and building brand equity in the longer run.
Laura Beaudin is a partner and Francine Gierak is a principal with Bain & Company's Customer Strategy & Marketing practice. Beaudin leads Bain's Marketing Excellence practice.
Laura Beaudin
Partner, San Francisco
It's About Time: Why Your Marketing May Be Falling Short
Discover how leading marketers are redefining time.
How CMOs Can Get—and Keep—Their Marketing Mix Right
All CMOs have access to the same tools, but the leaders focus on the full range of conditions necessary for results.
Laura Beaudin: Don't Miss the Moment on Mobile Marketing
Three ways marketers can capitalize on the prevalence of mobile marketing to connect with consumers.
Consumers Move Online to Buy Appliances
Covid-19 is accelerating the shift to digital marketing of white goods.
Covid-19 Catapults White Goods Makers into the Digital Future
As consumers shift to buying appliances online, leaders develop direct-to-consumer channels.
Direct marketing excellence through experimental design
A major retailer rejuvenates sales by thinking local
How to sell more dog food
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Merino share of lamb slaughter increasing, with intentions to increase further
By Angus Brown | Source: MLA, AWI
The MLA-AWI sheep supply survey is conducted every four months, and produces a range of data.
There appears to be a trend towards slaughtering more merino lambs, especially in Victoria.
Strong wool prices could see lamb supply ease when combined with a good season.
Every four months, Meat and Livestock Australia and Australian Wool Innovation survey sheep producers regarding their sheep production and their intentions for the coming four months. While the number of respondents vary, and some of the results can be misleading, there are some trends we can identify that have impacted markets.
There is plenty of data produced in the MLA-AWI survey, including ewes joined, lambs marked, and wether and lamb selling intentions, all broken down by state. One interesting trend has been the increase in Merino lambs that are intended to be sold for slaughter.
Figure 1 shows the lamb sales intentions per respondent over the last four years. Interestingly, the average operation size of respondents is large compared to the 'average' sheep and lamb operation. Lamb turnoff per respondent ranges between 3,000 head and 6,500, depending on the season.
The number of Merino lambs intended to be sold for slaughter has risen 9% from the Jun-11 to Feb-12 period through to Jun-14 to Feb-15. This increase has compensated somewhat for the 18% fall in 'other' lamb slaughter intentions over the same period.
The fall in 'other' lamb slaughter intentions highlights some of the issues with the survey data, as 2014-15 is likely to be a record slaughter year. Intentions can change, however, and this may have something to do with the discrepancy. It is possible that the season has led to more lambs being sold than initially intended.
Contrary to what you would think, the big Merino states of NSW and South Australia did not increase their Merino slaughter intentions, with their share of Merinos relatively steady. Rather, Victoria (figure 2), the state with the largest proportion of meat sheep, was where growers indicated their intention to slaughter more Merinos.
Looking at longer term sheep producer intentions, there has been a general trend over the last four years of producers intending to increase slaughter lamb turnoff. Figure 3 shows the proportion of respondents who are intending to increase, maintain or decrease slaughter lamb turnoff.
In general, 40-45% of respondents intend to increase slaughter lamb turnoff, while only 10-15% intend to decrease turnoff. This would suggest rapidly growing slaughter lamb supply. However, it probably has more to do with the type of producers who respond to the survey, and overestimates the proportion who intend to increase turnoff.
While the data from the MLA-AWI survey may not be totally reliable, it does seem to fit with some of the anecdotal trends we are seeing. The next survey is being conducted now. It will be interesting to see if Merino lamb slaughter intentions fall because of the good recent rain and improved wool prices. These factors may encourage more growers to hold onto Merinos. This could weaken total lamb supply over the coming year. However, the general intention to increase slaughter lamb turnoff should limit falls in lamb supply, with much depending on marking rates. We will have a look at this next week.


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Clove is a new shoe designed specifically for heathcare workers
Designed for healthcare workers, the Clove shoe is easy to slip on and fluid-repellent. It's the winner of the Fashion and Beauty category in the 2020 Innovation by Design Awards.
[Photo: Clove]
When Philadelphia hospitals filled up with COVID-19 cases, front-line workers would wipe themselves down from head to toe with sanitizing wipes between patients, terrified of transmitting the virus within the hospital or bringing it home to their families. Clove, a shoe brand created expressly for healthcare workers, was ready for this moment with sneakers that can withstand industrial-strength cleaners. Before its shoes hit the market in late 2019, the startup had spent a year conducting focus groups with hundreds of medical professionals to create its first product, a shoe that is perfectly tailored to their needs: easy to slip on and fluid-repellent. The brand also got input from shoe designer Stuart Weitzman to ensure a chic profile. When the pandemic arrived in America, many doctors and nurses bought multiple pairs to prepare for the long days ahead, quadrupling Clove's business. "They were preparing for battle, and Clove was part of their armor," says Joe Ammon, Clove's founder and CEO, who was inspired by his wife, a nurse, to launch the business.
See more honorees from the 2020 Innovation by Design Awards here.
WATCH: 7 innovative designs helping us through the pandemic
Elizabeth Segran, Ph.D., is a Senior staff writer at Fast Company. She lives in Cambridge, Massachusetts


Makarova was born in 1940 in Leningrad in former Soviet Russia. At the age of 12, she auditioned for the Leningrad Choreographic School (formerly the Imperial Ballet School), and was accepted although most students join the school at the age of 10.
Makarova was a permanent member of the Kirov Ballet in Leningrad from 1956 to 1970, achieving prima ballerina status during the 1960s. Soon after Makarova defected to the West in 1970, she began performing with the American Ballet Theatre in New York and the Royal Ballet in London.
When she first arrived in the West, Makarova was eager to expand her choreography by dancing ballets by modern choreographers. At the same time, she remained most identified with classical roles such as Odette/Odile in Swan Lake and Giselle. She was featured in the 1976 live American Ballet Theatre production of Swan Lake, simulcast from Lincoln Center on both PBS and NPR.
Makarova continued to excel in many different roles,most notably, her title role in Giselle. She was slim and slight, and combined a delicacy and lyricism with impeccable classical training. The History of Dance, published in 1981, notes that "Her performances set standards of artistry and aristocracy of dance which mark her as the finest ballerina of her generation in the West."
In 1989, Natalia Makarova returned to her home theater of the Kirov Ballet and was reunited with her family and with former colleagues and teachers. Her emotional homecoming was documented in the film Makarova Returns. After her performance at the Kirov, she retired from dancing, donating her shoes and costumes to the Kirov Museum. Today Makarova stages ballets such as Swan Lake, La Bayadère, and Sleeping Beauty for companies across the world.
In the opening week of On Your Toes at the Kennedy Center in Washington D.C., Natalia Makarova was accidentally struck and injured during a performance. With her arm in a cast, she returned to New York, unable to finish the engagement. To recover from this unfortunate accident and prepare for her debut on Broadway, she sought the private and personalized help of her good friends Lynn Stanford and David Howard. For more than two months, David prepared special exercizes to aid in Nataliya's recovery while Lynn created music to help her forget the pain and make her want to dance. "In Private" is the result of this extremely unique situation and contains a wonderful variety of melodies and rhythms for the ballet class. In addition to being an internationally renowned ballerina, Makarova won a Tony Award for her performance in the show On Your Toes.
She appeared as Lydia Lopokova (Lady Keynes) in Wooing in Absence, compiled by Patrick Garland. It was first performed at Charleston Farmhouse and then at the Tate Britain.


Angels hurler Jered Weaver threw MLB's second no-hitter of the season, striking out nine and walking one in a 9-0 win over the Twins. "Why couldn't you be perfect?" screamed Weaver's mother, who was actually Weaver himself wearing a wig and staring in a mirror. "You're nothing! You'll always be nothing!" Man, Jered Weaver is complicated.
Marian Goborik scored in the third overtime to give the Rangers a 2-1 sudden death win over the Capitals and a 2-1 series lead. Hey, anyone else up for calling him "Ranger 'Rik"? You know, from the kid's nature magazine? With the conservation and the birds and- ah FORGET IT YOU GUYS ARE LAME.
Predators goalie Pekka Rinne came up with 32 saves to shut out the Coyotes in a 2-0 win. As the tension builds and the series winds to a conclusion, the cities of Phoenix and Nashville are considering the addition of 10,000 riot police to their downtown areas, and rumors are spreading in Washington that Barack Obama may impose martial law as a precaution.
Chipper Jones hit a walk-off two-run homer to end a wild 15-13 Braves win over the Phillies. "I feel like I'm 40 years old again!" yelled an ecstatic Jones from the ice baths.
Sources report that Stanford Athletic Director Bob Bowlsby will accept an offer to become the next commissioner of the Big 12. In light of this news, let me be the first to congratulate the Big 12 on their impending success in sports like roller blading and rafting, and let me also offer to ghost write the novel about how they used to have good football teams.
Barcelona's Lionel Messi notched a hat trick in a win over Malaga to set the all-time European club scoring record with 68 goals on the season. The previous record-holder, German striker Gerd Mueller, still holds the record for most goals in a season without smiling.
I'm putting the call out for Reader's Revenge, the feature where we take stories from you, the reader, and feature the best ones in a top ten list. This week's topic is Your Worst Moment with the In-Laws/Significant Other's Family. Click the link to see the last installment and get more info if you have no idea what I'm going on about, and otherwise send your best stories to tobaccordblues@gmail.com by Sunday. As of today, this topic is wide open.
What's Going On With Matt Kemp?


Karen Taylor Fine Art returns to Illustrationcupboard Gallery for a second summer show, bringing a selection of British 18th and 19th century paintings and drawings this month.
Among the highlights is a fresh group of watercolours by Louis Francia (1772- 1839), a French artist specialising in shore landscapes who spent much of his earlier life in the UK, and others by the Norwich School painter John Sell Cotman (1782-1842).
The works have not been seen in public since they were painted in the 1830s. They were collected together into a friendship album compiled by Baron Francois Akerman (1809-90), regent of the Banque de France, for his wife Louise Bouquet de Saint Simon after their marriage in 1836.
Since then, the pictures have been kept out of the light in a thick leather binding at the family home, the Château de Coulonges in the Loire.
Such albums were fashionable in the 19th century, and guests would bring a drawing or watercolour to commemorate a visit. The book in question included a group of unmounted drawings and watercolours by artists including Richard Parkes Bonington, William Wyld and Alexandre-Gabriel Decamps.
Karen Taylor specialises in British art from all periods and normally operates by appointment. However, the annual show, which runs from June 29-July 6, is a chance to display a number of pictures together in one space.
Other highlights include a rare gouache conversation piece on vellum attributed to Thomas Bardwell, a portrait by Sir Joshua Reynolds and a group of topographical paintings and drawings from Greece and Turkey.


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Le toueur est un type de bateau amphidrome se propulsant par touage d'une chaîne ou d'un câble reposant sur le fond.

Historique
Le toueur apparaît au , avec l'introduction progressive du machinisme dans les procédés de propulsion. Il va jouer un rôle majeur dans la traction des bateaux en navigation intérieure sur les fleuves, les rivières et aussi les canaux. Rarement porteur, le toueur va surtout être utilisé comme remorqueur de bateaux porteurs non motorisés, tels que des chalands ou des péniches.

Principe
La vapeur créée actionne un moteur à piston qui actionne lui-même un engrenage autour duquel s'enroule une chaîne immergée au fond du fleuve. Selon le principe du touage, l'engin se hale lui-même grâce à un treuil embarqué motorisé et à l'aide d'une chaîne (parfois un câble) immergée et fixée solidement au sol aux deux extrémités du parcours sur lequel il travaille. Un toueur peut emmener ainsi une trentaine de bateaux chargés à 250 tonnes chacun.

L'utilisation de moteurs électriques sur les toueurs va les rendre aptes au tractage dans les tunnels-canaux de grande longueur non ventilés à l'intérieur desquels les péniches motorisées ne pouvaient pas utiliser leur moteur. Ils seront une alternative aux locotracteurs circulant sur une voie métrique du chemin de halage, comme à Liverdun.

Patrimoine

En France il existe encore quelques toueurs en fonction. Ils tirent les péniches sur le canal de Saint-Quentin et sur le canal de la Marne au Rhin. Celui qui faisait traverser les  mètres du Tunnel de Mauvages, dans la Meuse (entre les communes de Mauvages et de Demange-aux-Eaux), a été désaffecté en 2013, et celui qui est encore en fonction dans la tunnel de Riqueval (5670 m), sera arrêté en 2023.

On peut voir des toueurs exposés à terre à Riqueval, à Saint-Léger-des-Vignes (Nièvre) ainsi qu'à Pouilly-en-Auxois (Côte d'Or).

Sur le Rhône, le toueur Ardèche, en rade à l'état d'épave dans le port de plaisance de l'Épervière à Valence, a été construit en 1896. À l'origine, il pesait 325 tonnes. Contrairement aux autres toueurs, tous partis à la casse, il a échappé aux ferrailleurs grâce à Pierre Bonnet, aujourd'hui décédé, alors maire de La Coucourde-Derbières, qui l'avait racheté dans les années 1970. Mais des menaces planent sur ce dernier représentant d'un patrimoine exceptionnel.

Notes et références

Voir aussi

Articles connexes
 Touage
 Chaland de Loire
 Fûtreau
 Gabare

 Ramberte
 Loire
 Scute

Liens externes
 Dictionnaire des bateaux fluviaux dans le projet Babel : le toueur
 Le tunnel de Mauvage

Type de bateaux
Type de bateau fluvial
Type de bateau de passagers
Type de bateau de transport
Câble

redpajama_set_name: RedPajamaWikipedia


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buy it for the architecture Nov. 3, 2020
A Brooklyn Heights Curiosity Is on the Market for the First Time
By Megan Barber
Photo: DDREPS, Courtesy of The Corcoran Group
On a leafy, one-block-long Brooklyn Heights street lined with well-preserved Greek Revival townhouses, a rare bit of mid-century modern history has gone up for sale. Close to — though still set back from — the oft-crowded Brooklyn Heights Promenade, 48 Willow Place is one of three mid-century townhouses designed by local architects Joseph and Mary Merz.
Location: Brooklyn Heights, Brooklyn
Architect: Joseph and Mary Merz
Specs: 6 beds, 7.5 baths, 5,000 square feet, plus an 1,480-square-foot garden
Listed by: Deborah Rieders, Sarah Shuken, and Raquel Lomonico / Corcoran
In the early 1960s, the couple purchased three vacant non-contiguous lots at auction for $11,000, a particularly low price since Robert Moses was considering adding a Brooklyn Queens Expressway exit ramp right next door on State Street (the ramp ended up a block away on Atlantic Avenue). The Merzs then began planning how to design the three homes with a distinctly modern aesthetic, but one that wouldn't clash with the group of four 1840s townhouses fronted by chunky white columns across the street. Eventually they landed on using redwood and custom-designed concrete blocks marked to resemble eight-inch squares. From the street, the cement facade is as imposing as the columns nearby, but the Merzs' use of plain cement cornice echoes the neoclassical neighbors.
The resulting townhouses earned a spot in Architectural Record's list of record houses of 1969, where the design was described as "sophisticated," "uncluttered," and "orderly." The Merzs themselves moved into No. 48, a 5,000-square-foot home on a corner lot wrapping around State Street (No. 40 is even larger at 6,581 square feet and last sold in 2013 for $7.6 million; No. 44 is much smaller and has been divided into two duplexes). The project contributed to the creation of the Brooklyn Heights Historic District (the city's first designated historic district) in 1965 and also led to many commissions for the couple, including a redesign of the headquarters for the Dreyfus Fund in the GM Building.
Over the years, the Merz residence has stayed in the family, and the interior features remain untouched (all the furniture currently shown is staged). The abundant, large windows are still framed in redwood, while custom yellow-hued maple millwork showcases built-in bookshelves and cupboards in the 34-foot-wide living room and kitchen. Common spaces feature Scottish terracotta stone flooring that, per listing agent Deb Rieders, shows almost no sign of wear after decades. And the kitchen retains paneled appliances and open shelving (unusual elements in the 1960s) on a single back wall to maintain an open floor plan.
Upstairs, there are currently six bedrooms and four baths, but the building design allows for flexible layout configurations. Unlike in most brownstones, the only structural walls in this townhouse are the four exterior ones; the rest are fixed but "floating," meaning that you could create any number of larger or smaller rooms by adding interior walls. There's also a windowed basement that had served as the headquarters of Merz Architecture, with two baths, three office rooms, and plenty of storage.
For all of the mid-century appeal inside, the house's outdoor spaces are also quite special. The Merzs believed in incorporating nature into design, and their home includes a Japanese garden with a stone path leading to a wooden deck with a pergola (and a carport underneath), which all take up the full width of the lot. And the couple worked on outdoor spaces outside of their own home, as well — according to the Brooklyn Daily Eagle's obituary for Joseph, the Merzs planted several hundred neighborhood trees, renovated a community center on Willow Place, designed a playground and garden at Columbia and State streets, and were named joint curators of Prospect Park in 1975.
buy it for the architecture
Brooklyn Mid-Century Townhouse Hits Market for First Time


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Pandemic shows science is only way to save the world, says Greta Thunberg
01 01 2021 | 11:10Bethany Dawso
We are not listening to climate scientists, we're not listening to scientists who work on biodiversity and that, of course, needs to change'
Although the coronavirus pandemic has highlighted the importance of following scientific advice, Greta Thunberg states climate experts continue to be ignored on the severity of the climate crisis.
The Swedish activist stated that the Covid-19 crisis had "shone a light" on how "we cannot make it without science", but people were "only listening to one type of scientists".
Her comments came in an alongside author Margaret Atwood, broadcast on BBC Radio 4 on Tuesday as part of the two-times Booker Prize-winning writer's guest-editing of the Today programme.
Thunberg was asked if people's new-found appreciation of science due to the pandemic would have an effect on climate information as well.
"It could definitely have. I think this pandemic has shone a light on how ... we are depending on science and that we cannot make it without science," the 17-year-old replied.
"But of course, we are only listening to one type of scientist or some types of scientists, and, for example, we are not listening to climate scientists, we're not listening to scientists who work on biodiversity and that, of course, needs to change."
The environmental campaigner expressed a lack of enthusiasm and trust when questioned about nations' pledges to reduce their carbon emissions, such as China which has committed to reaching a net-zero target by 2060.
"That would be very nice if they actually meant something," Thunberg said.
"We can't just keep talking about future, hypothetical, vague, distant dates and pledges. We need to do things now. And also net-zero ... that is a very big loophole, you can fit a lot in that word net."
Thunberg said the election of Joe Biden as US president, who has pledged to rejoin the Paris climate accord on the first day of his presidency, sent a "signal" that "it could be a good start of something new".
"Let's hope that it is like that, and let's push for it to become like that," she said.
The coronavirus pandemic has prevented the Fridays for Future movement that Thunberg inspired from holding its mass rallies in recent months.
Thunberg started her solo protests outside Sweden's parliament in Stockholm in August 2018.
She told the Today programme that climate striking had moved online.
Asked by Atwood about what she would do if given a magic wand and three wishes to use, Thunberg said: "I don't think anyone should have that amount of power."
She added: "Maybe I would just go for something easy, like, make everything become alright, and make sure that everything turns out the way it should, or make the world a better place or something like that."
Bethany Dawson@bethanymrd
Greta Thunberg: 'We are speeding in the wrong direction' on climate crisis.
'Hypocrites and greenwash': Greta Thunberg blasts leaders over climate crisis.
Greta Thunberg: don't dump climate crisis on children to fix.


Trained volunteers are key to gathering the core of our research data. Each spring we recruit new volunteers. A one-evening training session and an in-the-field orientation are required for volunteers. Volunteers are expected to visit their assigned colonies at least once every week during late May, June, July and August.
The survey also includes more concentrated study during the period when the young are being fed in the burrows. Focused research at this time will shed light on the success of fledglings and the species of fish used to feed the young.
Our volunteer team has great enthusiasm for and interest in stewarding their colonies. The survey experience offers them an opportunity to contribute in a meaningful way to a community science project. Many also find it a delightful excuse for observing and enjoying our nearshore environment.
Please visit the "Breeding Survey Resources and Data" tab to view more details about the volunteer duties and protocol.
1. Document the species of fish being delivered to chicks in burrows.
2. Determine the frequency of fish delivery and the burrow to which it is delivered.
3. Monitor and map the burrows in chosen colonies around Whidbey Island.
2. Arrive at the colony within one hour after sunrise.
3. Make observations for 4 hours.
4. Count the total number of adults every 30 minutes.
5. Note the time when you see a bird with a fish.
6. Identify and record the species of fish type: gunnel, sculpin, perch, cod, or other.
7. Record the burrow receiving the fish delivery and the time.
8. Designate the burrow by a number. Use the same numbering system as the volunteer for that site.
2. Submit data to the coordinating team every 2 weeks.
3. Write up report at the end of the survey to be submitted to the Island County MRC, and Whidbey Audubon Society.
4. Power Point presentation to the funders.


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«June 12 |June 14 »
June 13 down the years
'How does it feel to drop the World Cup?'
South Africa's costliest dropped catch
Herschelle Gibbs' century was overshadowed by the costly drop of Steve Waugh	© Getty Images
"How does it feel to drop the World Cup, Herschelle?" That's what Steve Waugh supposedly said to Herschelle Gibbs on this day, when Gibbs, in prematurely celebrating catching Waugh at midwicket, threw away a chance to put Australia out of the tournament, at Headingley. Waugh had 56 at the time - he'd come to the crease with Australia on the brink at 48 for 3, chasing 272 - and went on to an awesome 120 not out. Not only did it keep Australia in the Cup, it put them above South Africa in the Super Six stage, an incidental detail at the time but one that would be of monstrous significance when the two sides tied in the semi-final four days later.
In Sarodar, India, one of England's most stylish batsmen is born. Kumar Shri Duleepsinhji - Duleep to most people - was the nephew of the great Ranjitsinhji, and had all of his wristy class and grace. His finest innings was a sensational 173 against Australia at Lord's in 1930. It was Duleep's first innings in an Ashes Test, but he ended up on the losing side after Don Bradman piled up 254. He only played 12 Tests because of poor health, but ended with an outstanding average of 58. Duleep also cracked 333 in a day for Sussex, against Northamptonshire at Hove in 1930. He later became High Commissioner for India in Australia. He died of a heart attack in Bombay in 1959.
Amelia Kerr, just 17, broke the record for the highest individual score in women's ODIs when she made 232 not out against Ireland in Dublin. Her innings, which went past Belinda Clark's 229 -- set way back in 1997 -- came in 145 balls, and it resulted in a 305-run thrashing for hapless Ireland, who were also at the receiving end when New Zealand racked up the highest total in all ODI cricket, 490, against them earlier in the series.
Raman Subba Row saved England with a three-day vigil after they had conceded a first-innings lead of 321 at Edgbaston. After a 171-minute half-century in the first innings - England were bowled out for 195, following which a hundred from Neil Harvey and a half-century from debutant Bill Lawry put Australia in a seemingly match-winning situation - Row batted between days three and five for a four-hour 112. But there was no danger of a collapse after his dismissal, even though England were still trailing, because Ted Dexter took up the task. His 180 took nearly six hours and saved the first Ashes Test in Edgbaston in 52 years.
In the days when cricket was not able to gorge on money from TV, a long-running dispute that threatened a blackout of English cricket on TV was resolved. The BBC upped their offer for rights to the 1975 Ashes and the 1976 West Indies series, as well as the 1975 World Cup and all English domestic one-day competitions for two summers, to £270,000.
Birth of Chris Cairns, the star of a largely faceless New Zealand team. With his clean, muscular hitting, athletic fielding and urgent new-ball bowling, Cairns was for a time arguably the only true allrounder in world cricket, until a string of injuries curtailed his bowling effectiveness. He became only the sixth man to achieve an allrounder's double of 200 wickets and 3000 runs in March 2004, and later that year broke the record for most sixes in Test cricket. He retired from international cricket in 2006, and played in the Indian Cricket League in 2008. In March 2012 he won a libel suit against the former IPL chairman Lalit Modi over match-fixing allegations that related to his stint in the ICL. Two years later he was charged with perjury but was acquitted after a nine-week trial in London in 2015.
An amazing performance from Alec Bedser for England against Australia in the first Test, at Trent Bridge - and an ultimately fruitless one. No bowler has returned better match figures than Bedser's 14 for 99 in a Test that his side has not won. Here he took 7 for 55 in the first innings, and England still trailed by 105. Bedser then blew the Aussies away a second time with 7 for 44, but with England seemingly on course for victory, rain washed out all of the fourth day and most of the fifth.
A historic victory for England at Old Trafford - their first in the opening Test of a home Ashes series for 42 years. It was a game they bossed from the moment John Snow and Geoff Arnold ripped out the Aussies for 142 in their first innings. England's debutant, one Tony Greig, had a match to remember. He top-scored in each innings, with 57 and 62, and took 4 for 53 in the second innings. Greig had already had some experience of the big time, though - he played in the unofficial series against Rest of the World in 1970.
A less auspicious start to an Ashes series for England. They only had to bat out 83 overs to save the first Test on a good Headingley pitch, but they were cleaned up inside two sessions as Terry Alderman, aided and abetted by a rapacious slip cordon, took his second five-for of the match. Amazingly, England started this series as favourites (they went on to lose 0-4; were it not for rain it would have been 0-6), but they blundered from the start. David Gower left out his spinner, John Emburey, and put Australia in on a belter. Two days later they were 601 for 7 - Steve Waugh made 177 of them without looking like getting out - and England never recovered.
A prodigy is born. It's amazing to think that Maninder Singh's last Test appearance came when he was 27. He was only 17 years 193 days old when he made his Test debut, against Pakistan in Karachi in 1982-83, and was India's youngest cricketer at the time. Maninder was a subtle slow left-armer who seemed a natural heir to Bishan Bedi, even down to the fact that he bowled in a patka. But at the top level he was a bit of a fair-weather performer. Against the heavyweights (Australia, West Indies and Pakistan), he took 45 wickets in 26 Tests at an average of 55. Against the rest (England, Sri Lanka and Zimbabwe) he took 43 wickets in only nine Tests at an average of 19. He was also the last man out in the tied Test against Australia in Madras earlier that winter.
A Jekyll-and-Hyde cricketer is born. Nottinghamshire's George Gunn mixed dashing strokeplay and dour defence with bewildering frequency, and as he walked to the crease, nobody quite knew which mood would take him. His zenith was his debut, when he made 119 and 74 against Australia in Sydney in 1907-08. But England lost that, as they did when Gunn made his other Test century, on the same ground that same winter. He hadn't played cricket for 17 years when he returned in the West Indies in 1929-30. Only John Tracios of South Africa and Zimbabwe has had a longer interval between Test appearances. Gunn's brother John and uncle William also played Test cricket. Gunn, who ended with a Test career of exactly 40, died in Sussex in 1958.
Other birthdays
1887 Neville Tufnell (England)
1888 Roy Minnett (Australia)
1931 Esme Irwin (England)
1967 Angus Mackay (Zimbabwe)
1970 Shaun Young (Australia)
1972 Alex Tait (New Zealand)
1976 Ian Redpath (Australia)


Engineering of single-species biofilms for enzymatic generation of fine chemicals is attractive. We have recently demonstrated the utility of an engineered Escherichia coli biofilm as a platform for synthesis of 5-halotryptophan. E. coli PHL644, expressing a recombinant tryptophan synthase, was employed to generate a biofilm. Its rapid deposition, and instigation of biofilm formation, was enforced by employing a spin-down method. The biofilm presents a large three-dimensional surface area, excellent for biocatalysis. The catalytic longevity of the engineered biofilm is striking, and we had postulated that this was likely to largely result from protection conferred to recombinant enzymes by biofilm's extracellular matrix. SILAC (stable isotopic labelled amino acids in cell cultures), and in particular dynamic SILAC, in which pulses of different isotopically labelled amino acids are administered to cells over a time course, has been used to follow the fate of proteins. To explore within our spin coated biofilm, whether the recombinant enzyme's longevity might be in part due to its regeneration, we introduced pulses of isotopically labelled lysine and phenylalanine into medium overlaying the biofilm and followed their incorporation over the course of biofilm development.
Through SILAC analysis, we reveal that constant and complete regeneration of recombinant enzymes occurs within spin coated biofilms. The striking catalytic longevity within the biofilm results from more than just simple protection of active enzyme by the biofilm and its associated extracellular matrix. The replenishment of recombinant enzyme is likely to contribute significantly to the catalytic longevity observed for the engineered biofilm system.
Here we provide the first evidence of a recombinant enzyme's regeneration in an engineered biofilm. The recombinant enzyme was constantly replenished over time as evidenced by dynamic SILAC, which suggests that the engineered E. coli biofilms are highly metabolically active, having a not inconsiderable energetic demand. The constant renewal of recombinant enzyme highlights the attractive possibility of utilising this biofilm system as a dynamic platform into which enzymes of interest can be introduced in a "plug-and-play" fashion and potentially be controlled through promoter switching for production of a series of desired fine chemicals.
The development of novel biocatalysts is crucial in order to provide greener and more sustainable solutions for synthesis of fine chemicals. Enzymatic transformations are becoming increasingly important to the chemical manufacturing and pharmaceutical industries and it is projected that by 2050, in order to move toward greater sustainability and cost reduction, 30 % of chemical syntheses in these sectors will be biocatalytic [1, 2]. One significant challenge to reaching this goal is that many enzymes demonstrate poor stability. Whilst immobilization technologies have been successfully utilised to improve enzyme viability and allow catalyst recycling [3–13], and evolution approaches have been successfully employed to afford enhanced stability [14–25], these approaches must be applied and optimised for each enzyme on a case-by-case basis and can add significant overall cost to the biocatalytic process. Whole cell biocatalysts offer a useful alternative to the use of isolated enzymes, especially for processes requiring the recycling of expensive cofactors [26–37], yet these biocatalysts are not able to operate for prolonged periods of time due to substrate and/or product toxicity and constant exposure to physical and chemical extremes, i.e. shear stress, high temperature, pH variation or solvents. This has driven the need to optimize a given whole-cell process, such as immobilization of cells which provide a more stable environment and the benefit of easy retrieval for reuse [29, 35, 38–45]. In contrast, the naturally immobilized cells or biofilms representing the predominant microbial lifestyle in the natural environment are well known for their resilience [46–51]. Their robustness, which in part is conferred by a well-organized 3-dimensional architecture where cells are embedded and protected by a matrix of secreted extracellular polymeric substances (EPS), allows them to withstand unfavourable conditions that planktonic cells and immobilised enzymes are not able to tolerate [52–56]. These properties lend them to being stable biocatalysts of utility to various industrial sectors: biofilms, mainly multi-species communities of bacteria, are employed in waste water treatment and bioremediation of polluted sites [57–61], and single species biofilms are used for the sustainable production of simple compounds, such as acetic acid, ethanol, butane-2,3-diol and succinic acid [60, 62, 63]. Recently, there is a growing interest in developing and utilising biofilm-based processes, and a new area of utilising engineered single species biofilms for synthesis of desired fine chemicals is beginning to emerge [64–70]. The utilisation of E. coli biofilms is attractive due to the wide industrial use of E. coli and the ease with which it can be genetically manipulated, e.g., overexpressing genes encoding target enzymes and constructing knockout mutants for control and maximization of target products and elimination of by-products. Previously we have demonstrated how the process of biofilm generation and maturation by E. coli K-12 PHL644, which readily forms biofilms due to overproduction of the adhesin curli , could be artificially accelerated by simply spinning cells onto glass plates coated with poly-l-lysine . After 6 days maturation of the engineered biofilm, well-organised mushroom-like structures are visible where cells are arranged in multilayers and embedded in the biofilm extracellular matrix. Deep pores and channels developed, conferring a large catalytic surface area to the biofilm [65, 72]. We previously showed that this system could be engineered by constitutive expression of the tryptophan synthase from Salmonella enterica (TrpBA) encoded by plasmid pSTB7 . The recombinant TrpBA is composed of two subunits associated as an α2β2 tetramer, and the β subunit is catalytically independent, mediating the synthesis of l-tryptophan from free indole and l-serine. The engineered biofilm has been employed to catalyse the enantio-selective conversion of haloindoles and l-serine to l-halotryptophans, a series of fine chemicals of value in the synthesis of medicinally relevant compounds [65, 70]. We previously observed that in the biotransformation of 5-chloroindole to 5-chlorotryptophan, strikingly, higher conversions were obtained with the 7 day-old biofilm (conversions of 78 %) than with the immobilised TrpBA enzyme (conversions of 49 %) or planktonic cells expressing recombinant TrpBA (conversions of 40 %) . It was also notable, in our previous experiments, that biotransformation of 5-chloroindole to 5-chlorotryptophan with the engineered biofilm still proceeded at the initial rate following 30 h incubation (Additional file 1: Figure S1a) . Moreover, the biofilm could be reused and, remarkably, there was no observable decrease in catalytic activity after three sequential reaction cycles (Additional file 1: Figure S1b) . Intrigued by the promising catalytic longevity and the potential application of this biofilm system in catalysing a range of required biotransformations, as simple to engineer, plug and play biocatalysts, we sought to investigate the reason for the observed biocatalytic longevity further. Through a pulse-chase SILAC (stable isotope labelling by amino acids in cell cultures) study, we set out to explore the fate of the recombinant enzyme introduced to the biofilm as a catalyst. Our aim was to investigate whether the catalytic longevity of the biofilm is in part due to the regeneration of the recombinant biocatalytic enzyme, beyond the protection conferred by the EPS.
Pulse-chase SILAC experiments involve dynamically following the metabolic behaviour of a cell culture in the presence of an isotopically labelled amino acid, followed by media replacement and the growth of the culture in the presence of the unlabelled amino acid. In this case we wished to explore whether the production of the recombinant enzyme tryptophan synthase was continuous, in engineered biofilms. In a typical SILAC experiment, at given time points, cells are harvested for preparation of protein samples and target proteins are isolated and tracked for label incorporation. Lysine and arginine are the most commonly used labelled amino acids for these studies as they present a trypsin cleavage site (provided they are not followed by a proline residue) and a convenient C terminal label for mass spectrometry (MS) analysis [74, 75]. Other stable isotope labelled amino acids are also frequently used  and the choice is usually dictated by the abundance of each amino acid in the protein under investigation. Importantly, the labelled form of the peptides resulting from the trypsin digestion of the parent protein must have a distinct mass, compared to the unlabelled peptide pool in order to be detected in the MS analysis. The optimal additional mass conferred by an isotopically labelled amino acid is plus 4 Da, when exploring expression profiles of a complex series of different proteins or an entire proteome . However, when exploring a single protein, the additional mass of plus 2 Da per amino acid incorporated is more than sufficient to enable ready detection of incorporation, as, for a doubly charged peptide fragment, the incorporation of such a label would result in an increase of 1 Da in the m/z. Conventional SILAC analysis involves a single pulse of one isotopically labelled amino acid followed by a single chase [76–78]. However, to gain a greater understanding of our biofilm and whether it regenerates its recombinant biocatalytic enzyme over the course of its maturation, we designed a multistep labelling regiment using a series of two labelled and unlabelled amino acids. To ascertain which two labelled amino acids should be utilised in the SILAC experiment, the amino acid composition and the tryptic digest of the β subunit of TrpBA were evaluated in silico (summarised in Additional file 1: Tables S1, S2, respectively). In TrpBA, lysine and phenylalanine are abundant, at around 5 and 3 % of all amino acids. Of the unique tryptic peptides with five or more amino acids, 71 % contain lysine and 33 % contain phenylalanine. Due to the observation of a high frequency of occurrence of lysine in the tryptic peptides of TrpBA, we therefore chose to use [4,4,5,5-2H4]-l-lysine as the SILAC amino acid. We chose [2,6-2H2]-l-phenylalanine as the second labelled amino acid due to our ability to generate the isotopic phenylalanine in plentiful supply, in a cost effective manner, in house. Growing the biofilm in the presence of each of these labels would provide an anticipated shift of 2 m/z unit per each [4,4,5,5-2H4]-l-lysine residue incorporated and a shift of 1 m/z unit per each [2,6-2H2]-l-phenylalanine residue.
To provide statistically robust results, in order to explore definitively whether the recombinant enzyme in the biofilm was regenerated, six series of pulse-chase experiments were carried out, in triplicate, as summarised in Fig. 1 (full details provided in the Methods section). Our labelling regimen involved supplementation of sets of individual cultures of the E. coli cells, to be used in biofilm formation, each with one of the two isotopically labelled amino acids. After the cultures had grown in the presence of the label, these cells were spin-coated onto glass slides that were then placed within deep-well plates, in the same manner as we have previously reported for the promotion of rapid biofilm formation . Fresh media containing the same labelled amino acid (either labelled lysine or phenylalanine), as the cultures had been initiated in, was reintroduced to each well, and then incubated for 3 days whilst the biofilms started to mature. After 3 days the medium was removed and replaced with fresh media supplemented now with the unlabelled variant of the initial amino acid that had been used for each culture/biofilm i.e. unlabelled phenylalanine (Fig. 1, rows 2, 3) or unlabelled lysine (Fig. 1, rows 5, 6). The biofilms from both rows 1 and 4 were harvested for preparation of target proteins subjected to tryptic digestion followed by liquid chromatography-mass spectrometry (LC–MS) analysis (as detailed in the ''Methods'' section). After a further 3 days of maturation, biofilms in rows 2 and 5 were harvested and processed for downstream analysis. Again, the media were removed from all of the other biofilms, and the isotopic labelling regimen was swapped with the biofilm in row 3, that had previously been cultured in the presence firstly of deuterated then unlabelled phenylalanine now being supplemented with labelled lysine. Conversely, the biofilm in row 6 that had previously being supplemented firstly with labelled then unlabelled lysine, now being supplemented with labelled phenylalanine. The incubation continued until day 9 with biofilms in rows 3 and 6 being harvested at this stage.
As described above, and in Fig. 1, we designed a labelling regime which included pulsing with labelled amino acids, followed by a chase phase in which unlabelled amino acids, then an alternative labelled amino acid was administered to maturing biofilms. At each time point, where the label was changed (as illustrated in Fig. 1) biofilms were harvested for preparation of target proteins subjected to tryptic digestion, and the tryptic peptides were submitted for LC–MS analyses. The MS data were analysed using the Mascot algorithm (Matrix Science, London, UK)  with the modified masses of SILAC amino acids, i.e., [2,6-2H2]-l-phenylalanine and [4,4,5,5-2H4]-l-lysine, added as variable modifications in the searches against an internal database at the BSRC (Biological Science Research Complex, University of St Andrews, UK). The data were also searched against the National Centre for Biotechnology Information (NCBI) database . The fragments of the β subunit of TrpBA identified by Mascot, based on at least five biofilm samples, are listed in Additional file 1: Table S3. In total, the identified fragments covered 53 % of all amino acids of TrpBA.
The aim of pulse-chase SILAC experiment performed in this study is to evaluate whether the recombinant enzyme expressed in the biofilm is replenished over time; if the system is static and the catalytic longevity is simply due to the protection of the initial enzyme population, little change or no incorporation in the label would be expected for target proteins coming from biofilms after 6 days of maturation. If the engineered biofilm enables catalytic longevity by renewal of the enzyme and the system is highly dynamic we would anticipate an observation of a change in the incorporation level of the respective labelled amino acid, at 6 days of maturation and beyond. After peptide identification, the MS spectra and ion chromatograms of two tryptic peptides (VGIYFGMK and DPEFQAQFADLLK) from the β subunit of TrpBA were extracted and examined at the MS level to quantify the incorporation level of each SILAC amino acid (the distribution of two peptides along the protein sequence is depicted in Additional file 1: Figure S2). The signal intensities (areas under the curves) from light and heavy peptides provide a quantitative comparison of their relative abundances [75, 78, 81]. The MS spectra of two reference peptides, which each gave nearly identical quantification results, are provided in Additional file 1: Figures S3, S4.
We examined the incorporation level of SILAC amino acid into target proteins from biofilm samples harvested at each time point (day 3, day 6 and day 9). The level of incorporation was quantified by calculating the median of the labelled (heavy, H)/unlabelled (light, L) ratios according the accepted procedure . In accordance with this standard literature procedure for determination of the median of H/L peptide ratios, the peak areas of the extracted ion chromatograms (XICs) of the labelled peptides were measured against the unlabelled peptides. The degree of labelling on target protein was quantified with two reference peptides, as listed above (see Additional file 1: Figures S5, S6 for the XICs of the two peptides). Labelling analysis of the recombinant tryptophan synthase produced by 3 day-old biofilms showed isotopic incorporation levels of ~93 % for biofilms that had been cultured in minimal M63 medium supplemented with [2,6-2H2]-l-phenylalanine (Fig. 1, row 1 samples), and isotopic incorporation levels of about 38 % for biofilms that had matured in [4,4,5,5-2H4]-l-lysine supplemented medium (Fig. 1, row 4 samples). The lower incorporation level of deuterated l-lysine was observed, even though supplemented at ten-fold higher concentration, potentially due to the direction of this amino acid towards alternative metabolic pathways, and highlights the benefits of utilising a pair of labelled amino acids.
To evaluate whether fresh enzymes were produced in both sets of samples, as described above, after 3 days maturation of the engineered biofilm we started the chasing by replacing the labelled medium with fresh medium supplemented with the respective unlabelled amino acids (Fig. 1, rows 2, 3 for phenylalanine and rows 5, 6 for lysine). After 3 more days of maturation, biofilms from rows 2 and 5 chased with unlabelled phenylalanine and lysine respectively, were harvested for preparation of target proteins. As seen in Fig. 2, for the row 2 samples, it was observed that the incorporation level of deuterated l-phenylalanine had dropped from 93 to 10 %; for the row 5 samples, the incorporation level of [4,4,5,5-2H4]-l-lysine was also observed to decrease from 38 to 16 %. The reduced level of SILAC amino acids observed for three reference peptides suggests that new recombinant enzyme was dynamically generated de novo. We also examined the MS spectra of two reference peptides, detected from the tryptic digest of recombinant enzyme coming from the biofilms chased with unlabelled phenylalanine (Additional file 1: Figures S3, S4; row 2) or unlabelled lysine (Additional file 1: Figures S3, S4; row 5). A drop in the incorporation level of the stable isotope labelled amino acids was observed as indicated by the ratios in the MS spectra of the correlated heavy and light peptides, which is consistent with the quantification analysis based on XICs.
To extend our studies on the enzyme regeneration we sought to explore whether a further change in labelling regiment, past day 6, would continue to differentially label the tryptic peptides. At day 6, biofilm that had previously been administered l-phenylalanine was now pulsed with 300 μM of [4,4,5,5-2H4]-l-lysine (Fig. 1, row 3) and conversely the biofilm that had been administered l-lysine was now pulsed with [2,6-2H2]-l-phenylalanine (Fig. 1, rows 6). After a further 3 days of maturation, the biofilms were harvested and processed for isolation of target proteins. Incorporation of labelled lysine into the recombinant tryptophan synthase was observed for the biofilms initially matured in medium containing phenylalanine and vice versa (see Additional file 1: Figures S3, S4 for incorporation of [4,4,5,5-2H4]-l-lysine and [2,6-2H2]-l-phenylalanine). Strikingly, incorporation of labelled phenylalanine is observed at ~90 % for biofilm samples in row 6, close to the incorporation level observed for the 3 day-old biofilms in row 1. Conversely, and in agreement with this result, the 9 day-old biofilms in row 3 that had been switched to the labelled lysine pulse now showed an incorporation level of labelled lysine at ~43 %, very similar to the incorporation level of ~38 % observed for the 3 day-old biofilms in row 4. This consistency in labelling level reveals that the recombinant enzyme within the biofilm continues to be completely replenished without perceivable difference in regeneration rates between the 3 day-old and 9 day-old biofilms. For our assessment of the ability of the biofilm samples to catalyse biotransformation, slides containing biofilms that had matured for 7 and 9 days were transferred to the reaction buffer supplemented with 5-chloroindole and incubated at 28 °C for 24 h in an orbital shaker incubator, as detailed in the Methods section. No significant difference was observed in terms of overall percentage chlorotryptophan yield between biotransformations with 7 and 9 days-old biofilms (54 vs 59 %; Additional file 1: Figures S7, S8), confirming that the generated recombinant tryptophan synthases in biofilms are catalytically active. Overall, these findings demonstrate that a key contributor to the striking and useful longevity of the biocatalytic biofilm is the regeneration of the active enzyme.
The urgent demand for the green, sustainable and cost-effective generation of fine chemicals continues to drive the need for new and effective tools for biotransformations. It is predicted that in the near future almost one-third of industrial chemical synthesis will be mediated by enzymes. To embrace this challenge, not only must new enzymes be discovered and developed, but robust and generic approaches toward the immobilisation, stabilisation and protection of the biocatalyst must be established. Readily engineered, single species biofilms represent an exciting opportunity as biocatalytic platforms into which genes encoding enzymes that catalyse valuable biotransformations can be introduced. The biofilm thereby produces, immobilises and protects the biocatalyst. We envisage the possibility of utilising this biofilm as a cell factory in which enzymes of interest can be introduced into this platform in a "plug and play" fashion for the generation of desired fine chemicals. We had previously observed the biofilm possesses striking biocatalytic longevity, this, we had postulated, was likely to be predominantly due to the protection by the EPS. Our studies of the lifespan and regeneration of the recombinant enzyme, using SILAC, demonstrate that the recombinant enzyme is being constantly regenerated in the matured biofilm, which we believe is fundamental to the longevity of its catalytic activity. Remarkably, we see similar levels of fresh label incorporation in a 9 day-old biofilm as in a 3 day-old biofilm indicating a similar generation level of the recombinant enzyme in the developed biofilm as in the newly formed biofilm. This result is surprising and exciting as it opens the potential, through the careful choice and application of promoters, to switch the biocatalytic function of a mature biofilm, and the products that it generates.
Chemicals used in this study were purchased from Sigma-Aldrich (St Louis, MO, USA), Fisher Scientific (Leicestershire, UK) and Apollo Scientific (Stockport, UK), unless stated otherwise. Solvents were purchased from Sigma-Aldrich and Fisher Scientific. E. coli K-12 strain PHL644 (MC4100 malA-kan ompR234)  transformed with pSTB7 plasmid containing S. enterica trpA and trpB genes and encoding ampicillin resistance  is employed in this study.
General microbiological work was performed in a sterile environment using either a laminar flow hood or working around a bunsen flame. A 2 % Distel solution was used to sterilise surfaces prior to work. Strains were routinely grown in LB (Luria–Bertani) broth (10 g L−1 NaCl, 10 g L−1 tryptone, 5 g L−1 yeast extract, pH 7.0) or agar plates supplemented with ampicillin at a final concentration of 100 μg mL−1. For solid medium, agar was added at a final concentration of 15 g L−1. For long term storage at −80 °C, LB broth containing 20 % of glycerol was used.
For biofilm maturation, M63 medium (100 mM KH2PO4, 15 mM (NH4)2SO4, 0.8 mM MgSO4.7H2O, 9 mM FeSO4.2H2O, 17 mM K-succinate, 10 mM glucose, adjusted to pH 7.0 and supplemented with 100 μg mL−1 ampicillin) was used as it has been shown to promote curli production. For SILAC experiment, M63 medium was supplemented with either [4,4,5,5-2H4]-l-lysine (Sigma-Aldrich, St Louis, MO, USA) or in house [2,6-2H2]-l-phenylalanine.
SILAC was explored as a tool to determine whether or not the tryptophan synthase was constantly regenerated over time within the biofilm. We set pulse-chase experiments in which biofilms were matured in M63 medium supplemented with stable isotope labelled amino acids (pulse phase) and subsequently with unlabelled amino acids (chase phase). [4,4,5,5-2H4]-l-lysine (Sigma-Aldrich) and in house [2,6-2H2]-l-phenylalanine were used during the pulse phases and supplemented to the medium at final concentrations of 300 and 30 μM, respectively. The same concentrations were used for unlabelled l-phenylalanine and l-lysine during the chase phases.
Biofilms were grown by employing a spin coating method as previously described  with slight modification. 75 × 25 mm microscope slides (VWR International, Leicestershire, UK) were cut into 25 × 25 mm squares, sterilized and coated with approximately 1.5 mL of 0.1 % w/v poly-l-lysine in water (Sigma-Aldrich). Slides were dried in an oven at 60 °C and then transferred into 9-deep-well plates. A culture of PHL644 pSTB7  was streaked onto a LB agar plate with 100 µg/mL of ampicillin and single colonies were inoculated into M63 medium containing ampicillin. When the exponential growth phase (OD600 of 0.2) was reached, 600 μL aliquots were inoculated in 60 mL volume of M63 supplemented with either 30 μM of labelled phenylalanine or 300 μM of labelled lysine in 250 mL Erlenmeyer flasks, incubated at 28 °C, 150 rpm, in an incubator with a 19 mm throw until stationary phase (OD600 of 2.0) was reached. Cultures were then centrifuged at 1851g for 15 min and resuspended in fresh M63 medium supplemented with either 30 μM of labelled phenylalanine (triplicate cultures illustrated in Fig. 1, rows 1–3) or 300 μM of labelled lysine (triplicate cultures illustrated in Fig. 1, rows 4–6) to an OD600 ~1.0. 4 mL Aliquots of these suspensions were transferred to each well of a 9-deep-well plate containing a 25 × 25 mm poly-l-lysine coated slide (the labelled phenylalanine containing culture was introduced 9 deep well plates, illustrated in Fig. 1, rows 1–3 and the labelled lysine containing culture introduced into a further 9 deep well plates (Fig. 1, rows 4–6), enabling each experiment to be carried out in triplicate). Cells were deposited onto the slides by centrifuging at 2200g for 15 min using a JS 5.3 rotor in an Avanti JXN-26 centrifuge (Beckman, Munich, Germany), media was removed and 4 mL of fresh M63 media supplemented with the same labelled amino acid (either 30 μM of labelled phenylalanine or 300 μM of labelled lysine), as the cells were previously grown in, was added. The biofilm samples were incubated at 28 °C in an orbital shaker incubator at 50 rpm with a throw of 19 mm and left to mature. The 18 biofilms were initially left to mature in the two sets of labelled M63 media for 3 days (pulse phase). At day 3, three of the samples that had been grown in media enriched with labelled phenylalanine and 3 of the samples that had been grown in media enriched with labelled lysine were harvested for analysis (see next section). Media was removed from all of the remaining 12 samples and replaced with 4 mL of fresh media supplemented with either 30 μM of unlabelled phenylalanine (Fig. 1, rows 2, 3) or 300 μM of unlabelled lysine (Fig. 1, rows 5, 6), these samples were returned to incubate at 28 °C in an orbital shaker incubator at 50 rpm with a throw of 19 mm. On day 6, six further samples corresponding to Fig. 1, rows 2, 5 were harvested; the media from the 6 remaining samples removed and replaced with 4 mL fresh media containing the alternative label i.e. the samples that had previously been supplemented with phenylalanine were now supplemented with 300 μM of labelled lysine (Fig. 1, row 3) and those supplemented previously with lysine were now supplemented with 30 μM of labelled phenylalanine (Fig. 1, row 6). The samples were incubated for a further 3 days, under the same conditions prior to their harvesting and analysis.
At each time point (every 3 days), slides containing the biofilms were removed from the deep well plates. Planktonic cells were removed from the biofilm by gentle submersion and washing in aliquots of phosphate buffer (0.1 M KH2PO4, 7 mM serine, 0.1 mM pyridoxal phosphate (PLP), adjusted to pH 7.0), having an average of optical density (OD600) of 0.65 for 3 day-old biofilm, 0.25 for 6 day-old biofilm and 0.31 for 9 day-old biofilm in 4 mL phosphate buffer. Slides were transferred to 50 mL centrifuge tubes, immersed in phosphate buffer and the biofilms were detached by vigorous vortexing. The empty slides were carefully removed and washed and the resultant suspended cells were recovered by centrifuging at 17,000g. The biomass was then suspended in lysis buffer (50 mM Tris–HCl, pH 8.0, 25 mM NaCl, 5 % glycerol, 2.5 mM EDTA, 1 mg/mL lysozyme, Sigma) to which 1 volume of 2× SDS loading dye (100 mM Tris–HCl, pH 6.8, 4 % (w/v) SDS, 0.2 % (w/v) bromophenol blue, 20 % (v/v) glycerol, 200 mM DTT) was added; samples were then denatured, loaded in a 4–12 % polyacrylamide gel (NuPage, Thermo Fisher Scientific). After electrophoresis, gels were immersed in staining solution (50 % methanol, 40 % water, 10 % acetic acid, 0.1 % Coomassie Brilliant Blue G-250), followed by destaining. The bands of interest were excised from the gel, cut into 1 mm cubes and subjected to in-gel digestion, using a ProGest Investigator in-gel digestion robot (Genomic Solutions, Ann Arbor, Michigan, USA) according to standard protocols . Briefly, the gel cubes were destained by washing with acetonitrile and subjected to reduction and alkylation before digestion with trypsin at 37 °C. The peptides were extracted with 10 % formic acid and concentrated down to 20 μL using a SpeedVac (Savant™, Thermo Fisher Scientific).
Extracted peptides were separated on an Acclaim PepMap 100 C18 trap and an Acclaim PepMap RSLC C18 column (Thermo Fisher Scientific), using a nanoLC Ultra 2D plus loading pump and nanoLC as-2 autosampler (Eskigent™, Woodlands, Singapore). The peptides were eluted with a gradient of increasing acetonitrile, containing 0.1 % formic acid (5–40 % acetonitrile in 15 min, 40–95 % in a further 1 min, followed by 95 % acetonitrile to clean the column, before reequilibration to 5 % acetonitrile). The eluate was sprayed into a TripleTOF 5600+ electrospray tandem mass spectrometer (Sciex, Massachusetts, USA) and analysed in Information Dependent Acquisition (IDA) mode, performing cycles of 250 ms of MS followed by 100 ms MS analyses on the 15 most intense peaks seen by MS.
The MS data file generated via the 'Create mgf file' script in PeakView v 2.1 (Sciex, Massachusetts, USA) was analysed using the Mascot algorithm (Matrix Science, London, UK) , against the NCBI database  with no species restriction, trypsin as the cleavage enzyme and carbamidomethyl as a fixed modification of cysteines and methionine oxidation as a variable modification, to confirm the presence of the S. enterica tryptophan synthase TrpBA in the E. coli background. The data file was then searched against an internal database which includes the tryptophan synthase sequence from S. enterica. Additional variable modifications of [2,6-2H2]-l-phenylalanine and [4,4,5,5-2H4]-l-lysine, were added to the search parameters. Extracted ion chromatograms (XICs) corresponding to the peaks of interest, identified by the Mascot search, as peptides from tryptophan synthase were obtained through PeakView software 2.2. The level of incorporation was quantified by calculating the median of the labelled (heavy, H)/unlabelled (light, L) ratios according to the formula: Incorporation % = [H/L]median/(1 + [H/L]median) × 100 . Ratios of labelled and unlabelled peptides were calculated based on the peak area of XICs (areas under the curves).
Biotransformations were carried out as previously described [65, 70] using engineered biofilms that had matured for 7 days and 9 days in a potassium phosphate reaction buffer (0.1 M KH2PO4, 7 mM Serine, 0.1 mM pyridoxal phosphate (PLP), adjusted to pH 7.0). The biofilm maturation M63 medium was carefully removed from the biofilm covered slide by using a syringe. Unadhered, planktonic cells were removed from the biofilm by gentle resubmersion and washing in aliquots of reaction buffer, which was then also removed. Each of washed biofilm slides removed from the media after 7 and 9 days of maturation was then submerged in 4 mL reaction buffer. To start the biotransformation, the washed slides coated with 7 day-old biofilms (approximately 1.70 mg of total cellular protein present in the biofilms) and 9 day-old biofilms (~1.83 mg of protein) were submerged in 4 mL reaction buffer supplemented with 5 % (v/v) DMSO and 2 mM 5-chloroindole and incubated in an orbital shaker incubator (28 °C, 50 rpm with a 19 mm throw) for 24 h. The total protein content of the biofilm (average of 3 repeats) was determined using a Bradford protein assay and a protocol from Li et al. . 5-chlorotryptophan concentrations in biotransformation samples were analysed using a Waters Acquity UPLC system equipped with a Waters Acquity BEH C18 1.7 µm 2.1 × 50 mm column and PDA detector monitoring absorbance at 280 nm (solvent A: 0.1 % TFA in water, solvent B: acetonitrile. Gradient: 0 min, 5 % B; 0.5 min, 5 % B; 2.0 min, 50 % B; 0.5 mL/min). For analysis, solutions were taken from the biotransformation reaction and centrifuged (16,060g, 5 min), and equal volume of methanol was then added. The concentration of 5-chlorotryptophan was calculated based on a standard curve of UPLC peak area versus concentration. Samples of 5-chlorotryptophan of known concentration, ranging from 62.5 to 500 µM, were analysed by UPLC using the method described above. A standard curve of the relationship between UPLC peak area (y) and concentration (x) was constructed and the relationship between peak area and concentration was found to be linear (y = 4866.1x − 596.4). From this standard curve the unknown concentration of 5-chlorotryptophan in the biotransformation samples was determined and an estimation of the overall yield (Y) was calculated (Y % = 5-chlorotryptophan concentration/initial 5-chloroindole concentration). For LC–MS analysis samples dissolved in water: methanol 50:50 were separated using a Dionex Ultimate 3000 RS system (solvent A: 0.1 % FA in water, solvent B: acetonitrile. Gradient: 0 min, 5 % B; 0.5 min, 5 % B; 5 min, 37.5 % B; 5.1 min, 90 % B; 7 min, 90 % B; 0.35 mL/min), ionised using a H-ESI source (positive ion mode, 3.5 kV, heater 325 °C, capillary 350 °C, S-lens RF 47 %, sheath gas 55, auxillary gas 20, sweep gas 2) and analysed using a Thermo Fisher Orbitrap Velos Pro (30,000 resolution, 100–800 m/z).
Conception and design of the study: TTB, RJMG, MJHS and TWO. Acquisition of data: TTB and XT. Analysis and interpretation of data: XT, TTB, CHB, SVS, MJHS, TWO and RJMG. Contributed analysis tools: CHB. Drafting the article: TTB, XT and RJMG. All authors were involved in the writing of the manuscript and contributed intellectually. All authors read and approved the final manuscript.
All materials described within this manuscript, and engineered strains are available on request.
This work was supported by BBSRC Grant BB/I008713/2 and European Research Council under the European Union's Seventh Framework Programme (FP7/2007–2013/ERC Grant agreement no 614779) to Goss and BB/I006834/1 to Simmons.
12934_2016_579_MOESM1_ESM.docx Additional file 1: Figure S1. Biofilm-mediated biotransformation (previous work). Table S1. Amino acid composition of the subunit β of TrpBA calculated by ProtParam (Expasy). Table S2 In silico tryptic digest of the subunit β of TrpBA with Peptide Cutter tool (Expasy). Table S3 Fragments of the subunit β of TrpBA identified by Mascot. Figure S3. MS spectra analysis of the peptide VGIYFGMK of TrpBA. Figure S4. MS spectra of the peptide DPEFQAQFADLLK of TrpBA. Figure S5. The XICs of the reference peptide VGIYFGMK. Figure S6. The XICs of the peptide DPEFQAQFADLLK. Figure S7. Percentage of 5-chlorotryptophan accumulation, monitored by UPLC. Figure S8. LC-MS analysis of formation of 5-chlorotryptophan.


Some 600 of the island's special athletes will converge on the grounds of the Montego Bay Community College next month, for the National Special Olympics meet.
A total of 53 athletes will be selected from the November 11 meet to represent the country at the International Special Olympics to be held in Shanghai, China from October 2 to11, 2007.
Chairman of the Montego Bay Special Olympics organizing committee, Lee Bailey, told JIS News that the athletes, from all over the island, would be participating in team events in various sports disciplines.
More than 100 volunteers drawn from educational institutions, service clubs and uniformed groups, will provide support during the meet, which will get underway at 9:30 a.m.
Pointing out that the meet would cost some $1.5 million, Mr. Bailey thanked the various government and private entities, which have pledged monetary assistance towards the staging of the event.
The organizing committee chairman issued an appeal for public support of the event. "I believe strongly that the games can show that Montego Bay does have a heart," he said, adding that the games could also send a clear message that Montego Bay was capable of handling tourists, who are physically and intellectually challenged.
Mr. Bailey pointed that the Jamaican Special Olympics team has always done well at the International Special Olympics.
More than 7,000 special athletes from 169 countries would be participating in the 2007 games in Shanghai.


Space dedicated to safety of your own home: safes, locks and safety boxes are essential for safe keeping our valuables and our savings. Nowadays, thanks to the development of materials, a safe with key approximately 25 x 25 mm of excellent quality costs about € 80. Prices rise with mechanical or digital combination safes. The main goal of any manufacturer in the security sector is to use certified materials to resist as long as possible in attempts at burglary. Below we selected the companies with better value for money.
Safes There are 23 products.


Stunning panoramic views across the mountains and the sea!
Large private swimming pool perfect for cooling off!
Villa Ballencia is a fabulous, modern villa featuring stunning views of the mountains and the sea. This spacious and bright property offers ample space for eight guests to come together and enjoy an unforgettable holiday in the Costa Blanca.
The villa features modern and tasteful furnishings throughout, with each expansive room flowing into another and through to the outdoor spaces. Villa Ballencia is the type of house where you can meander between the interior and exterior with ease, soaking in every ray of sunshine that you wish! The main living areas are perfect for family style living, with a large lounge, boasting a comforting, open-hearth fireplace, satellite television, Hi-Fi and air conditioning. The lounge is the perfect place to spend some quality time together, reading, playing games or enjoying a movie on the DVD player. There is a small dining area, open-plan to the lounge, perfectly suited to morning breakfasts, as guests get up in their own time. The kitchen can be found through a couple of archways, offering all the equipment you may need to whip up some family feasts or a picnic to carry down to the beach! Throughout the villa there are two bathrooms, two double bedrooms and two twin bedrooms. One of the double bedrooms is a master suite, with a luxury bathroom, as well as a dressing area.
When guests step outside at Villa Ballencia, the remarkable, panoramic views of the sea, the mountains, the valley, the countryside, the forest and the bay truly take your breath away! There is a fabulous private swimming pool on the edge of the terrace, where guests can take a dip, cooling off on hot summer days. Villa Ballencia offers something for every guest, whether you desire days by the pool, water sports at the beach or a round of golf, this villa and amazing location leaves nothing to be desired!
When in Spain, act as the Spanish do and remember the sacred Siesta!
Villa Ballencia is spread over one floor. With a capacity of up to 8-guests, this is a great choice for a family holiday. The villa has a large pool and spacious exterior, perfect for meals in the sun.
Villa Ballencia is located in a quiet, sunny area just outside of the main resort on a slope. The villa is 8 km from the centre of Dénia, the secluded area has little traffic and offers a calm, relaxed atmosphere for guests, perfect for families. The villa itself sits surrounded by forest, 6 km from the sea and 6 km from the popular beach. This location is incredibly well connected, with trains, bus connections and even a ferry that can take you to Ibiza in only 3 hours!
Nestled between mountains and the sea, this jewel of the ever-popular Costa Blanca has charming waterfronts and beaches, and there is a huge range of local shops and markets along with supermarkets for more basic necessities, and a range of international cuisine if you fancy something different. The prestigious 18-hole golf course "La Sella", is so close to the villa, that even beginners might have to give it a go! This area is perfect for children and adults alike, with everything you might need for a fabulous holiday in Spain.
A family-friendly area. Spacious grounds and swimming pool ideal for kids.


text:
Fernandes breaks silence on tense handshake with Man Utd team-mate Ronaldo
Bruno Fernandes has attempted to downplay the speculation over his seemingly tense handshake with Manchester United team-mate Cristiano Ronaldo.
On Monday, footage emerged of Fernandes joining up with his Portugal team-mates ahead of the Qatar World Cup.
The moment he was greeted by Ronaldo raised eyebrows as Fernandes initially appeared to ignore Ronaldo's outstretched hand before belatedly shaking it and making a comment that left the five-time Ballon d'Or winner stony-faced.
Fernandes then walked away to greet other team-mates – doing so in a far more cheery manner.
The footage led to speculation that Fernandes was left annoyed by some of the comments Ronaldo made in his explosive interview with Piers Morgan.
Just hours before Fernandes arrived for World Cup duty, the first clips of the interview were released, with Ronaldo saying he feels 'betrayed' by Manchester United and does not trust Erik ten Hag.
But Portugal midfielder Joao Mario was quick to dismiss talk of a rift in the dressing room, claiming Ronaldo and Fernandes shared a joke during the aforementioned clip.
https://twitter.com/ESPNFC/status/1592434781555400704?ref_src=twsrc%5Etfw%7Ctwcamp%5Etweetembed%7Ctwterm%5E1592434781555400704%7Ctwgr%5E54358a14afead2eee3ac2fd02fc8169984ab821a%7Ctwcon%5Es1_&ref_url=https%3A%2F%2Fmetro.co.uk%2F2022%2F11%2F15%2Fman-utd-news-bruno-fernandes-breaks-silence-cristiano-ronaldo-handshake-17763450%2F
'I was in the locker room at that moment,' Mario said. 'I saw pictures. It was a joke between them. Bruno Fernandes was one of the last to arrive. Cristiano asked: 'Did you come by boat?'"
Fernandes has now appeared to corroborate Mario's comments, writing on social media: 'Oh Joao Mario, there you are ruining the newspaper headlines', with a facepalm and laughing emoji.
While Fernandes has attempted to downplay his eyebrow-raising greeting with Ronaldo, a number of Manchester United players are said to be 'furious' at the timing of Ronaldo's interview with Morgan.
The first clips were aired on Sunday night, just a few hours after Manchester United scored an injury-time winner to beat Fulham at Craven Cottage.
Fernandes attempted to downplay the incident
Ronaldo is considered a legend at Old Trafford but former Liverpool defender Jamie Carragher believes the 37-year-old now 'wants to be sacked'.
'This was always going to happen,' he told Sky Sports. 'It was always going to end like this.
'That's exactly why I said 18 months ago: it was a massive mistake for Manchester United to bring Cristiano Ronaldo back to the club.
'I just thought the comments [revealed on Sunday] were bizarre. They were bang out of order considering the timing of it: just hours after Manchester United fans being delighted after getting a winning goal at Fulham with the last kick of the game.
'It should give them excitement over the next month for when the league starts again. They should be enjoying themselves and then the Ronaldo interview comes out and boom – it's about negativity.
'Not the team, not the manager – it's about Cristiano Ronaldo. And it's been like that since he's come back. It's basically been the Cristiano Ronaldo show.
'At times that's been good on the pitch, last season he got a lot of goals. But this season he's not involved and he's craving the attention that he can't get because he's not on the pitch.
'The only way he's going to do that is by causing trouble and that was exactly his intention with the interview.
Cristiano Ronaldo hit out at Manchester United in a stunning interview
'He wants to be sacked, that's obvious. He told the club in the summer that he wanted to leave, which is football – these things happen. There were no takers, maybe because of his wages or the price Manchester United wanted.
'I think he knows that's going to be the same in this next transfer window and maybe the following summer and he is now trying to be sacked or wants to leave on a free. That's the only way he's going to get out.
'And I think United should sack him or move him on by giving him a free transfer. At the moment, they're trying to build something that is not trying to win the Premier League or the Champions League right now.
'The manager will want to push for those honours in the next two or three years and Ronaldo is not going to be around.
'He is going to be a distraction, this will not go away. As soon as a player comes out and says he doesn't respect the manager, how is that going to work when he walks into the first day of the post-World Cup season?
'It can't work that going forward – there will be people taking pictures, we will be asking about Ronaldo to the manager in interviews for Sky. United don't need the hassle.'


The PetParent app is a super fun and interactive way for pet owners to find new, local activities for their furry friends and interact with other pet owners through a central newsfeed.
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A pet's health is of top importance, which is why the app provides push reminders for things like vet appointments, shots, and medication refills. Is Fido due for his checkup? Ding, the app puts a reminder on your phone so he'll always make his appointments on time! Plus, PetParent works with veterinarians and animal specialists to offer its users exclusive tips and articles on things like preventive care and dietary advice.
Users really liked the events listings and photo contests, but one challenge we had was getting them to treat the app as a social site where they could follow and interact with each other. A major hurdle of any social community is rebuilding the social connections users already have on other platforms. A solution we implemented was having the app import connections of the user's Facebook friends who also had the app, therefore users didn't need to re-follow people they already knew. (This was a huge benefit of having people sign up through Facebook.) The newsfeed aspect of the app also helped boost engagement, since users saw other people's activity every time they logged in.
With minimal input from the client, the Jackrabbit team handled full design and development for both iOS and Android native apps.
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The app's social aspect is key for getting users to interact more.
PetParent produces content for pet lovers in big cities, and needed a way to reach a bigger audience and get current readers more involved. We developed a social app for iOS and Android that gives pet owners an easy way to find local, pet-friendly businesses and events.