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Welcome! Login
This United Arab Emirates Dirham and Macedonia Denar convertor is up to date with exchange rates from May 23, 2019.
Enter the amount to be converted in the box to the left of United Arab Emirates Dirham. Use "Swap currencies" to make Macedonia Denar the default currency. Click on Macedonia Denars or United Arab Emirates Dirhams to convert between that currency and all other currencies.
The United Arab Emirates Dirham is the currency in United Arab Emirates (AE, ARE, UAE). The Macedonia Denar is the currency in Macedonia (The Former Yugoslav Republic, MK, MKD). The symbol for AED can be written Dh, and Dhs. The symbol for MKD can be written MKD. The United Arab Emirates Dirham is divided into 100 fils. The Macedonia Denar is divided into 100 deni. The exchange rate for the United Arab Emirates Dirham was last updated on May 22, 2019 from The International Monetary Fund. The exchange rate for the Macedonia Denar was last updated on May 23, 2019 from Yahoo Finance. The AED conversion factor has 6 significant digits. The MKD conversion factor has 4 significant digits.
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It's Friday, so as promised time to announce the winner of my giveaway!
A big congratulations to Abbie from Black Lace And Glitter!
Abbie, please email me your address to firstname.lastname@example.com and I'll get that sent over to you!
A big thank you to everyone else who entered, I really appreciate all your support!
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subset: commoncrawl


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National & World Ag News Headlines
U.S. September Tractor, Combine Sales Up
USAgNet - 10/11/2018
According to the Association of Equipment Manufacturer's monthly "Flash Report," the sale of all tractors in the U.S. in September 2018, were up 2% compared to the same month last year.
Agri Marketing magazine reports that for the nine months in 2018, a total of 181,515 tractors were sold which compares to 168,651 sold thru September 2017 representing an 8% increase for the year.
Two-wheel drive smaller tractors (under 40 HP) were up 2% from last year, while 40 & under 100 HP were down 7%. Sales of 2-wheel drive 100+ HP were up 30%, while 4-wheel drive tractors were up 51%.
For the year, two-wheel drive smaller tractors (under 40 HP) are up 10% from last year, while 40 & under 100 HP are up 2%. Sales of 2-wheel drive 100+ HP are up 9%, while 4-wheel drive tractors are up 19%.
Combine sales were up 9% for the month. Sales of combines for the year total 3,560 compared to 2,936 in 2017, a 21% increase.
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Other National Headlines
USDA Announces Strategy for Tariff Support to Farmers
AccuWeather See Substantial Shortfall in Corn & Soybean Yields
Congress Agrees to $19 Billion Disaster Aid Package
USDA Alerts Vendors to Fraudulent DocuSign Email
Hormel Cuts Forecast on Rising Pork Prices
Higher Limits Now Available on USDA Farm Loans
Bill Would Place Moratorium on Ag Mergers
Copyright © 2019 - USAgNet.com. All Rights Reserved.


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We were in the mood for some porridge. Honkee immediately came to mind. But we could not find them any more. Their outlet in PJ Uptown was no longer there. So was their branch in USJ10, Subang Jaya. Both had closed shop. We wondered if Honkee, Klang Valley wide, had finally called it a day. While we cruised USJ10 with blank minds, wondering what to have for dinner, we stumbled on this place - Tasty Porridge.
The place is located on Jalan USJ10/1C, opposite the car park complex in USJ10. It was just our luck, our crave for porridge was not to be in vain. But the place was packed. This apparently was a popular makan place. We were going to find out if their food was that good to attract all these people.
Porridge was obviously the mainstay of what they had to offer. But they also served rice. And there were quite a number of dishes that went with rice - including, of all things in a porridge shop, fish head curry.
We came for porridge and we stuck with that. I ordered a bowl of my favourite - the pork innards porridge (chu chap chok - 猪杂粥). It was good. The porridge was typically Cantonese (Hongkong style), boiled to a gruel. In my bowl were adequate variety of pork innards. On top were pieces of deep fried pork intestine. These fried intestines are absolutely my favourite - one of the reasons why I love this pork innards porridge. But I found the fried intestine insufficient. I wish they had been more generous on that. Still I enjoyed the porridge. It did not disappoint.
My wife's choice was rather unique - minced pork and salted egg. I had a taste of it. It was good. The salted egg blended very well with the minced pork and the porridge was flavourful. She liked it enough to order it again on a subsequent visit.
The yew char kueh (油炸鬼) to go with the porridge was surprisingly good. It appeared freshly fried, still crispy and not at all soft or flaccid. Dipped into the porridge, the crispy YCK tasted very good indeed. I was impressed that the shop serves such fresh YCK. I think they made their own YCK as against buying them off the street which by then would be limp and totally not the same.
On the table were various condiments - sesame oil, pepper, fried shallots in oil and soy sauce - to complement our porridge.
We liked the porridge so much that we re-visited the restaurant a couple more times. And each time we ordered one or two side dishes. This salty chicken dish had a thumb-up in the menu. And indeed, it tasted very good. It was served piping hot in a claypot. The aroma was mouth-watering. The sauce was salty and tasty - with some goji (wolfberries). And the chicken meat was tender.
This mui choy stewed pork (梅菜肉) also had a thumb-up. And it too was served hot in a claypot. It was a very well done. The flavour was really good. They blended the mui choy (梅菜) very well with the meat. It was not at all salty. The belly meat had some layers of fat in it - sinful but what good meat is without some fat?
This soft tofu (水豆腐) is more suited for rice. But we had it anyway. It was quite ordinary.
So was the blanched vege. There was a good amount of deep fried shallots. Certainly made a difference to the flavour.
This place is a porridge haven. Besides the pork innards, and the salted egg and mince meat, they have a good variety of other porridge flavours. And if your dining partner is not in the mood or dislike porridge, there is always rice and dishes for him or her.


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Disney Signs, Disney Props, Disney Park used signs, Disney Park Used Props, Animal Kingdom, Epcot, Magic Kingdom, Magic Kingdom Signs
In stock
This is the only known remaining directional sign from the American Idol Experience at Disney's MGM/Hollywood Studios. It features the attractions logo and includes a removable arrow. Perfect for the Disney or American Idol Fan! Measures 32 1/2" X 22" and is made of a sturdy plastic. Don't miss out on an opportunity to own a piece of a bygone attraction!


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Small and moderate-sized endeavors ( SMEs ) play a really of import function in heightening economic development in Laos. However, SMEs in Laos are little and lack fight. The influx of foreign direct investing ( FDI ) and trade liberalisation in Laos bring chances and challenges for SMEs in Laos. Furthermore, issues of SMEs and appraisal of SMEs publicity policy are non good understood for deficiency of surveies. This paper survey provides an overview of the current state of affairs, issues, and policies facing SMEs in Laos. It evaluates the impact of FDI and trade liberalisation on SME development. Although the authorities of Laos has defined policies and scheme to back up SMEs, it lacks specific plans that could flesh out these policies. SMEs are therefore faced with the jobs of high revenue enhancement aggregation, high rising prices, unstable exchange rate, and unequal support. Furthermore, SMEs are confronted with jobs of innovativeness, deficiency of fight, limited market, and networking. FDI provides SMEs with chances to increase their production through farm outing production linkages. The impact of trade liberalisation on SMEs is non good understood but trade liberalisation seems to hold negative impact instead than positive impact on SMEs.
Lao people's democratic republic joined the Association of Southeast Nations ( ASEAN ) in 1997 and ASEAN Free Trade Area ( AFTA ) in 1998. As portion of its committedness under the AFTA understanding, Laos has to cut down the duty rates to 0 to 5 per centum in 2008. Reducing duty rates would increase imports, and have a negative impact on domestic economic system. Furthermore, increasing FDI is expected to play an of import function in increasing SME productiveness. On the other manus, FDIs may cut down the figure of domestic investings, particularly those of SMEs. Most SMEs in Lao face challenges and restraints, such as deficiency of fiscal support and inadequate skilled labour, and low degree of fight.
It is widely accepted that SMEs play a really of import function in Lao 's economic system. In 2004, medium endeavors numbered about 722 and little endeavors, 25,271. In term of employment, SMEs offer more than 60,000 occupation chances, which accounted for 40 per centum of the state 's entire employment. To salvage Lao SMEs from the inauspicious impact of globalisation, it is important to place the issues facing SMEs and measure the authorities policies and the impact of trade liberalisation and FDIs on SMEs.
For deficiency of surveies, nevertheless, on SMEs and appraisals of SMEs publicity policies in Laos, bing policies and plans toward SMEs are non good understood. In this respect, this paper provides an overview of SMEs in Laos and assesses SMEs publicity policies in Laos. The specific aims of this paper are:
a. To supply an overview of the authorities policies refering SME development and their present state of affairs in Laos and Vietnam
B. To analyze the innovativeness, market enlargement, fight and networking affecting Lao SMEs compare to neighbour states
c. The impact of trade liberalisation and FDI on SME development
The Lao authorities 's Policies ( SMEs )
The Lao authorities began advancing gradual economic liberalisation through the debut of the New Market Mechanism in 1986. This was followed by the Law on Promotion and Management of Foreign Investment and Business Law in July 1994. To advance and supply more inducements to FDI, the Law on Promotion and Management of Foreign Investment was amended in October 2004. In add-on, Lao authorities besides began carry oning the Business Forum in Laos to ease duologues with the concern sector. Government attempts toward SME publicity were expressed in Primary Office Decree No. 42/PM, the ends of which were as follows: a ) better the regulative environment ; B ) enhance the fight of constitution ; degree Celsiuss ) expand domestic and international market entree ; vitamin D ) better entree to finance ; e ) promote the development of concern organisation ; f ) enhance entrepreneurial attitudes and features within the society. The National Small and Medium Sized Enterprise Office ( SMEPDO ) was established by virtuousness of the Prime Minister 's Decree No. 42/PM. Its chief aim was to advance the constitution and sustainable development of Lao SMEs.
Since SMEs in Laos are in the early phase of development, innovativeness slowdowns behind adjacent states. Worse, informations coevals on innovativeness is non yet good developed. A figure of establishments exist to supply invention preparation for SMEs, but their attempts are non plenty to raise the degree of SME innovativeness. They had seen the demand for SME proprietors to spread out their merchandise lines.
Among SME merchandises, merely handicrafts appear competitory pricewise. The cost of handcraft merchandise is cheaper than those of other states in the ASEAN part, since labour costs in Laos are really low. The mean pay of Lao worker is around US $ 30 a month while the mean pay of Vietnamese worker and Kampuchean worker is 40 US $ and US $ 35, severally ( ALGI 2002 ) . However, the productiveness of Lao workers is besides lower compared to neighbouring states like Vietnam and Cambodia.
Market enlargement
In the absence of informations on SME market portion in the domestic market, this survey opted to utilize the figure of constitutions as a step of local market enlargement. The figure of new SME constitutions could explicate the enlargement of the domestic market. In 2005 there were 77,651 registered constitutions. Two old ages subsequently, in 2007, SME constitutions, harmonizing to SMEPDO, were estimated to figure about 135,000. This translates to about 73.5 per centum compared to 2005.
With discriminatory GSP intervention from other states, Lao 's chance for its SMEs to increase its production and heighten its exports, particularly garments and handcrafts. In 2005, the garment export sum to 142 million US $ . Garment SMEs portion of entire export was about 30 per centum. Export market seems to hold expanded compared to the old twelvemonth.
Past networking between SMEs, contractors, and providers seemed to be really hapless. Many SMEs were run as household concerns. Some succeeded, but some failed. With the coming of trade liberalisation and FDI influx, SME fight was enhanced ; so was networking between SMEs, contractor, and providers. Yet, the deficiency of back uping authorities and private institutes including Bankss do non portend good for SME development in Laos, even where networking is concerned. Hence, external networking of SMEs in Laos remains weak and need more support from authorities.
Impact of FDIs on SMEs
FDIs have played a really of import function in Lao economic system. It can speed up economic growing and income of the people. FDIs besides help SMEs develop by easing the transportation of engineering, bettering cognition and accomplishments, among other benefits.
Impact of Trade Liberalization on SMEs
The authorities of Laos continues to implement assorted trade reforms to increase growing. Duty decrease is being pursued as portion of the AFTA committedness. Furthermore, it plans to fall in the World Trade Organization by 2010. Trade liberalisation has positive and negative impacts on SMEs in four major ways: 1 ) by increasing competition ; 2 ) by take downing production costs due to cheaper imported inputs ; 3 ) by increasing export chances ; and 4 ) by cut downing handiness of local inputs. However, Lao SMEs are really little and non every bit competitory as those in adjacent states. Therefore, trade liberalisation might hold negative impact on SMEs. On the other manus, Lao SMEs could increase their fight by diminishing the cost of input and market enlargement.
In amount, it is non clear if Lao SMEs will profit from trade liberalisation. Furthermore, the benefits of trade liberalisation will hold a negative impact on SMEs if the authorities of Laos disregards to advance SMEs to better their comparative advantage for increasing exports and to reform the domestic revenue enhancement and aggregation system. Furthermore, as Laos is a landlocked state, transit cost is one of the most of import issues environing SMEs ' fight. It is of import for authorities of Laos to happen ways to cut down transit costs.
Obstacles of SMEs
Lack of skilled labour, deficient stuff, rigorous ordinances, deficiency of market, expensive natural stuff, rival of the same concern, unstable exchange rate, deficiency of fiscal support, high revenue enhancement, high rising prices


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Estate homes
On this page you will find the solution to Estate homes crossword clue. This clue was last seen on LA Times Crossword November 19 2018 Answers In case the clue doesn't fit or there's something wrong please contact us.
Estate homes
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- » Separate lounge inc.TV
- » Two fridge/freezers
- » Off-road parking
- » Shower room
- » Gas central heating
- » Burglar alarm
- » En-suite shower
- » Lrge kitchen inc.dining area
Bills inclusive option available. See Utilities section on our website.
This student property is located just off the main Crookes high street which has many shops and services including Sainsburys & the Co-Op. This area is very popular with students.
The property benefits from double glazing, gas central heating and hot water by combi boiler and burglar alarm.
The large fitted kitchen is of a high standard and includes a 2xlarge fridge/freezers, dining table, chairs and washer&dryer. The kitchen leads through to the lounge which is bright and spacious with two sofas, further fold out dining table, chairs, coffee tables and a 37" TV.
There is a a shower room on the first floor and the first floor front bedroom has an en-suite shower. All bedrooms are of a good size and feature double or three quarter sized beds, wardrobes, chests, desk and chairs.
There is also an off road private parking space.
EPC Rating – E 44


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GREG TARR/Observer-Reporter/via AP Ralph Ullum gets kisses from his Siberian huskies at his property in East Finley Township near Claysville, Pa. A Pennsylvania breeder of Siberian Huskies has accused the American Kennel Club of defamation for suspending him from club shows based on an allegation that he once drugged another dog to give his a competitive edge. Ralph Ullum, a ...Read More »
Top Stories
Ariel Castro neighbor charged with 1990s rape and murder of two women
Cuyahoga County Sheriff's Office Elias Acevedo Sr., 49, confessed to the murders of his 30-year-old neighbor Pamela Pemberton and 18-year-old Christina Adkins. Increased scrutiny of missing person cases in a Cleveland neighborhood following the arrest of kidnapper Ariel Castro led to charges against a neighbor for the murders of two women in the 1990s, the FBI said on Friday. Elias ...Read More »
A-Rod shelled out $305G for evidence linking him to doping: source
David Karp/AP A source says Yankees star Alex Rodriguez paid $305,000 for evidence that could have incriminated him or that would be used in his defense in an arbitration or court proceeding. As Major League Baseball closed in on Tony Bosch's doping operation, a desperate Alex Rodriguez paid $305,000 for evidence involved in the investigation linking him to multiple violations ...Read More »
Same-sex marriages to begin Monday in New Jersey
Published: October 18, 2013 10:25 PMBy THE ASSOCIATED PRESS Photo credit: AP | Same-sex marriages can begin within days in New Jersey after the state's highest court ruled unanimously Friday to uphold an order that they must start Monday and to deny a delay that had been sought by Gov. Chris Christie's administration. Same-sex marriages can begin within days in ...Read More »
Lunar eclipse brings hundreds to mysterious NY site
Published: October 19, 2013 8:00 AMBy WATERTOWN DAILY TIMES Photo credit: AP | The moon seen from the Belarusian capital Minsk, during a partial lunar eclipse. (April 25, 2013) WATERTOWN, N.Y. – On the very night of a rare partial lunar eclipse, believers and non-believers alike came together on Friday night to pay tribute to Thompson Park's so-called vortex, an ...Read More »
Joseph Jenkins and Charles Walker, who escaped from Fla. prison, used forged documents
Originally published: October 19, 2013 9:56 AMUpdated: October 19, 2013 9:57 AMBy THE ASSOCIATED PRESS Photo credit: AP | This undated photo provided by the Florida Department. of Corrections shows Joseph Jenkins. Jenkins and Charles Walker were mistakenly released from prison in Franklin County, Fla., in late September and early October. According to authorities, the the two convicted murderers were ...Read More »
Kronwall taken off on stretcher as Avs-Red Wings gets ugly again
DENVER – Jonas Gustavsson stopped just about everything, including the Colorado Avalanche's six-game winning streak. The backup goalie turned in a 38-save performance and Johan Franzen broke a third-period tie with his second goal of the game as the Detroit Red Wings beat the Avalanche 4-2 on Thursday night to hand Patrick Roy his first loss as a coach. "He ...Read More »
PIC: $1.6 million supercar painted GOLD
Alex Penfold / SWNS.com/Alex Penfold / SWNS.com The supercar, which can race as fast as 252 mph, is registered in Saudi Arabia and sports a '1' license plate. Owning a $1.6 million supercar just wasn't enough for this oil head. No, the showoff with the keys to this Bugatti Veyron decided his ride also had to be painted an ostentatious ...Read More »
Malala meets Queen Elizabeth II at Buckingham Palace in London
POOL/REUTERS Malala Yousafzai gives a copy of her book, 'I am Malala,' Britain's Queen Elizabeth during a Reception for youth, education and the Commonwealth at Buckingham Palace in London. LONDON — Malala Yousafzai skipped school for the day but she had a pretty good excuse: she was meeting Queen Elizabeth II. The 16-year-old advocate for girls' education and survivor of ...Read More »
Obama surveys shutdown damage and blames GOP
Originally published: October 17, 2013 10:39 AMUpdated: October 17, 2013 3:33 PMBy THE ASSOCIATED PRESS Photo credit: AP | President Barack Obama speaks in the State Dining Room of the White House in Washington. (Oct. 17, 2013) The government unlocked its doors Thursday after 16 days, with President Barack Obama saluting the resolution of Congress' bitter standoff but lambasting Republicans ...Read More »


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Our neighbours across the Tasman have been waiting years for it, but the day has finally come… Vuly trampolines (and now swing sets) are available for purchase throughout New Zealand. The Lift, Thunder and Thunder Summer are ready for backyards this Christmas, and we think that they'll find as much love in NZ as they have in Australia!
We want our New Zealand fans to have the same fantastic online shopping experience as local Vuly customers. NZ shoppers can purchase Lift and Thunder from VulyPlay.com/en-NZ in the same way as those in Australia, and 360 Pro is also available in size M with the unique Spin and Yoga swings. It won't be long before we expand our range based on any and all valuable feedback.
Our Australian Sales and Service team is already standing by to assist via phone (toll free at 0800 661 238), LiveChat on our website or through social media (Facebook.com/Vuly)! With our freight partner, Mainfreight – New Zealand's largest freight company – Vuly will be distributing its products in the most efficient manner possible, and with the lowest cost to the consumer.
Vuly is also working with Torpedo7 and No. 1 Fitness stores to bring the incredible Thunder Summer to New Zealand. This particular version of Thunder Summer features the enclosure of the next generation Thunder Pro, combined with the tried-and-true Leaf Spring design of the Australian Thunder Summer. First sold in Australia in 2014 – and now upgraded just for NZ – it's the ideal top-tier trampoline for springless bouncing at its best. This model is exclusive to Torpedo7 and No. 1 Fitness; visit our store locator to find your nearest store.
We're so excited to finally have a presence in New Zealand; just in time for Christmas 2017 and our biggest year at Vuly yet!
Vuly owners on Instagram


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Award Objective
Everyday people in our community are helping Twisted Pink make an impact towards our mission to fund metastatic (stage IV) breast cancer research. These people provide valuable donations and advocacy work along with the precious gift of time and talents.
The Dace Polk Brown Ambassador Award was established in 2017 in honor of Dace Polk Brown. Dace Polk Brown is a breast cancer survivor and a member of the Twisted Pink board of directors. She has been a vital part of our mission since joining Twisted Pink as a donor and volunteer board member. Twisted Pink has donated over $423,000 to metastatic breast cancer research since June 2014. In 2017 we were founding members of a metastatic breast cancer research collective. Twisted Pink, along with Hope Scarves and The Cancer Couch secured a 1:1 foundation match which amounted to 1.2 million dollars in metastatic breast cancer research funding at Dana Farber Cancer Institute and Memorial Sloan Kettering Cancer Center.
The Dace Polk Brown Ambassador Award is given annually to one individual in each category who shows commitment and support to the mission of Twisted Pink. The award is given to volunteers, donors, patient advocates, and others who believe in our mission to fund metastatic breast cancer research. Nominations are accepted between July 1, 2018 and September 1, 2018. Nominees will be notified by October 1, 2018 and winners will be announced at the 2019 Masquerade Ball on Saturday, January 26, 2019 at The Speed Art Museum.Past Award Recipients
Nominee Qualifications
Any volunteer living in the United States is eligible for the Dace Polk Brown Ambassador Award. Only one person may be nominated on a form.
Award Categories
Breast Cancer Survivor Award Recipient – One breast cancer survivor is eligible to receive the award annually.
Advocate Award – One community advocate is eligible to receive the award annually. This person does not have to be a breast cancer survivor.
Community Impact Award – A small business or corporation is eligible to receive the award annually.
Judging Criteria
The Twisted Pink Nominations & Awards Committee will select the Dace Polk Brown Ambassador Award Recipient from the nominations submitted to Twisted Pink. The committee considers:


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Making a spectacle of EMF
In the many glass cabinets at Cyberdog were lots of laser cut acrylic bangles and spectacles. I thought it would be cool to make some of these things up at camp but in the end I spent the whole time cutting badge cases (literally 12 hours a day). I did make this one prototype though which somebody loved enough to take home again. The arms were bent using a hot air gun, although I have done 'off focus' laser bending before it's definitely quicker doing it this way.


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Victorian Porcelain Dolls
Victorian Porcelain Dolls
Porcelain baby dolls continue to be quite popular collectible figurines. Meanwhile, a number of individuals retain a powerful interest in the period and also its style. Therefore, it isn't shocking that a wide variety of businesses produce a large range of Victorian porcelain dolls.
Don't assume all Victorian porcelain dolls are made equally, however. Many of the available for sale dolls which claim to be symbolic of the era are susceptible to horrifying errors. You should become aware of the best way to detect a porcelain doll which thoroughly copies the fashion of the Victorian era prior to choosing one as a present or for your own private lineup.
Victorian daytime garments tended to be made from lighter fabric. Linen was widely used during warmer months. Flannels and also muslins were fabrics of choice through the cool season. One will often find Victorian porcelain dolls clothed in daywear created with fabrics that quite frankly wouldn't normally have been used for that purpose. The noble evening dresses in the Victorian period included fabrics including taffeta, organdy and silk. It is smart to choose Victorian porcelain dolls that use those identical fabrics. You might choose to consult some of the remarkable available references on Victorian dress to find out whether a doll's apparel is genuinely representative of the age.
. Instead of simply buying porcelain dolls for your children at the start of their hobby, you should encourage them to pay at least part of the cost. They can engage in summer jobs or work errands for you so that they can pay for the dolls that they like. This will instill a sense of value within your kids and also help them appreciate each new doll as they engage in doll collecting with a greater sense of maturity. This will also help you to save larger amounts of money when you embark on buying collectible porcelain dolls that are quite costly to buy as compared to regular dolls.


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Email: email@example.com
Both residential complexes and shopping malls have something in common: the need for daily communal areas cleaning. No matter how many advantages your building has to offer in terms of amenities or attractions, the first thing that potential customers see are the outside windows, the entrance door and the main hallway.
Unless they are perfectly clean, you will not even get the chance to take your potential client for the grand tour. You may be able to hide some flaws or draw your clients' attention away from them, but cleanliness cannot be overlooked. A cluttered, dirty and bad smelling building will chase everyone away.
Professional companies have developed their communal areas cleaning services after years of analysing and understanding their clients' needs and the restrictive conditions these operate under.
For example, in a residential building, it is absolutely the worst moment ever to start cleaning the corridors at 7 a.m., when everyone is preparing to leave for work. The 2 to 4 in the afternoon period is the nap hour for small children and senior citizens, and you must avoid any noisy activities, such as vacuuming the hallway carpets.
Shopping malls need to be sparkling clean when they open for business and stay like this throughout the day. The important aspects to consider in this situation are how to plan the general daily communal areas cleaning in order to finish on time and how to maintain cleanliness during the day without major impediments to clients walking up and down the corridors.
Professional cleaning companies in Guildford are ready to meet all exigencies imposed by corporate and residential clients. Besides the knowhow required to ensure high level services in communal areas cleaning, these companies have developed a series of work procedures that help them get the job done efficiently and consistently, with minimum disturbance to the people living, working or shopping in the respective building.
Here are a few reasons for working with cleaning companies:
Before hiring a cleaning company, you need to do a background check. These days, you can easily find reviews and customer testimonials on various internet forums or on the companies' websites. Also, you should check their clients portfolio and assess if they have experience in large scale cleaning jobs.
Star Shiners has all the qualifications required to become your trusted cleaning partner. Their staff is carefully selected and trained, and they are committed to delivering superior services for affordable prices. If you are looking for efficient and high quality communal areas cleaning in Guildford, look no further than Star Shiners – the company that keeps its promises.
If you are fed up with doing all that cleaning and you are considering hiring one of the trustworthy, efficient and affordable cleaning services Guilford offers, you should hire the best – you should hire Star Shiners. For us, cleaning is much more than a business to live by – it is our passion, too, so with us you can be sure we will make everything in your home or office sparkle and shine with cleanliness.
Call: 01483
Email: firstname.lastname@example.com
Partner Bournemouth Sites
2015 Copyright Star Shiners | Powered by SEO Express | SEO Bournemouth


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Diep — Lahaina Maternity Photographer
What a beautiful belly! Diep is due in 3 weeks and took the plunge for maternity photos on baby #3. She barely looks old enough to have 1, much less 3!!! Little Alex hung around and tried all sorts of various ways to make sure I couldn't get his photo.. but I got it anyway :)


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Register for: KubeCon + CloudNativeCon North America 2018 from December 10-13, 2018!
Don't miss KubeCon + CloudNativeCon North America 2018 from December 10-13, 2018 in Seattle, Washington.
The Cloud Native Computing Foundation's flagship conference gathers adopters and technologists from leading open source and cloud native communities in Seattle, WA from December 10-13, 2018. Join Kubernetes, Prometheus, OpenTracing, Fluentd, gRPC, containerd, rkt, CNI, Envoy, Jaeger, Notary, TUF, Vitess, CoreDNS, NATS, and Linkerd as the community gathers for three days to further the education and advancement of cloud native computing.
December 10: Community Co-Located Events & Conference Lightning Talks
- Cloud Foundry Day: KubeCon + CloudNativeCon is the place for the cloud-native community, and Cloud Foundry is synonymous with cloud-native. From startups to the Fortune 500, Cloud Foundry is used by enterprises around the globe to automate, scale and manage cloud apps throughout their lifecycle. Cloud Foundry Day at KubeCon + CloudNativeCon Europe is an educational event designed for the best minds of the cloud-native world attending KubeCon + CloudNativeCon.
- Kubernetes Contributor Summit: The Contributor Summit brings together new and current Kubernetes contributors alike to connect and share face-to-face. It is an opportunity for existing contributors to help shape the future of community development and governance and offers a welcoming space for new community members to learn, explore and put the contributor workflow to practice.
- OpenShift Commons Gathering: Join key stakeholders, customers, upstream project leads, and contributors that make up the Red Hat OpenShift ecosystem for a full day of talks, demos and case studies from stakeholders, customers, and contributors from across the OpenShift ecosystem. The OpenShift Commons Gathering brings together experts from all over the world to discuss real-world implementations of container technologies, best practices for cloud native application developers and the upstream open source software projects that make up the OpenShift ecosystem.
December 11-13: Conference
About the Linux Foundation:
The Linux Foundation is dedicated to building sustainable ecosystems around open source projects to accelerate technology development and industry adoption.
Founded in 2000, The Linux Foundation provides unparalleled support for open source communities through financial and intellectual resources, infrastructure, services, events, and training. Working together, The Linux Foundation and its projects form the most ambitious and successful investment in the creation of shared technology.
For more information or to learn more about the event, visit "KubeCon + CloudNativeCon North America 2018".


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Well, they've done it again. This time its the folks at Heroclix, who put out some teasers for their new line celebrating DC Comics 75th Anniversary. Heroclix has really done a tremedous job in capitalizing on the Blackest Night and Brightest Day events by releasing sets very quickly.
So who is set to become a White Lantern? The answer is Oliver Queen, the Green Arrow.
Now Green Arrow was a White Lantern for a couple of pages in Blackest Night as well, and while that was a cool moment, I don't think that moment alone gets you a heroclix, but maybe I'm wrong.
What I do know is that at the end of the most recent issue of Green Arrow, we see Oliver Queen laying on the ground near the tree with the White Lantern Symbol. He also coincidently has an arrow shot directly through his skull. That's hard to recover from, even if you have a good health care plan.
So, right now he is dead, but I don't really think he's going to stay that way? Do you?
So instead you bring back Green Arrow in the next issue -- as a White Lantern.
Is this yet another case of the tail wagging the dog, and scooping the story with related merchandise advertising? Seems like it to me. This doesn't seem like a hard problem to solve, but it seems that DC Comics is having a very hard time figuring it out lately. Hopefully that changes soon.


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Professional Community for Car Dealers, Marketing, Advertising and Sales Leaders
The word "why" is popularly used in essay questions so that students can demonstrate their knowledge, reasoning and understanding of a particular subject. The Oxford Advanced Learners Dictionary defines "why" in this context as being "used in questions to ask the reason for or purpose of something".
Asking a student "why" allows them to demonstrate a more advanced knowledge of their subject matter, for example by analysing the processes involved in a chemical reaction, rather than just writing a list of reasons. It's easy to get into a mess trying to order your thoughts onto paper, so this essay strives to give you the top five tips on structuring an essay that attempts to respond to a "why" question.
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- Research
- Open Access
Amphipathic DNA polymers exhibit antiviral activity against systemic Murine Cytomegalovirus infection
- Rhonda D Cardin1Email author,
- Fernando J Bravo1,
- Andrea P Sewell1,
- James Cummins2,
- Louis Flamand3,
- Jean-Marc Juteau4,
- David I Bernstein1 and
- Andrew Vaillant4Email author
© Cardin et al; licensee BioMed Central Ltd. 2009
- Received: 24 August 2009
- Accepted: 2 December 2009
- Published: 2 December 2009
Phosphorothioated oligonucleotides (PS-ONs) have a sequence-independent, broad spectrum antiviral activity as amphipathic polymers (APs) and exhibit potent in vitro antiviral activity against a broad spectrum of herpesviruses: HSV-1, HSV-2, HCMV, VZV, EBV, and HHV-6A/B, and in vivo activity in a murine microbiocide model of genital HSV-2 infection. The activity of these agents against animal cytomegalovirus (CMV) infections in vitro and in vivo was therefore investigated.
In vitro, a 40 mer degenerate AP (REP 9) inhibited both murine CMV (MCMV) and guinea pig CMV (GPCMV) with an IC50 of 0.045 μM and 0.16 μM, respectively, and a 40 mer poly C AP (REP 9C) inhibited MCMV with an IC50 of 0.05 μM. Addition of REP 9 to plaque assays during the first two hours of infection inhibited 78% of plaque formation whereas addition of REP 9 after 10 hours of infection did not significantly reduce the number of plaques, indicating that REP 9 antiviral activity against MCMV occurs at early times after infection. In a murine model of CMV infection, systemic treatment for 5 days significantly reduced virus replication in the spleens and livers of infected mice compared to saline-treated control mice. REP 9 and REP 9C were administered intraperitoneally for 5 consecutive days at 10 mg/kg, starting 2 days prior to MCMV infection. Splenomegaly was observed in infected mice treated with REP 9 but not in control mice or in REP 9 treated, uninfected mice, consistent with mild CpG-like activity. When REP 9C (which lacks CpG motifs) was compared to REP 9, it exhibited comparable antiviral activity as REP 9 but was not associated with splenomegaly. This suggests that the direct antiviral activity of APs is the predominant therapeutic mechanism in vivo. Moreover, REP 9C, which is acid stable, was effective when administered orally in combination with known permeation enhancers.
These studies indicate that APs exhibit potent, well tolerated antiviral activity against CMV infection in vivo and represent a new class of broad spectrum anti-herpetic agents.
- Antiviral Activity
- Plaque Assay
- MCMV Infection
- Plaque Reduction Assay
- Phosphorothioate Oligonucleotide
Cytomegalovirus (CMV) is a ubiquitous β-herpesvirus that asymptomatically infects immunocompetent individuals but leads to serious illness and mortality in immunocompromised individuals [1, 2]. Currently licensed drugs for the treatment of CMV infection in the United States include Foscarnet, Cidofovir, Ganciclovir and Fomivirisen. These compounds are effective in controlling CMV infection, but emergence of resistance and potentially serious side effects limit their use [2–4]. As such, the need for well-tolerated and potent antiviral compounds with activity against CMV is well recognized.
Amphipathic DNA polymers (APs) are a new class of antiviral compounds based on the sequence-independent activity of phosphorothioated oligonucleotides. The antiviral activity of these compounds against Human Immunodeficiency Virus (HIV-1), Herpes Simplex Virus (and other herpesviridae), arenaviruses, and Hepatitis C Virus (HCV) has been previously demonstrated [5–8]. These compounds inhibit both the entry/fusion and attachment of these viruses [5, 6, 8], and in the case of HIV-1, the amphipathic nature of these compounds was shown to directly mediate their interaction with the core amphipathic α-helices in gp41 [5]. The optimal polymer size (40 mer) for antiviral activity and the specific requirement for phosphorothioation as an enhancer of amphipathic character are well conserved in these unrelated viruses. Moreover, the amphipathic α-helical domains HIV-1 gp41 show a high degree to structural homology to analogous amphipathic domains in the surface glycoproteins of many type 1 fusion viruses [9–15], suggesting that these amphipathic interactions between APs and viral fusion glycoproteins underlie the broad-spectrum antiviral activity of these compounds.
Although the target of AP interaction in viruses other than HIV-1 is not yet known, the structural conservation of amphipathic alpha helical domains in the fusion glycoproteins of most enveloped viruses strongly suggests that the amphipathic domains in surface glycoproteins in herpesviridae are a potential target for AP interaction. Thus, the mechanism of action of these compounds appears unrelated to the published mechanism of action for antisense compounds, such as fomvirisen, which is an antisense phosphorothioate oligonucleotide that blocks CMV-specific protein synthesis by hybridizing with mRNA from the major immediate-early transcriptional unit of the CMV genome [16].
APs have been shown to have broad spectrum in vitro antiviral activity against herpesviridae including HSV types 1 and 2 (HSV-1, HSV-2), Varicella Zoster Virus (VZV), Human Cytomegalovirus (HCMV), Epstein-Barr Virus (EBV), and Human Herpesvirus 6 types A and B (HHV-6 A/B) [6, 17], demonstrating that APs might provide advantages to the currently available therapies to treat herpesvirus infections. Moreover, two amphipathic polymers, REP 9 (a 40 mer degenerate phosphorothioate oligonucleotide) and REP 9C (a 40 mer poly C phosphorothioate oligonucleotide) were shown to be effective topically against HSV-2 infection in a murine model of genital herpes [17], suggesting the potential for antiviral activity in vivo against CMV. Here we report that APs possess anti-CMV activity against two animal CMV strains in vitro and potent, well tolerated prophylactic in vivo efficacy in a murine model of systemic CMV infection via multiple routes of administration and at parenterally administered doses as low as 0.5 mg/kg/day.
Oligonucleotide synthesis
All oligonucleotides for in vitro use were synthesized as described previously [5, 7]. Nucleic acid sequences of oligodeoxynucleotides used in this study are as follows:
REP 9 (phosphorothioate) and REP 2015 (phosphodiester): N40 (random incorporation of A, G, T and C); REP 9C (phosphorothioate) and REP 2110 (phosphodiester): C40; CPG7909: 5' TCGTCGTTTTGTCGTTTTGTCGTT 3' (B-class CPG oligonucleotide, [18]).
Compounds used for in vivo experiments (REP 9, REP 9C) were synthesized under contract with Girindus America Inc. under GMP-like conditions to yield high purity sodium salts.
Nuclease resistance testing
Nuclease resistance of REP 9, REP 9C and their non-phosphorothioated analogs
Nuclease resistance (- = fully degraded, ++++ = fully resistant)
Phosphodiesterase II
S1 Nuclease
Bal 31
Exo I
N40 (degenerate)
Cells and virus
NIH 3T3 cells (ATCC CRL1658) were grown in Dulbecco's modified Eagle's medium (DMEM, Mediatech, Herndon, VA) supplemented with 10% fetal bovine serum (FBS, Hyclone, Thermo-Fisher Scientific), 7.5% Sodium Bicarbonate, 4 mM HEPES, 2 mM L-glutamine, and gentamicin in a humidified 5% CO2 incubator at 37°C. Parent stocks of murine cytomegalovirus (MCMV, strain K181+) were kindly provided by Dr. Edward Mocarski (Emory University School of Medicine, Atlanta Georgia). The K181+ virus stocks were grown in NIH 3T3 cells and the resulting tissue culture-passaged stocks were used in all of the in vitro MCMV studies and the majority of in vivo studies. The Smith strain of MCMV was purchased (ATCC, VR-1399) and a salivary gland-passaged virus stock was used in one in vivo study as described in the text. The salivary gland virus stock was prepared from infected salivary glands at 21 days after infection of 6 week old BALB/c mice with the Smith strain [19]. MCMV (Smith) titers were determined by plaque assay on murine embryonic fibroblasts (MEF) from BALB/c mice. MCMV (K181+) titers were determined by plaque assay on NIH 3T3 cells [20, 21]. Guinea pig CMV (GPCMV, Strain 22122) was purchased (ATCC, VR-682) and was grown in guinea pig lung fibroblast (GPL) cells (ATCC CCL-158). GPL cells were maintained in F-12 media (Invitrogen Corporation, Grand Island, NY) supplemented with 10% FBS (Hyclone, Thermo-Fisher Scientific) and penicillin/streptomycin in a humidified 5% CO2 incubator at 37°C. GPCMV titers were determined by plaque assay on GPL fibroblast monolayers overlaid with media containing 50% Basal Medium Eagle (Sigma-Aldrich Corporation) and 1.5% methylcellulose [22]. All virus stocks were stored at -70°C and re-titered before use in experiments.
Virological assays
For MCMV plaque assays, dilutions of virus stocks or 10% mouse tissue sonicates were adsorbed onto 70% confluent NIH 3T3 monolayers for one hour at 37°C, then overlaid with 1.5% carboxymethyl cellulose (CMC): 2× modified Eagle's medium (1:1) as previously described [20, 21]. After incubation at 5% CO2 for 6 days, the CMC overlay was removed and the monolayers were fixed with methanol and stained with Giemsa to determine the number of plaques. To measure the antiviral activity of the compounds against MCMV and GPCMV, plaque reduction assays were performed. For the plaque reduction assay, 100 plaque-forming units (pfu) of MCMV (K181+) or GPCMV were adsorbed onto NIH 3T3 cells or GPL cells and known amounts of compound ranging from 0.01 μM to 1.0 μM were included in the one hour infection of the NIH 3T3 cells (for MCMV) and GPL cells (for GPCMV) and also included in the overlay media of the plaque assay. The assays were performed in duplicate wells in three separate experiments. The IC50 for each compound was calculated as the concentration of compound which reduced the number of plaques by 50% compared to the untreated control. Cytotoxicity of compounds was assessed by Giemsa staining of uninfected murine fibroblasts or Crystal Violet staining of uninfected guinea pig fibroblasts that were incubated with increasing concentrations of REP 9 compound ranging from 1.0 μM to 100 μM and compared microscopically to control monolayers that were not exposed to compound.
Time-of-Addition assay
To investigate the antiviral activity of REP 9 on MCMV replication when the compound was added at different times during infection, time-of-addition assays were performed using the plaque reduction assay as described above with the following modifications. Briefly, in each well, 10 μM REP 9 or no compound was added with 200 pfu of MCMV (K181+) at the time of infection. After incubation for 2 hours, the virus inoculum was removed and the wells were overlaid with CMC containing 10 μM REP 9 (thus present during entire infection time) or no compound as a control. Additional wells were infected with MCMV for 2 hours or 10 hours in the absence of REP 9, then the virus inoculum was removed, and overlaid with CMC containing 10 μM REP 9, (thus present only at 2 hours or 10 hours after infection). As a control, duplicate sets of wells were set up and cells were washed twice with media to remove excess non-absorbed virus or to remove excess compound. Lastly, similar wells containing 10 μM Ganciclovir (Sigma) added at 2 hours or 10 hours as a control for inhibition of MCMV replication were used. The wells were then incubated at 37°C and 5% CO2 for six days and stained with Giemsa to enumerate the average number of plaques in triplicate wells for each inhibition assay. The time-of-addition studies were performed twice and the % reduction of plaques was determined by comparison to the number of plaques in the control wells.
Mice and infection
Female BALB/c mice were purchased from Jackson Laboratories (Bar Harbor, ME). Three-week old or five-week old mice (8 mice/group) were infected with 1 × 105 pfu of MCMV (K181+) by intraperitoneal (i.p.) inoculation and maintained under specific pathogen-free conditions at Cincinnati Children's Hospital Medical Center. In one study, 5-7 week old mice were infected with 1 × 103 pfu of MCMV (Smith) by i.p., intravenous (i.v.), or subcutaneous (s.c.) inoculation and housed in the animal facility at Laval University, Quebec. At various days post infection (dpi), mice were sacrificed, tissues were collected and 10% tissue sonicates were prepared for virus titration by plaque assay.
In vivo antiviral activity
APs (REP 9, REP 9C), or saline control, were administered to mice by i.p., i.v., or s.c. injection daily at the indicated doses beginning 2 days prior to infection and continued daily for 5 to 6 days as described in the text. For i.p. injection, APs were administered 3 hours prior to infection. On day 0, 3 hours post treatment, the mice were inoculated with virus, and at 3 days post infection (dpi), the mice were sacrificed and the virus titers in the spleen and liver were determined. The spleens were placed in pre-weighed tubes to determine the weight of the spleens at 3 dpi. In some studies, REP 9C was delivered by oral dosing (p.o., 100 μl volume) starting at 2 days prior to infection or in dose response experiments by i.p. injection to 3 week old BALB/c mice starting at 8 days prior to infection. In oral dosing studies, mice were fasted for 3 hours before dosing each day and for 1 hour following dosing. Mice were weighed daily prior to treatment to determine the dose of compound administered to the mice. As a control in some studies, mice were treated daily for 5 consecutive days by i.p. injection of 25 or 50 mg/kg Ganciclovir (GCV, Sigma). In all studies, mice were also monitored for signs of toxicity by weight loss, ruffled fur, and level of activity. For oral REP 9C formulations, the intestinal absorption enhancer n-tetradecyl-β-D-maltopyranoside (TDM, Anatrace, Inc) was prepared at 0.25% in normal saline. The appropriate amount of REP 9C was dissolved in the TDM solution to yield a 400 mg/kg dose per 100 μl solution. A second intestinal absorption enhancer, sodium caprate (C10, Sigma) was formulated in normal saline to yield 100 mg in each oral dose in combination with 400 mg/kg of REP 9C. These solutions were filter sterilized and stored at 4°C and then allowed to come to room temperature prior to administration.
AP stimulation of cytokine production in human PBMC cells
Human peripheral blood mononuclear cells (PBMCs) were isolated from Leukopaks obtained from Biological Specialty Corporation (Colmar, PA) by centrifugation on a Ficoll-Hypaque density gradient. The cells were washed and resuspended in AIM V (InVitrogen; Carlsbad, CA) serum-free medium (containing 50 μg/ml streptomycin sulfate and 10 μg/ml gentamicin sulfate) at a concentration of 4 × 106 cells/ml. For cytokine stimulation, the cells were diluted in an equal volume of AIM V medium (containing 2× final drug concentration) to yield a final concentration of 2 × 106 cells/ml. PBMCs were stimulated in a 48-well format (0.5 ml cells + 0.5 ml drug) for 48 hr with 32 nM of each compound. At study endpoint, cell supernatants were tested in each of the assay platforms to determine the specific cytokines which were stimulated. The following cytokines were measured using kits from Meso Scale Discovery (MSD, Inc., Gaithersburg, MD): Human IFN-γ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-8 IL-10, IL-12p70, IL-13, TNF-α and IFN Inducible Protein 10 (IP-10). These cytokine levels were determined in culture supernatant by using a multiplex platform developed by MSD. Custom-coated MULTI-SPOT plates (7-, 4-, or 1-Spot 96-well format) were used in which the cytokines could be simultaneously detected within each well of the plate. For each, 10 μl of a calibrator (containing all human cytokine standards) or supernatant was added to appropriate wells of the plate and incubated at room temperature for 1-2 hours with vigorous (300-1000 rpm) shaking. This was followed by three washes with PBS-0.05% Tween-20, addition of 20 μl/well of a 1 μg/ml detection antibody solution (containing antibodies to the cytokines in each kit type), and incubation of the plate at room temperature for 1-2 hours with vigorous shaking. After three washes (PBS-0.05% Tween), 150 μl/well of 2× MSD Read Buffer T was added and the plate was analyzed on a Sector 6000 instrument. Cytokine concentrations in each sample were calculated from calibration curves based on four-parameter logistic algorithms and expressed as pg/ml. In addition, a human IFN-α kit from BioSource (Camarillo, CA) kit was used for this study. The high sensitivity protocol (0-500 pg/ml) was used to determine IFN-α levels in culture supernatants. A volume of 100 μl of calibrator or supernatant was added to appropriate wells of the plate and incubated at room temperature for 1-2 hours. The wells were then washed once in the kit wash buffer, followed by addition of 100 μl of antibody, and incubation of the plate at room temperature for 1-2 hours. After three washes in kit wash buffer, 100 μl of horseradish peroxidase reagent was added, and the plate was incubated at room temperature for 1 hour. After four washes in kit wash buffer, 100 μl of TMB substrate was added, and the plate was incubated for 15 minutes at room temperature. To stop the reaction, 100 μl acid stop solution was added, and the well absorbance were determined at 450 nm. Cytokine concentrations in each sample were calculated from calibration curves based on four-parameter logistic algorithms and expressed as pg/mL.
Statistical analysis
All virus titer data shown are expressed as mean virus titer (log10 pfu/ml) +/- standard error. Statistical analysis was performed using the Student's t test (GraphPad Instat Program). P values of < 0.05 (two-tailed) were considered to indicate a significant difference.
APs exhibit in vitro antiviral activity against animal cytomegaloviruses
The strict species specificity of HCMV infection limits the study of HCMV in animal models. To further explore the in vivo antiviral activity of APs against CMV infection, it was first necessary to determine whether REP 9 (a 40 mer degenerate AP) showed antiviral activity in plaque reduction assays against CMV for which small animal models exist. In our assays, REP 9 showed potent activity against both MCMV and GPCMV, with IC50s of 0.045 ± 0.004 μM and 0.16 ± 0.07 μM, respectively, that were comparable to the previously established IC50 of REP 9 against HCMV [17]. In addition to reducing the numbers of plaques observed with increasing concentrations of REP 9, the plaque size also appear to be reduced. A second compound, REP 9C, also showed similar activity against MCMV, with an IC50 of 0.048 ± 0.005 μM. Neither compound demonstrated significant toxicity on fibroblasts, with CC50s > 100 μM. A similar lack of toxicity was reported for REP 9 toxicity on human foreskin fibroblasts (HFFs) [17].
In vivo effect of REP 9 against murine cytomegalovirus
Immunostimulatory activity vs. antiviral activity of APs
In vivo effect of REP 9C against murine cytomegalovirus
Nuclease stability of REP 9 and REP 9C
Both REP 9 and REP 9C exhibited comparable antiviral activity in the liver and, to a lesser degree, in the spleen. Since all phosphorothioate oligonucleotides in general are known to accumulate preferentially in the liver and to a lesser extent in the spleen [29–34], the differential activity of these two compounds in the spleen was most likely not due to differences in pharmacokinetic behavior but rather to differences in compound stability. Since nuclease activity is the major degradative pathway of these compounds in vivo, the nuclease stability of both REP 9 and REP 9C were examined (table 1). Both REP 9 and REP 9C and their non-phosphorothioated counterparts were exposed to a variety of nucleases under optimal degradation conditions (see methods) and then subjected to qualitative analysis by denaturing gel electrophoresis. Under our testing conditions, the control, non-phosphorothioated (phosphodiester) degenerate oligonucleotide (REP 2015) had no detectable stability to any of the nucleases tested as expected. REP 9 was fully resistant to Bal31 and Exo 1 digestion but had only moderate stability to Phosphodiesterase II and was not stable to S1 nuclease activity. The poly C phosphodiester oligonucelotide was completely stable to phosphodiesterase digestion but was only moderately stable to S1 nuclease digestion and displayed no resistance to Bal 31 and Exo 1 activity. REP 9C, however, was completely resistant to all enzymes except for S1 nuclease to which it still displayed a high degree of resistance. Thus, REP 9C has a superior nuclease resistance profile to REP 9 and this may account for the better antiviral activity with this analog.
Antiviral dose response of REP 9C
Antiviral activity of REP 9C following oral dosing
The REP 9C compound has been shown to be highly acid resistant [17], making it well suited for oral administration in the absence of a controlled release formulation. To determine whether REP 9C exhibited antiviral efficacy following oral dosing, an oral formulation of 400 mg/kg of REP 9C in 0.25% (w/v) TDM (tetradecyl-β-D-maltopyranoside) or combined with 100 mg of sodium caprate (C10) was used. TDM belongs to a family of alkylated saccharides known to enhance the intestinal absorption of high molecular weight compounds with poor oral bioavailability [35, 36] and sodium caprate has previously been shown to substantially enhance to the oral bioavailability of phosphorothioate oligonucleotides in human patients [37, 38]. A total volume of 100 μl of either REP 9C + TDM or REP 9C + C10 was delivered orally by gavage with feeding tubes once daily for 5 consecutive days starting at 2 days prior to infection. Control mice were treated with 100 mg C10 in normal saline. Mice were infected i.p. with 1 × 105 pfu MCMV at day 0 and the livers were collected and titered at 3 dpi. As shown in figure 7b, REP 9C delivered orally with 0.25% TDM or 100 mg C10 significantly reduced the MCMV viral loads in the livers. The reduction in viral titers and number of mice with undetectable MCMV in mice treated with REP 9C + TDM or REP 9C + C10 were comparable to the control group which received 20 mg/kg REP 9C via i.p. injection. Oral administration of REP 9C at 400 mg/kg was shown to have no antiviral activity when administered without an intestinal absorption enhancer (data not shown). Mice treated orally with C10 alone or with C10 in combination with REP 9C developed weight loss, inactivity and ruffled fur but none of these complications were observed in mice which received the TDM + REP 9C oral formulation. The animal deaths which occurred in this study (one mouse in the C10 alone group, one mouse in the REP 9C + C10 group, and two mice in the REP 9C + TDM group) were attributed to mechanical trauma from the oral administration procedure and were not considered to be drug related.
The studies of REP 9 and REP 9C presented here demonstrates for the first time the in vivo activity of APs against systemic MCMV infection in two important tissues, the spleen and liver, where CMV replicates and establishes long term persistent infection. APs have been shown to have antiviral activity in vitro against HSV-1, HSV-2, HCMV, VZV, EBV and HHV-6A/B [6, 17], and REP 9 and REP 9C have been previously shown to prevent the topical transmission of HSV-2 [17]. Thus, the in vivo prophylactic activity of REP 9 and REP 9C against MCMV infection in mice suggests that APs may have the potential to be broad spectrum anti-herpetic prophylactic agents in vivo. These initial murine studies presented here are considered an initial proof of concept of the ability of REP 9 and REP 9C to target CMV infection in the liver and spleen.
Traditionally, the activity of phosphorothioate oligonucleotides in rodent models has been difficult to achieve at doses lower than 10 mg/kg/day due to the weaker organ accumulation and much shorter half-life of these compounds compared to primate species [33, 34]. Phosphorothioate oligonucleotides accumulate primarily in the liver, kidney, spleen and lung of mammalian species [29–34] and as phosphorothioate oligonucleotides, APs share these same general characteristics (data not shown) and are thus ideally suited to treat viral infections in these organs. Moreover, at equivalent mg/kg dose, phosphorothioate oligonucleotide accumulation in these target organs is known to be substantially increased in primates compared to rodent species while the half life in these target organs increases from 5 days in rodents to 28 days in primates [33, 34]. In fact, efficacious dose levels of phosphorothioate oligonucleotides of 10 mg/kg/day in rodent species have translated into effective dosing at 200 mg once weekly in humans [39, 40]. This suggests that effective antiviral activity with APs could be achievable with a reasonable dosing regimen in patients.
APs in general have been shown to be entry/attachment inhibitors in HIV-1, HSV-1/2, arenaviruses, and more recently, HCV [5–8]. The inhibition of viral entry/attachment by APs appears to be well conserved in enveloped viruses from divergent families. Similarly, the results from figure 1 indicates that the majority of the REP 9 activity against MCMV appears to occur at early times after infection and thus most likely affects binding and entry as shown for HSV-2 [6]. Some REP 9 activity is also observed when drug is added as late as 10 hours after infection. Thus it seems reasonable to assume that mechanism (s) similar to that reported for REP 9 against HSV-2 are responsible at least in part for the antiviral activity of APs against MCMV infection.
APs have been shown to directly interact with amphipathic alpha helical core fusion domains in HIV-1 [5] which are structurally analogous to the amphipathic alpha-helical structures which are present in gB of HSV [11] and the gH of all herpesviruses [41]. Moreover, these amphipathic domains in gH can be complemented by the core amphipathic alpha-helical domain from HIV gp41 [10]. This heterologous complementation demonstrates the presence of amphipathic alpha-helical structures on the HSV virion surface (as well as all herpesviruses including CMV) with comparable functionality to similar structures in gp41 of HIV-1. Thus, gB and gH may contain domains amenable to AP interaction whose inhibition would be consistent with the entry and attachment inhibition observed with these compounds in HSV-1/2 [6] and with the antiviral activity consistent with entry inhibition in CMV. While the target of AP interaction in CMV has not yet been defined, these observations strongly suggest that interactions of APs with gB and or gH in CMV may be responsible for the inhibition of CMV viral entry.
In murine models, treatment with CpG ODN reduces the severity and time course of infection and facilitates the clearance of viruses such as HSV type 2, Friend retrovirus and influenza [42–44]. Since several studies have shown the involvement of Toll-like receptors (and specifically TLR-9) in the progression of CMV infection [45–48], it is also possible that CpG mediated immunomodulation might be another mechanism whereby REP 9 could indirecty affect viral replication via cytokine stimulation. However, the activity of REP 9C (which is devoid of CpG motifs) in this model system argues that the efficacy of APs in vivo against CMV infection is mainly derived from the direct antiviral activity of APs. Interestingly, REP 9C still maintained residual IL-1β and TNF-α induction in human PBMCs which have pro-viral [49] or antiviral activity [50], respectively. While this cytokine induction in human PBMCs in vitro cannot be directly translated into potential immunomodulatory activity in murine models in vivo, it is possible that some form of cytokine induction may influence the overall antiviral activity of REP 9C in vivo. Splenomegaly was also observed in these studies but only with REP 9 and only in MCMV-infected animals treated with REP 9. It is possible that splenomegaly induced by REP 9 only during MCMV infection could be due to cytokine induction. Further studies are needed to address the underlying mechanism of REP 9-induced splenomegaly.
Phosphorothioate oligonucelotides are known to have limited stability to acid hydrolysis below pH 5, and even when delivered interduodenally to bypass gastric acidity, require intestinal absorption enhancers to achieve systemic exposure [37]. REP 9C is resistant to acid hydrolysis for 24 h at pH1 [17]. This acid resistance is due to the i-plex DNA structure adopted by polypyrimidine sequences [51]. This i-plex structure may also be a factor in the enhanced nuclease stability of REP 9C. The acid stability of REP 9C is predicted to overcome the need for protection from gastric degradation, one of the hurdles in the development of an oral formulation with phosphorothioate oligonucleotides. This is supported by the antiviral activity of an oral formulation of REP 9C with either of two different intestinal absorption enhancers in the absence of any protection from acidic degradation in the digestive tract. The activity of 400 mg/kg of orally administered REP 9C was comparable to that of 20 mg/kg of REP 9C administered parenterally, demonstrating that REP 9C can maintain its bioactivity after passing though the digestive tract. Although the calculated minimum oral bioavailability of REP 9C with these formulations was only 5%, it should be stressed that these experiments were performed with crude formulations which were not optimized for efficient intestinal absorption. Moreover, given the long half-life of these compounds in primates and their preferential accumulation in organs where CMV infection and replication occurs, only modest improvements in oral bioavailability may be necessary to have an oral formulation that would be viable as a prophylactic treatment for CMV in the clinic.
The studies reported here have some limitations. We only evaluated prophylactic administration because CMV infections in rodent systems such as the murine model employed in this study only display transient infections of the spleen and liver, making the therapeutic activity of antiviral compounds difficult to assess. While the use of immunocompromised murine models of CMV infection are also considered useful for evaluating the efficacy of antiviral compounds against CMV infection [52], the efficacy of REP 9 or REP 9C was not further assessed in these models. Investigating the therapeutic activity of APs against CMV infection is the next important step in assessing the clinical potential of these compounds against established CMV infection.
There is a need for antivirals which are effective against multiple human herpesviruses that could be used prophylactically for transplant and severely immunocompromised patients. APs have been shown to have antiviral activity in vitro against HSV-1, HSV-2, HCMV, VZV, EBV, and HHV-6A/6B [6, 17]. Importantly, REP 9 and REP 9C appear to be well tolerated and show in vivo activity against HSV-2 [17]. The experiments reported here extend these observations to a systemic infection with another herpesvirus that is an important pathogen in the immunocompromised. Thus, REP 9 and REP 9C may have the potential to be broad-spectrum anti-herpetic prophylactic agents.
These studies indicate that APs exhibit potent, well tolerated antiviral activity against CMV infection in vivo and represent a new class of anti-herpetic agents.
As a well-tolerated prophylactic agent, APs may be useful to prevent CMV reinfections or reactivations in CMV-positive liver or lung transplant recipients or to prevent primary infection in CMV negative transplant recipients. These compounds may also be useful when combined with currently available drugs for the treatment of CMV. Moreover, since APs are equally effective in vitro against acyclovir and foscarnet resistant herpesviridae [17], these compounds may be useful to treat drug-resistant CMV strains in vivo.
The authors wish to thank Janelle Allen for technical assistance in performing plaque assays and Nadine Bertholet for technical assistance in performing the nuclease resistance assays. This work was supported by REPLICor Inc. and contract NO1-AI-15438 from the Virology Branch, NIAID, NIH.
Authors' Affiliations
- Pass RF: Cytomegalovirus infection. Pediatr Rev 2002, 23: 163-170. 10.1542/pir.23-5-163View ArticlePubMedGoogle Scholar
- Biron KK: Antiviral drugs for cytomegalovirus diseases. Antiviral Res 2006, 71: 154-163. 10.1016/j.antiviral.2006.05.002View ArticlePubMedGoogle Scholar
- Boivin G, Goyette N, Gilbert C, Humar A, Covington E: Clinical impact of ganciclovir-resistant cytomegalovirus infections in solid organ transplant patients. Transpl Infect Dis 2005, 7: 166-170. 10.1111/j.1399-3062.2005.00112.xView ArticlePubMedGoogle Scholar
- Gilbert C, Boivin G: Human cytomegalovirus resistance to antiviral drugs. Antimicrob Agents Chemother 2005, 49: 873-883. 10.1128/AAC.49.3.873-883.2005PubMed CentralView ArticlePubMedGoogle Scholar
- Vaillant A, Juteau JM, Lu H, Liu S, Lackman-Smith C, Ptak R, Jiang S: Phosphorothioate oligonucleotides inhibit human immunodeficiency virus type 1 fusion by blocking gp41 core formation. Antimicrob Agents Chemother 2006, 50: 1393-1401. 10.1128/AAC.50.4.1393-1401.2006PubMed CentralView ArticlePubMedGoogle Scholar
- Guzman EM, Cheshenko N, Shende V, Keller MJ, Goyette N, Juteau JM, Boivin G, Vaillant A, Herold BC: Amphipathic DNA polymers are candidate vaginal microbicides and block herpes simplex virus binding, entry and viral gene expression. Antivir Ther 2007, 12: 1147-1156.PubMedGoogle Scholar
- Lee AM, Rojek JM, Gundersen A, Stroher U, Juteau JM, Vaillant A, Kunz S: Inhibition of cellular entry of lymphocytic choriomeningitis virus by amphipathic DNA polymers. Virology 2008, 372: 107-117. 10.1016/j.virol.2007.10.016PubMed CentralView ArticlePubMedGoogle Scholar
- Matsumura T, Hu Z, Kato T, Dreux M, Zhang YY, Imamura M, Hiraga N, Juteau JM, Cosset FL, Chayama K, et al.: Amphipathic DNA polymers inhibit hepatitis C virus infection by blocking viral entry. Gastroenterology 2009, 137: 673-681. 10.1053/j.gastro.2009.04.048PubMed CentralView ArticlePubMedGoogle Scholar
- Weissenhorn W, Dessen A, Harrison SC, Skehel JJ, Wiley DC: Atomic structure of the ectodomain from HIV-1 gp41. Nature 1997, 387: 426-430. 10.1038/387426a0View ArticlePubMedGoogle Scholar
- Gianni T, Martelli PL, Casadio R, Campadelli-Fiume G: The ectodomain of herpes simplex virus glycoprotein H contains a membrane alpha-helix with attributes of an internal fusion peptide, positionally conserved in the herpesviridae family. J Virol 2005, 79: 2931-2940. 10.1128/JVI.79.5.2931-2940.2005PubMed CentralView ArticlePubMedGoogle Scholar
- Heldwein EE, Lou H, Bender FC, Cohen GH, Eisenberg RJ, Harrison SC: Crystal structure of glycoprotein B from herpes simplex virus 1. Science 2006, 313: 217-220. 10.1126/science.1126548View ArticlePubMedGoogle Scholar
- Eschli B, Quirin K, Wepf A, Weber J, Zinkernagel R, Hengartner H: Identification of an N-terminal trimeric coiled-coil core within arenavirus glycoprotein 2 permits assignment to class I viral fusion proteins. J Virol 2006, 80: 5897-5907. 10.1128/JVI.00008-06PubMed CentralView ArticlePubMedGoogle Scholar
- Wilson IA, Skehel JJ, Wiley DC: Structure of the haemagglutinin membrane glycoprotein of influenza virus at 3 A resolution. Nature 1981, 289: 366-373. 10.1038/289366a0View ArticlePubMedGoogle Scholar
- Zhao X, Singh M, Malashkevich VN, Kim PS: Structural characterization of the human respiratory syncytial virus fusion protein core. Proc Natl Acad Sci USA 2000, 97: 14172-14177. 10.1073/pnas.260499197PubMed CentralView ArticlePubMedGoogle Scholar
- Dimitrov DS: Virus entry: molecular mechanisms and biomedical applications. Nat Rev Microbiol 2004, 2: 109-122. 10.1038/nrmicro817View ArticlePubMedGoogle Scholar
- de Smet MD, Meenken CJ, Horn GJ: Fomivirsen - a phosphorothioate oligonucleotide for the treatment of CMV retinitis. Ocul Immunol Inflamm 1999, 7: 189-198. 10.1076/ocii.184.108.40.20607View ArticlePubMedGoogle Scholar
- Bernstein DI, Goyette N, Cardin R, Kern ER, Boivin G, Ireland J, Juteau JM, Vaillant A: Amphipathic DNA polymers exhibit antiherpetic activity in vitro and in vivo. Antimicrob Agents Chemother 2008, 52: 2727-2733. 10.1128/AAC.00279-08PubMed CentralView ArticlePubMedGoogle Scholar
- Krieg AM, Efler SM, Wittpoth M, Al Adhami MJ, Davis HL: Induction of systemic TH1-like innate immunity in normal volunteers following subcutaneous but not intravenous administration of CPG a synthetic B-class CpG oligodeoxynucleotide TLR9 agonist. J Immunother 7909, 27: 460-471. 10.1097/00002371-200411000-00006View ArticleGoogle Scholar
- MacDonald MR, Li XY, Stenberg RM, Campbell AE, Virgin HWt: Mucosal and parenteral vaccination against acute and latent murine cytomegalovirus (MCMV) infection by using an attenuated MCMV mutant. J Virol 1998, 72: 442-451.PubMed CentralPubMedGoogle Scholar
- Cardin RD, Brooks JW, Sarawar SR, Doherty PC: Progressive loss of CD8+ T cell-mediated control of a gamma-herpesvirus in the absence of CD4+ T cells. J Exp Med 1996, 184: 863-871. 10.1084/jem.184.3.863View ArticlePubMedGoogle Scholar
- Cardin RD, Schaefer GC, Allen JR, Davis-Poynter NJ, Farrell HE: The M33 chemokine receptor homolog of murine cytomegalovirus exhibits a differential tissue-specific role during in vivo replication and latency. J Virol 2009, 83: 7590-7601. 10.1128/JVI.00386-09PubMed CentralView ArticlePubMedGoogle Scholar
- Bravo FJ, Cardin RD, Bernstein DI: Effect of maternal treatment with cyclic HPMPC in the guinea pig model of congenital cytomegalovirus infection. J Infect Dis 2006, 193: 591-597. 10.1086/499603View ArticlePubMedGoogle Scholar
- Wagner M, Michel D, Schaarschmidt P, Vaida B, Jonjic S, Messerle M, Mertens T, Koszinowski U: Comparison between human cytomegalovirus pUL97 and murine cytomegalovirus (MCMV) pM97 expressed by MCMV and vaccinia virus: pM97 does not confer ganciclovir sensitivity. J Virol 2000, 74: 10729-10736. 10.1128/JVI.74.22.10729-10736.2000PubMed CentralView ArticlePubMedGoogle Scholar
- Bale JF Jr, O'Neil ME: Detection of murine cytomegalovirus DNA in circulating leukocytes harvested during acute infection of mice. J Virol 1989, 63: 2667-2673.PubMed CentralPubMedGoogle Scholar
- Cardin RD, Boname JM, Abenes GB, Jennings SA, Mocarski ES: Reactivation of murine cytomegalovirus from latency. In Multidisciplinary Approaches to Understanding Cytomegalovirus Disease. Edited by: Plotkin SA, Michelson S. Amsterdam: Elsevier; 1993:101-110.Google Scholar
- Collins TM, Quirk MR, Jordan MC: Biphasic viremia and viral gene expression in leukocytes during acute cytomegalovirus infection of mice. J Virol 1994, 68: 6305-6311.PubMed CentralPubMedGoogle Scholar
- Hudson JB: The murine cytomegalovirus as a model for the study of viral pathogenesis and persistent infections. Arch Virol 1979, 62: 1-29. 10.1007/BF01314900View ArticlePubMedGoogle Scholar
- Stoddart CA, Cardin RD, Boname JM, Manning WC, Abenes GB, Mocarski ES: Peripheral blood mononuclear phagocytes mediate dissemination of murine cytomegalovirus. J Virol 1994, 68: 6243-6253.PubMed CentralPubMedGoogle Scholar
- Agrawal S, Temsamani J, Tang JY: Pharmacokinetics, biodistribution, and stability of oligodeoxynucleotide phosphorothioates in mice. Proc Natl Acad Sci USA 1991, 88: 7595-7599. 10.1073/pnas.88.17.7595PubMed CentralView ArticlePubMedGoogle Scholar
- Saijo Y, Perlaky L, Wang H, Busch H: Pharmacokinetics, tissue distribution, and stability of antisense oligodeoxynucleotide phosphorothioate ISIS 3466 in mice. Oncol Res 1994, 6: 243-249.PubMedGoogle Scholar
- Geary RS, Leeds JM, Henry SP, Monteith DK, Levin AA: Antisense oligonucleotide inhibitors for the treatment of cancer: 1. Pharmacokinetic properties of phosphorothioate oligodeoxynucleotides. Anticancer Drug Des 1997, 12: 383-393.PubMedGoogle Scholar
- Phillips JA, Craig SJ, Bayley D, Christian RA, Geary R, Nicklin PL: Pharmacokinetics, metabolism, and elimination of a 20-mer phosphorothioate oligodeoxynucleotide (CGP 69846A) after intravenous and subcutaneous administration. Biochem Pharmacol 1997, 54: 657-668. 10.1016/S0006-2952(97)00190-1View ArticlePubMedGoogle Scholar
- Yu RZ, Geary RS, Leeds JM, Watanabe T, Moore M, Fitchett J, Matson J, Burckin T, Templin MV, Levin AA: Comparison of pharmacokinetics and tissue disposition of an antisense phosphorothioate oligonucleotide targeting human Ha-ras mRNA in mouse and monkey. J Pharm Sci 2001, 90: 182-193. 10.1002/1520-6017(200102)90:2<182::AID-JPS9>3.0.CO;2-FView ArticlePubMedGoogle Scholar
- Yu RZ, Kim TW, Hong A, Watanabe TA, Gaus HJ, Geary RS: Cross-species pharmacokinetic comparison from mouse to man of a second-generation antisense oligonucleotide, ISIS 30 targeting human apolipoprotein B-100. Drug Metab Dispos 1012, 35: 460-468. 10.1124/dmd.106.012401View ArticleGoogle Scholar
- Yang T, Arnold JJ, Ahsan F: Tetradecylmaltoside (TDM) enhances in vitro and in vivo intestinal absorption of enoxaparin, a low molecular weight heparin. J Drug Target 2005, 13: 29-38. 10.1080/10611860400020191View ArticlePubMedGoogle Scholar
- Maggio ET: Intravail: highly effective intranasal delivery of peptide and protein drugs. Expert Opin Drug Deliv 2006, 3: 529-539. 10.1517/17425247.3.4.529View ArticlePubMedGoogle Scholar
- Raoof AA, Chiu P, Ramtoola Z, Cumming IK, Teng C, Weinbach SP, Hardee GE, Levin AA, Geary RS: Oral bioavailability and multiple dose tolerability of an antisense oligonucleotide tablet formulated with sodium caprate. J Pharm Sci 2004, 93: 1431-1439. 10.1002/jps.20051View ArticlePubMedGoogle Scholar
- Tillman LG, Geary RS, Hardee GE: Oral delivery of antisense oligonucleotides in man. J Pharm Sci 2008, 97: 225-236. 10.1002/jps.21084View ArticlePubMedGoogle Scholar
- Crooke RM, Graham MJ, Lemonidis KM, Whipple CP, Koo S, Perera RJ: An apolipoprotein B antisense oligonucleotide lowers LDL cholesterol in hyperlipidemic mice without causing hepatic steatosis. J Lipid Res 2005, 46: 872-884. 10.1194/jlr.M400492-JLR200View ArticlePubMedGoogle Scholar
- Kastelein JJ, Wedel MK, Baker BF, Su J, Bradley JD, Yu RZ, Chuang E, Graham MJ, Crooke RM: Potent reduction of apolipoprotein B and low-density lipoprotein cholesterol by short-term administration of an antisense inhibitor of apolipoprotein B. Circulation 2006, 114: 1729-1735. 10.1161/CIRCULATIONAHA.105.606442View ArticlePubMedGoogle Scholar
- Gianni T, Menotti L, Campadelli-Fiume G: A heptad repeat in herpes simplex virus 1 gH, located downstream of the alpha-helix with attributes of a fusion peptide, is critical for virus entry and fusion. J Virol 2005, 79: 7042-7049. 10.1128/JVI.79.11.7042-7049.2005PubMed CentralView ArticlePubMedGoogle Scholar
- Olbrich AR, Schimmer S, Heeg K, Schepers K, Schumacher TN, Dittmer U: Effective postexposure treatment of retrovirus-induced disease with immunostimulatory DNA containing CpG motifs. J Virol 2002, 76: 11397-11404. 10.1128/JVI.76.22.11397-11404.2002PubMed CentralView ArticlePubMedGoogle Scholar
- Dong L, Mori I, Hossain MJ, Liu B, Kimura Y: An immunostimulatory oligodeoxynucleotide containing a cytidine-guanosine motif protects senescence-accelerated mice from lethal influenza virus by augmenting the T helper type 1 response. J Gen Virol 2003, 84: 1623-1628. 10.1099/vir.0.19029-0View ArticlePubMedGoogle Scholar
- Harandi AM, Eriksson K, Holmgren J: A protective role of locally administered immunostimulatory CpG oligodeoxynucleotide in a mouse model of genital herpes infection. J Virol 2003, 77: 953-962. 10.1128/JVI.77.2.953-962.2003PubMed CentralView ArticlePubMedGoogle Scholar
- Compton T, Kurt-Jones EA, Boehme KW, Belko J, Latz E, Golenbock DT, Finberg RW: Human cytomegalovirus activates inflammatory cytokine responses via CD14 and Toll-like receptor 2. J Virol 2003, 77: 4588-4596. 10.1128/JVI.77.8.4588-4596.2003PubMed CentralView ArticlePubMedGoogle Scholar
- Krug A, French AR, Barchet W, Fischer JA, Dzionek A, Pingel JT, Orihuela MM, Akira S, Yokoyama WM, Colonna M: TLR9-dependent recognition of MCMV by IPC and DC generates coordinated cytokine responses that activate antiviral NK cell function. Immunity 2004, 21: 107-119. 10.1016/j.immuni.2004.06.007View ArticlePubMedGoogle Scholar
- Tabeta K, Georgel P, Janssen E, Du X, Hoebe K, Crozat K, Mudd S, Shamel L, Sovath S, Goode J, et al.: Toll-like receptors 9 and 3 as essential components of innate immune defense against mouse cytomegalovirus infection. Proc Natl Acad Sci USA 2004, 101: 3516-3521. 10.1073/pnas.0400525101PubMed CentralView ArticlePubMedGoogle Scholar
- Szomolanyi-Tsuda E, Liang X, Welsh RM, Kurt-Jones EA, Finberg RW: Role for TLR2 in NK cell-mediated control of murine cytomegalovirus in vivo. J Virol 2006, 80: 4286-4291. 10.1128/JVI.80.9.4286-4291.2006PubMed CentralView ArticlePubMedGoogle Scholar
- Tian P, Shen X, Feng H, Gao B: IL-1B Attenuates IFN-aB-induced antiviral activity and STAT1 activation in the liver: involvement of proteasome-dependent pathway. J Immunol 2000, 165: 3959-3965.View ArticlePubMedGoogle Scholar
- Anderson KP, Lie YS, Low MA, Fennie EH: Effects of tumor necrosis factor-alpha treatment on mortality in murine cytomegalovirus-infected mice. Antiviral Res 1993, 21: 343-355. 10.1016/0166-3542(93)90012-8View ArticlePubMedGoogle Scholar
- Kanehara H, Mizuguchi M, Tajima K, Kanaori K, Makino K: Spectroscopic evidence for the formation of four-stranded solution structure of oligodeoxycytidine phosphorothioate. Biochemistry 1997, 36: 1790-1797. 10.1021/bi961528cView ArticlePubMedGoogle Scholar
- Kern ER: Pivotal role of animal models in the development of new therapies for cytomegalovirus infections. Antiviral Res 2006, 71: 164-171. 10.1016/j.antiviral.2006.05.018View ArticlePubMedGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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Producer/Host: Dave Evans
Studio Engineer: Matt Murphy
Topic: Conservation & Footprint
What are the most-conserving changes that people can make? What information sources can help with evaluating our impacts and changes? What is the carbon footprint of computer use?
Guest: Jim Merkel, Author of "Radical Symplicity"
Call in show
Standard Podcast [ 58:15 ] Play Now | Play in Popup | Download (113)


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After viewing her friend's engagement photos on Facebook, Sarah decided to contact us about her wedding photography and we are so glad she did. Sarah said she found it hard to initially start researching photographers because "everyone with a Canon EOS Rebel thinks they're the next Ansel Adams, so there's no shortage of portfolios to review." But at the same time it was easy "because you could really see who had an eye for certain things" while viewing their work.
Sarah and her fiancé Ryan said of their initial meeting with us: "we felt we all just "clicked" as people which was huge." As we tell all the couples we meet with, "your wedding photographer is with you basically every second at your wedding, so you want to be comfortable and confident around who you choose." Ryan added that "the engagement shoot was a great preview of what we could expect at the wedding. After that, we were sure that everything was going to be fine."
Ryan and Sarah's engagement was 14 months which they felt was the perfect amount of time to find a venue and get everything in order. They were even able to personalize their wedding too. Ryan says: "The theme going into it, if it could be called a theme, was "simple but elegant." That pretty accurately describes us, I think. Nice, but not in an in-your-face kind of way. There was the general elegance of it, as well as LEGO table numbers that we built. I don't think you should have to give up on what you were into as a kid, just because you're an adult, so I routinely buy and build LEGO models as a stress reliever. Sarah got into it as well, and it's something we both do together after work sometimes. Our friends know that about us, and we had to incorporate it into the wedding somehow." <– those LEGO table numbers were really cool!
October 21st was a GORGEOUS day that didn't feel like fall at all. The weather was 74 and sunny – perfect for photos at Indian Pond Country Club. Ryan and Sarah had scouted out all the locations where they wanted to do their photos at – which was great. And there were sooo many options!
Paul and I coordinated their first look out on the deck overlooking the golf course. I would say it was pretty epic actually. 2017 has been the year of emotional first looks and Sarah and Ryan's was no different. I believe when Ryan saw Sarah he said, "wow!" In his words: "The first look will always be one of my happiest memories. I remember standing there, looking out at the golf course, talking to Paul about how excited I was. I never get excited, certainly never like I did as a kid. In that moment, it was like the anticipation of every birthday and Christmas and summer vacation all rolled into one and amplified. I was so sure that I wouldn't cry, like maybe I could will myself not to. Partially because I was going to be photographed, but mostly because I didn't want to be cliché. I was incorrect. I cried. She was too pretty not to."
After the first look, the venue drove the bridal party {and us} to this really nice stone bridge by one of the holes on the golf course so we could do most of the formal photos. The lighting was perfect, the scenery was gorgeous and we didn't get hit by a golf ball once!
They had an unplugged wedding ceremony that was performed by their good friend Buddy. The ceremony was personalized to the couple so guests got a good vibe of who Ryan and Sarah were. At the end of their ceremony and kiss, we caught the infamous "elbow" photo that they wanted. The meaning behind the elbow notion occurred one night when the couple pondered by "I love you" had to be followed by a kiss? Why not an elbow touch? Kind of genius don't you think?
Immediately following the ceremony, we took a few more photos with Ryan and Sarah before they joined their guests in cocktail hour.
Sarah says her favorite moment was: "There are so many its hard to pick a favorite. Since Ryan went with the first look I will chose our first dance. We realized about 6 weeks before our wedding, we needed dance lessons so we didn't look like we belonged at a middle school dance just standing in one spot swaying back and forth. So we went on a crash course in ballroom dancing and had a planned set of moves we practiced over and over again for the 2 minutes that everyone would be watching us. We started out just as choreographed but then I got caught up in the moment and we messed up a move. I tried to cover it up with an extra twirl and don't think anyone but us knew it was a "mistake". I felt like we were the only two in the room and felt so special and loved in that moment."
After the formalities of their reception, DJ Mike Amado from Entertainment Specialists got EVERYONE on the dance floor for the rest of the night. There were some songs where I wanted to put my camera down and dance! Here are some more fun reception photos from their wedding night – including a photo of my with Bryan and Erica who originally referred us.
Venue & Food: Indian Pond Country Club
Day of Wedding Planner: Jill Hall of Mosaic Events
Photographer: Nicole Gesmondi Photographer, LLC
Cake / Cupcakes: Veronica's Sweet Cakes
Florals: Divine Orchids
Entertainment: Mike Amado of Entertainment Specialists
Officiant: Buddy Hutton {friend}
Uplighting / Photo Booth: Mike Amado of Entertainment Specialists
Transportation: ROCKSTAR LIMO
Invitations: The Write Approach
Bridal Gown / Veil: purchased from Mariella Creations
Groom's Attire: Customized Tux from Alton Lane
Honeymoon: Sandlas Resort in St. Lucia. Coordinated by Jayne from My Honeymoon Planner
Nicole Gesmondi Photographer, LLC is a wife-and-husband photography team located in Providence, Rhode Island. We've been providing wedding and portrait photography services in and around Providence and Newport, Rhode Island, New York, Connecticut, Boston and more for over a decade.
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Joe Again? Say It Ain't So.
HEMPSTEAD, N.Y., Oct. 15
Before Wednesday night's final presidential debate, the big question had been whether John McCain would hit Barack Obama with Bill Ayers. Instead, he hit him with Joe the Plumber.
Ayers, the "washed-up terrorist," as McCain called him, had but a bit role; the Republican nominee instead focused on a plumber from Toledo who fears that Obama will make him pay higher taxes.
"Senator Obama was out in Ohio and he had an encounter with a guy who's a plumber. His name is Joe Wurzelbacher," the Republican nominee said at the start of the debate. Turning to Obama, McCain leveled a severe accusation: "What you want to do to Joe the Plumber and millions more like him is have their taxes increased and not be able to realize the American dream of owning their own business."
"Is that what you want to do?" the moderator, CBS's Bob Schieffer, asked Obama.
"That's what Joe believes," McCain maintained.
Obama, the front-runner, suddenly found himself on the defensive on the Joe issue. He sought to clarify "the conversation I had with Joe the Plumber."
McCain would not yield. "Small-business people like Joe the Plumber are going to create jobs unless you take that money from him and spread the wealth around," he said.
Obama, still embattled on the Joe front, acknowledged that "my friend and supporter, Warren Buffett, for example, could afford to pay a little more in taxes."
"We're talking about Joe the Plumber!" McCain interjected.
An hour later, they were still turning Joe into a real-life version of "Swing Vote," the Kevin Costner film in which one man single-handedly decides the presidency.
The Joe maneuver was emblematic of McCain's tactics in the final debate: He answered critics' demands that he go after Obama, but he did it with apparent ambivalence.


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Stafford Road Club kit is ordered usually twice a year for delivery in Spring/ summer season and autumn / winter. Club tops (polo and sweat shirts) are available all year round.
An email will be sent out to all members with up to date details of club kit available and asking for your specific requirements. Due to the cost of clothing and the variety of items and sizes available, the club does not hold kit in stock and the orders placed are to meet specific request from members. So please plan ahead to ensure you have the appropriate items of clothing you may require for the next 6 months.
Details of club kit and latest prices are shown below (prices are subject to change and you will be notified of the current prices before orders are placed).
The new and exclusive Stafford Road Club kit design is shown below


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KSA Landbridge to link the Red Sea with Arabian Gulf
It will be the largest BOT rail project ever undertaken in the region, cutting across the peninsula
The Saudi Railways Organisation (SRO) has finalised details of the Saudi Landbridge project, which proposes for the first time to link by rail the eastern and western coasts of the Gulf Cooperation Council's largest member state. SRO is now looking for prospective investors and partners and is inviting private sector participation in the infrastructure project on the basis of a Build-Operate-Transfer (BOT) concession. The Saudi institution held a 'Project Day' in London earlier this month to brief investors and other potential private sector participants on the Saudi Landbridge project. Information was provided to shipping lines, port and rail operators, equipment suppliers and other potential bidders. Though no value has yet been attached to the project, top Saudi officials said that Landbridge will be one of the largest BOT projects ever undertaken in the Middle East. "It is a key initiative in the Railway Expansion Programme approved by the kingdom's Supreme Economic Council," said Dr Jobarah Al Suraisry, Saudi Minister of Transport and SRO chairman. "The project is part of the kingdom's privatisation strategy, the objectives of which include generating economic benefits through effective private sector participation in the national economy, creating employment opportunities and rationalising public expenditure," Dr Al Suraisry added. He pointed out that due to the complexity and multi-disciplinary aspects of the project, SRO anticipates that interested parties may wish to form consortia to bid for the project. Landbridge is the second large rail project within the GCC that has come off the drawing boards; Dubai is already well advanced on its multibillion-dollar Metro project, with the final submission of bids scheduled for 28th February. However, the scope of the Saudi project is much larger. "The Landbridge project will create a new dimension in land transport across the Saudi Arabian Peninsula, transforming the existing rail network into a world-class freight and passenger rail system," said the kingdom's Transport Minister. The Landbridge rail network will link Saudi Arabia's three largest ports: the Jeddah Islamic Port in the west through to Riyadh's Dry Port, and on to Dammam's King Abdul Aziz Port in the east. "The interconnected railway tracks will allow large quantities of cargo to be transported across the country at competitive rates, resulting in considerable cost savings in the movement of goods to and from North America/Europe and the Gulf," the minister said. "The east-west rail link will also facilitate the development of a safe, comfortable and fast overland passenger transport service." The Landbridge project will involve the construction of a new 950 km railway line between Riyadh — the capital city of Saudi Arabia — and the Red Sea port of Jeddah. Additionally, it will create a new 115 km railway line between the industrial city of Jubail in the Eastern Province and the Gulf port of Dammam. The project also calls for the upgrade of the existing rail link between Riyadh and Dammam and the integration of the new lines with Jeddah Islamic Port, King Abdul Aziz Port in Dammam and Riyadh Dry Port. The minister said that the successful BOT bidder will design, finance, build and operate the Saudi Landbridge for a fixed period of time. The obligations of the selected consortia will include designing and constructing the rail network to meet the SRO specified criteria, and subsequently maintaining the network at required levels of operational efficiency. Project financing is expected to comprise investor equity and debt from the Saudi Arabian and international financial markets. The Kingdom of Saudi Arabia will provide the land required for the project and core assets of SRO will be transferred to the BOT concessionaire. UBS Investment Bank, Saudi Arabia's National Commercial Bank and SNCF International of France, are providing financial and technical advisory services to the SRO for the project. Linklaters and the law office of Abdulaziz H Fahad, are providing legal advice. SRO president Khalid H. Al Yahya said that prospective bidders should carefully consider the skills and capabilities required to implement a project of this nature when forming their consortia. "The grand scale and complexity of the project and the exacting operational requirements mean that we would expect consortia to comprise an appropriate group of different disciplines," he pointed out. A pre-qualification document describing the project in more detail will be published in two months' time, he added. The SRO expects to announce the pre-qualification process in the second quarter of 2005. Invitations to bid will be issued shortly thereafter, most likely in the second half of 2005.


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Agriculture and rural community has always played a crucial role not only in conserving the environment, but also in preserving cultural heritage. It is also the core industry as the basis of BT (BioTechnology) and ET(Environment Technology) leading the 21st century.
Competitive agriculture, clean environment, safe agricultural products, and living and breathing rural community with tradition and culture which are our desirable goals. About 400 researchers are working hard to achieve the goals day and night.
We perform extensive studies in various fields such as investigation for change of agricultural environment & soil resources, research for plant nutrition & soil microbe, pests management & security of safe agricultural products, preservation of agricultural ecosystem & culture, mechanization of agriculture & development of technology for energy reduction, research on agricultural bio-technology, modernization & globalization of Korean food, efficiently management of agricultural genetic resource and so on. Through these activities, we make every effort to provide a benefits for the people, consumers who care about and support agriculture, as well as farmers.
Finally, we will do our best to post sufficient information in the areas of agricultural science technology such as agricultural environment, agricultural biology, safety of agricultural products, agricultural engineering, biology resource, globalization of Korean food, and agricultural genetic resource on our homepage.
We will sincerely appreciate your particular attention and support.


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There are so many bad cake recipes that are out there for the holidays that we thought we would treat you to one that everyone loves, rum cake.
Christmas is a festive time and there is no better way to get the festivities rolling than with great Christmas rum cake recipes. This is a recipe that we found years ago in the Internet and have used it every year to everyone's delight.


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Bite-sized Goodness
We're going light and bite sized with the spotlight on the words and tunes themselves.
Sneaking up this September as promised!
Project OMG!: An Evening of Musical & Literary Snacks
Sunday, 23rd September 2007
at The Dram Projects (next to Food Foundry) (map)
Donations at the door (to help keep us alive!)
Edible snacks will be served as well. Yes, the real stuff.
For further enquiries email us at email@example.com or you can contact Patricia (012 7839006) or Priya (016 6606460)
See you all there then?
*Updates! Line up!
Zlikha Batsheba, Catalina Rembuyan, Sheena Baharuddin
with music by Ray Cheong and Rina S.
and featuring a ragtag band with a certain Daphne Lee in it, and a djembe (now known as Mojo Pin)
Sep 16, 2007
Bite-sized Goodness
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