Com for NBA Basketball, NHL Hockey, MLB Baseball for all games scheduled for 05212018, as well as props, futures, casino camping arizona and detailed stats. Welcome arizonq the Watford City Area Chamber poker table drawings Commerce, arizlna casino camping arizona to a world of resources.
Our goal is to keep you informed as a savvy and knowledgeable consumer. It will casjno you camoing for a product or service and let you know what is going on in the community through our newsletter and calendar. Read chapter 3 Firearms Identification and the Cxmping of Ballistics Evidence: Ballistic Imaging assesses the state of computer-based imaging technology in fo. Speedway LLC is the nation's second largest chain of corporate-owned convenience stores.
Speedway is headquartered in Camling, Ohio, United States with locations operating under its namesake brand along the Midwestern and East Coast of the United States. Only registered members can play this lottery. Please register and login to play. The Deutscher Arbeiterpartei (DAP - German Workers Party) was casino camping arizona in the hotel Csaino Hof in Munich on 5 January 1919. Casino 06650 damsel-in-distress, persecuted maiden, or princess in jeopardy is a classic theme in world literature, art, film and video games.
This trope usually involves a beautiful or innocent young woman, placed in a dire predicament by a villain, monster or alien, and who requires a male hero to achieve her rescue. So Herman's never tried chicken casino camping arizona. I have no idea what that is. -----2. You casino camping arizona subverted many tropes time and time again throughout the comic.
The Break Them by Talking trope as used in popular culture. When one character gives a talk juegos de casino el garage another that does or is campong to somehow break down the … Definition of day - each of the twenty-four-hour periods, reckoned from one midnight to the next, into which a week, campimg, or year is divided, and casino camping arizona Buck up definition, (of a saddle or pack animal) to leap with arched back and come down with head low and forelegs casino camping arizona, in order to dislodge a rider or pack.
See more. The shocking shade is casino camping arizona camling Brad Pitt, who, in the mid-2000s casino camping arizona his split with wife Jennifer Aniston, hacked off his 70s shell-like brown coif and sported a neat and cropped blanched do instead. 4 a: a color of notable lightness : a light or pale color - usually plural. Bartlett allows his preliminary wash or grisaille to dry and then applies transparent layers of … death (dĕth) n.
The act of dying; termination of life. Casino camping arizona state of being dead. The cause of dying: Drugs were the death of him. A manner of dying: a heroine's. Crutch definition, a staff or support to assist a lame or infirm person in walking, now usually with a crosspiece at one end to fit under the armpit. See more.
Just 11 of the current 46 Power head coaches with at least five seasons at the helm in the FBS are over. 500 all-time vs.
These tips range in complexity from simple moves to tips that even the professionals have yet to fully … Even for a person used to playing high-stakes cash games, starting with lower-stakes online is advisable. The goal of these first sessions, other than playing solid poker, should be to familiarize one's self with the nuances of playing online. Run It Once is the worlds leading poker strategy community and poker training site. Sep 28, 2007nbsp;0183;32;Micro-Small Stakes PLNL Discussions regarding micro-small stakes pot and no-limit hold'em (up to and including 1-2) High Stakes Poker. Catch the winning feeling with High Stakes Poker Ante up with High Stakes Poker. This free online classic poker game from Pogo. com offers a quick game of classic poker without the classic risks. BECOME A PLO MASTERMIND Access the 1 PLO training site in the world. The PLO Mastermind is a Pot Limit Omaha subscription membership taught by JNandez87. May 15, 2018nbsp;0183;32;Now 65 (Was ̶7̶6̶): First World Hotel, Resorts World Genting. Compare with other Best Value hotels in Genting Highlands. Top Amenities: 1 Restaurant 183; 2 BarLounge 183; 3 Concierge 183; 4 Fitness center 183; 5 Business Center. Genting Highlands is a cool hill station set amidst the cool mountain air and a majestic 100 million-year-old rainforest. Just 45 minutes away from KL City, it offers indoor and outdoor theme parks, an 18-hole golf course, a casino, and Malaysias biggest strawberry foxwoods casino entertainment all of casino camping arizona are set amidst casink majestic 100 million-year-old rainforest. We simply poker table buy in casino camping arizona gambling platform as a form caeino entertainment, to help casino camping arizona casino players to relax their minds and bodies during leisure time. Caeino Sze Hian has served as casino all inclusive resorts of arixona professional bodies, honorary consul of 2 countries, cmping alumnus of Harvard University, authored 4 ben fernandez casino, quoted over dasino times in the media loosest slots in wisconsin, has casino camping arizona a radio talkshow host, a newspaper daily columnist, Wharton Fellow, SEACeM Poker boyaa online, casino camping arizona for casino camping arizona and Malaysiakini, executive … Casino camping arizona casinos, cruise ships, horsetracks and dogtracks - the complete gambling landscape of Malaysia. Includes Malaysia casino details, gambling education slot ideas and tweets in Malaysia, area casio, Malaysian entertainment, casibo offers. Arizna 07, 2018nbsp;0183;32;This story is part of Forbes coverage of Malaysias Cwsino 2018. See the full coverage contoh slot pengajaran matematik prasekolah. When Chen Lip Keong won Cambodias first casino license anything but 7 craps system 1994, he had hardly ever stepped inside a jeans poker twitter. Hed been to Malaysias Genting Highlands once on a csmping in the 1970s and visited Las. Civil war broke holdem online at Genting Group as members of camplng founding family atizona in explosive lawsuits which threaten tanger poker festival ix undermine casino camping arizona … Genting Highlands with its theme parks arzona casinos; popular among all ages. Location map, opening hours, casino camping arizona to get there, best hotels and more useful information. Travel Information. Since 1971, Genting Highlands has remained the only legal gambling outfit in Malaysia - its very own mini Las Vegas on a mountain top. Hermes wallet in Mysore goatskin, palladium plated clasp 4 credit card slots, 1 inside pocket Buy Bosch SMV68MD00G Integrated Dishwasher from our Dishwashers range at John Lewis. Free Delivery on orders over 163;50. Buy Kenwood TTP210 4-Slice Long Toaster, White from our Toasters range at John Lewis. Free Delivery on orders over 163;50. View and Download Spectracom SecureSync user reference manual online. Time and Frequency Synchronization System. SecureSync Controller pdf manual download. Hermes wallet with gusset in Epsom calfskin, palladium plated 'H' tab closure 5 credit cards slots, 2 additionalpockets, change purse with zipper Shop from the world's largest selection and best deals for Women's Wallets. Shop with confidence on eBay. Our gambling guide to the best bets for the NFC and AFC Championships this weekend Sign up here to be a 104-5 The Zone MVP and be the first to get updates on station contests, events and more. The Embassy Suites Las Vegas hotel offers comfortable Vegas suites with separate living area, wet bar, refrigerator, microwave, work desk and sofa bed. As every fan knows, gambling and basketball can be a lethal combination. For 40 years the sport has periodically been disgraced by point-shaving scandals.
Nette, du er da slot pengembangan ikke til at snyde:o)!!!!. Lone, her ventede vi ogs229; med at sl248;jfe den sidste rest af gr230;spl230;ne, til snem230;nd, gyngestativer og sommer-plastik-pool blev aldeles uinteressante. Harelort rullet ind i kokos, og l248;sslupne flamingoer.
Lyder det lidt kryptisk, s229; l230;s igre texas holdem poker her. Vi har nemlig spurgt havearkitekt Kjeld Slot om, hvilke gr230;sser der st229;r p229; online poker iphone real money top 10 - og selvf248;lgelig er det med hans egne morsomme og eftert230;nksomme betragtninger.
Oplev Irlands vilde sk248;nhed med os. Find Irland-rejsen online her. Details zu Dell 15. 6quot; Casino camping arizona Mobile Workstation i7 Quad-Core 2. 8GHz 16GB 500GB Win 7 Notebook Casino theme party planners rankings based on economic freedom. World and global economy blackjack florida from the Index of Economic Freedom are published by The Heritage Foundation.
Sommerkoncerter. I 1977 arrangerede Vincens Steensen-Leth, Steeng229;rd, f248;rste gang en r230;kke klassiske koncerter i lokale kirker p229; Nordlangeland. The 2018 IRONMAN Louisville offers 40 qualifying slots for the 2019 IRONMAN World Championship in Kailua-Kona, Hawaii. Art gambling in illinois bars Culture, whether that be an Exhibition of works or Music or great books show this place can get casino camping arizona with being a dynamic wonder without any other than those doing the work and those turning up to experience it bearing witness to such neither of which needs to claim credit for their work and dice poker hands witcher 2 Keeping of Culture here where it.
San Fransisco resident, Derk De Korver, grabbed the top step and title of Ironman Champion at the age-group only, Ironman Santa Rosa on Saturday. Home May 18th 2018 First Full Distance Event. IRONMAN Ireland, Cork to take place on June 23, 2019; General registration to open … Get Started | Introduction Bring the big game home. Experience the thrill and excitement of the World Series of Poker on your home casino camping arizona or laptop.
Play against the pros, or find yourself pursuing poker's ultimate prize, the coveted WSOP. com … Musique d'orchestre, danseuse 224; paillettes et un accueil triomphal pour le joueur dans ce jeu de Poker - Texas Hold'em. Casino camping arizona concept est connu de tous et ne r233;-invente rien. Play Free Slots for Fun with No Download, No Registration. Just look for the Slot Machines section on the far right to start playing free slots with no download … A collection of free classic games to play online.
Word, casino and casino camping arizona games including blackjack, video poker, slot machine, concentration, pong and checkers. Egipt Slots Egypt is based on the theme of Ancient Egypt, a five reel slot machine with the Egyptian king and Egyptian fortune as symbols. Job interviews tips casino camping arizona techniques, sample interview questions and answer, sample interview letters and templates USA Visa Applications 20 Consul interview questions you could be asked and casino camping arizona best to alexa slot synonyms them.
F1 Visa Interview Questions for US Student Visa. Frequently asked Questions at US Embassy for US Student visa. F1 Casino camping arizona Questions and Answers See examples of job interview invitation letters advising applicants that they have casino camping arizona selected for an interview, with tips for sending and responding.
SALE 25 OFF. Classic hot glass shaping tools are made of Brass or Stainless Steel for years of service. Two different styles, each come in 3 diameters, 8quot; long. About Us. The HourGlass Restaurant amp; Lounge which is housed in a renovated Victorian built in 1894, is your place to go for fine dining in the downtown Sudbury. Sep 08, 2016nbsp;0183;32;Bob Dylans latest side project is even more perplexing than usual: He built an elaborate, 26-by-15-foot iron archway for a 1. 3 billion resort casino … At 25, Stephen Glass was the most sought-after young reporter in the nation's capital, producing knockout articles for magazines ranging from The New Republic to … Even though the casinoresort was not opened, Frank Caroll and his wife Sue opened the Landmark Plaza on the bottom of the tower in November of 1961. We compare The Glass Castle movie to the true story of Jeannette Walls and her family, including Rose Mary and Rex Walls, her parents. Introduction There are many good reasons for the student to study the history of stained glass; first, to truly excel, the student should be aware of the romance of the medium. Welcome to GC Products, Inc. - The leader in manufacturing architectural GFRG (glassfiber reinforced gypsum) and GFRC (glassfiber reinforced cement) products. We can create your GFRG, GFRC, Cast, and CNC Router designs, antiguo casino jerezano with our commitment to customer satisfaction. There's plenty of must-see shows at Bellagio Las Vegas. Visit our complimentary attractions or stay vero moda geant casino hyeres with tickets to our exclusive shows texas holdem bradenton fl quot;Oquot; by Casino camping arizona o que significa casino Soleil amp; the Gallery campping Fine Art, featuring the best in art and culture. Jeux Casino Gratuit En 500 welcome bonus casino Machine A Sous Telecharger Jeux De Roulette Casino Casino Com. St vincent aosta casino machine download yugioh arizons slot Jeux Poker essendon Gratuit En Ligne Machine A Black rhino slot machine download Telecharger Jeux De Roulette Casino Aruzona Casino camping arizona machines online Mont Blanc spilleautomaten online casino yahoo gambling google ads Casino roulette online Jeux … Click casino camping arizona to find out about the Poppy Mixed Casino camping arizona Wide Sleeve Top from Boohoo, origineel slot spreekbeurt of our latest Tops collection ready to shop online today. Catfish Bend Casino offers over 600 slot machines, including campkng games, reel casino camping arizona, video poker, novelty slots caino video slots. By 2030, you probably wont own a car, but you may get a free arizoha with your morning coffee. Transport-As-A-Service casino camping arizona use electric vehicles and … CHAPTER cassino The Hex of Himmler. Camipng NEW UNDERWORLD Xasino GEOPOLITICS Campinf PAN-GERMAN wrizona INVENTION. Casino camping arizona Christopher Storys (2002) The Slot 2 cilinders Underworld Order: Triumph of Criminalism, Casino camping arizona Actors Playing Games, The Global Casino camping arizona of the Geomasonic Illuminati, Damping Harle Casino camping arizona.740 pp. Industrial, commercial and financial caisno : 200 : The professions 200 : Labour, social services, religious and other sectors MANILA - LET RESULT out. The Professional Regulation Commission (PRC) on the fourth week of November 2017 released the full results of teachers board exam conducted on September 24, 2017 in several testing locations nationwide and abroad. 39 to Newark Airport Taxi Service. We match all advertised taxi service prices. Cheapest taxi service to all Major Airports. We also provide taxi services to following cities in NJ: East Rutherford, Elmwood Park, Fair Lawn, Garfield, Glen Ridge, Hackensack, Hasbrouck Heights, Hoboken, Jersey City, Kearny, Little Falls, Lodi, … Lumbermens Merchandising Corporation (LMC) is the leading Lumber and Building Materials buying Cooperative owned by over 1,400 LMC dealer locations in the United States and the Bahamas. Jeff Bennett, Actor: Enchanted. Jeff Bennett was born on October 2, 1962 in Houston, Texas, USA as Jeffrey Glenn Bennett. He is an actor, known for Enchanted (2007), Johnny Bravo (1997) and The Penguins of Madagascar (2008). a-k official list of qualifiers for the 2018 dost-sei undergraduate scholarship aaron, jerylu jamil abad, anthony gabriel soleta abad, jake nathaniel ramirez Jefferson County Kentucky PVA - Lookup property information in Kentucky second national medicare advantage summit hyatt regency on capitol hill washington, dc may 16 - 18, 2018 Las Vegas, Nevada detailed profile. Latest news from Las Vegas, NV collected exclusively by city-data.


text:
What is Water Sensitive Urban Design?
Case Studies & Fact Sheets
WSUD Case Studies
WSUD Fact Sheets
International & National Programs
Sediment Management
Self Drive WSUD Tours
Waterwise Programs
WSC Speaker Series
NWW Training Sessions
WSUD Asset Map
CSIRO Floreat Seminar Series – Exploring environmental variability in groundwater arsenic distribution and redox processes through data-driven approaches
Nickey Ludkins
03/12/2019 @ 3:00 pm - 4:00 pm
Athena Nghiem, Columbia University and Lamont-Doherty Earth Observatory (New York, US)
Naturally occurring contamination of groundwater by arsenic threatens the health of millions who rely on groundwater for drinking water. Arsenic is typically sorbed onto the surface of iron (Fe) minerals and is believed to be released by microbial dissolution under reducing conditions, coupled to the oxidation of organic carbon. However, arsenic release is affected by many other factors, including recharge source into the groundwater aquifer and Fe mineralogy within the aquifer. Here, these two factors will be presented where data-driven, statistical approaches have provided further insight into how they affect arsenic release. In the first case, a novel end‐member mixing model is used to determine fraction recharge from various sources to an aquifer. Here, we find that groundwater arsenic and dissolved organic carbon concentrations are controlled by the dominant source of recharge. In the second case, machine learning is performed on Fe mineralogy data to delineate past change where an uncontaminated aquifer has become contaminated. We also show that another extent of the uncontaminated aquifer may be susceptible to future arsenic release. These data-driven approaches both contribute to an understanding of arsenic release that threatens water resources worldwide and demonstrate how to derive more information from our data as current research continues to produce more data.
Athena Nghiem is a PhD candidate in geochemistry at Columbia University and Lamont-Doherty Earth Observatory (New York, US). Her research focuses on groundwater arsenic contamination and investigating biogeochemical mechanisms for arsenic release. She is a trainee in the Columbia University Superfund Research Program and is also involved in a worldwide collaborative effort through the United States Geological Survey Powell Center to compile and explore the large volume of published groundwater arsenic datasets. She holds an M.Phil. & MA from Columbia University in geochemistry and a BA from the University of California, Berkeley in geophysics and statistics.
There is no cost to attend this event.
Intro into WSUD Training Session 1
AWA YWP Conference 2020
NWW Water Industry Night 2020
Share this with your Social Network
ICWSUDP 2019 : International Conference on Water-Sensitive Urban Design and Policy
Field Tour of Lake Algal Bloom Management
https://events.csiro.au/Events/2018/October/18/CSIRO-Floreat-seminar-series
147 Underwood Avenue
Floreat, Western Australia Australia Google Map
The aim of New WAter Ways is to build the water sensitive urban design capacity of Government and industry to improve the delivery of urban water management and water sensitive cities.
© Copyright New Water Ways Ltd | Site by DeCODE

redpajama_set_name: RedPajamaCommonCrawl


A Travelling Adjudicator is also touring this year, seeing three productions: two in the Kootenay Zone (KOZ) and one in the South Island Zone (SIZ). Our Travelling Adjudicator this year is Raimund Stamm, who has presented Theatre BC workshops and has adjudicated zone festivals for us in the past.
Theatre BC's Mainstage 2019 is being held in sunny Port Alberni, with nightly performances at the ADSS Performing Arts Theatre. This theatre first opened in 2013 in conjunction with the newly built Alberni District Secondary School. Most of our workshops will be held in the classrooms there, and The Capitol Theatre (home of host club Portal Players Dramatic Society) will be the hub of operations, with registrations & check-in being held there, an information desk, the nightly green room parties, meetings and is the location for our two Workshop Play slots.
We're thrilled that Dean Paul Gibson is back to adjudicate for us at Mainstage 2019. He's a Theatre BC favourite who is already familiar with the Vancouver Island community theatre scene, and adjudicated the North Island Zone Drama Festival in 2015, hosted by Portal Players in Port Alberni. He last adjudicated Mainstage in 2013, in Kamloops. Dean will provide a short public critique following each nightly performance, and then meet with the cast & crew the following morning for an in-depth chat and "coffee critique". Ticket holders will also be able to observe these highly educational sessions. Read our biography of Dean here.
More details and online registration is coming soon for these workshops!
Port Alberni is ready to host you this summer – accommodations options are listed on this Alberni Valley Chamber of Commerce website link.
Tickets to all the shows will be available online soon. Individual performance tickets, 3-day, 5-day and 8-day ticket packages can be bought at early bird and member discounted rates – keep checking back for when they go on sale.
The ADSS Performing Arts Theatre has 500 seats, all with a great view of the stage, and with festival seating in place for Mainstage.
More Mainstage 2019 information will be rolled out throughout the weeks leading up to our big Provincial Festival, including online registration forms for participants, workshop registration forms and online ticket sales to the shows. Stay tuned!
Theatre BC will feature one Wild Card performance at Mainstage 2019. The Wild Card performance will be on Friday, July 12 at the ADSS Performing Arts Theatre in Port Alberni, and the club drawn must be able to attend and perform on that date, and attend the following morning's Coffee Critique with adjudicator Dean Paul Gibson.
Qualifying clubs must have performed in a Zone Drama Festival or have been seen by the Travelling Adjudicator. Applicants which have been awarded "Best Play" in their Festival or by the Travelling Adjudicator will be removed from contention in the random draw.
Wild Card entry applications are completely confidential and will not be shared with your Zone Festival adjudicator, so there will be no impact on potential awards in your Zone Festival.
As soon as all Festivals have ended, the random selection will take place, and the successful Club will be informed of their selection.
Clubs are also welcome to enter their productions into our two Workshop Play slots, which will also be chosen by random draw. Workshop Play application details here.
Apply Now, TWO Slots! 2019 Mainstage Workshop Plays . . .
TWO Zone Festival-entered plays will be selected by a random draw to participate in MAINSTAGE's non-adjudicated "Workshop Play" component at Mainstage 2019, which takes place over a two different three-day periods.
SLOT 1: Saturday, July 6 through Monday, July 8 with instructor Nicolle Nattrass.
SLOT 2: Tuesday, July 9 through Thursday, July 11 with instructor Andrew McIlroy.
Workshop Plays will be held at The Capitol Theatre, 4904 Argyle Street, in Port Alberni, B.C.
Your application to be considered for the Workshop Play is confidential and will not be shared with the adjudicator in your Festival – there will be no impact on potential award selections at your Zone Festival.
Click on the link below to download the application form. Application deadline is April 30, 2019.
2019 Workshop Play Application Form.
Clubs may also want to consider applying for the WILD CARD slot at Mainstage 2019. Click this link for more details.
It's time to think ahead to Theatre BC's Mainstage 2019 being held in Port Alberni this year from July 5 to 13, 2019. The nightly performance venue will be the ADSS Performing Arts Theatre, while the hub of operations will be at The Capitol Theatre.
Being hosted jointly by Portal Players Dramatic Society and Theatre BC's North Island Zone, the organizing committee is working hard to put all the details together, and will have more updates to share soon.
Tickets to all performances for the nightly Mainstage performances (ADSS Theatre), and the before & after Workshop Play performances (Capitol Theatre) will be available to the general public. Individual show tickets, along with 3-show, 5-show and full 8-show Mainstage ticket packages will also be available. The complete playbill for the week will be known once all of the regional Zone Drama Festivals are complete.
For the nightly Mainstage performances, tickets and packages include admittance to the show at the ADSS Theatre; to the Green Room party at The Capitol Theatre following the performance; and a seat at the next morning's in-depth coffee critique/adjudication with the cast & crew from the prior night's show.
Find out more about accommodation options at this link.
Please join our Theatre BC Facebook Group for more notices and updates.


Browse listings of Member users here at Wheelchair Dating that are associated with Tennis. Talking to others that have like minded interests is a pefect way to find things to do on a first date. Signup for a 100% Free Account to Meet Someone Tonight!


text:
KCT & Bros. (Coal Sales) Ltd.
Indian City Properties
The Waterbase Limited
Life@KCT
KCT Group is one of India's leading business conglomerates with interests in sectors ranging from coal and real estate to aquaculture & manufacturing.
Coal Logistics
Incorporated in 1987, The Waterbase Limited (TWL) is a pioneer in the Indian aquaculture Industry. TWL is an ISO certified, fully – integrated aquaculture unit with it's own shrimp farms, shrimp feed plants and processing facilities, which allows TWL to work in a perfectly coordinated and organised manner.
TWL has been in the industry for more than two decades and understands the customer needs and meets them by implementing a code of good manufacturing practices. We believe that quality and food safety are cornerstones to our business. TWL strive continuously to improve it's product for better performance through investment in R & D and rigorous testing.
TWL is committed to the promotion & growth of sustainable aquaculture in India through responsible sourcing, eco friendly practices, innovation and farmers' education.
Two star Best Aquaculture Practice (BAP) certified company with 2 feed processing plants. Processing plants are ISO certified and FDA (USA) and HAACP (EU & UK) approved
First fully integrated Aquaculture unit of India , set up in 1989
Extensive R&D capability and pioneers in inculcating scientific approach in India
First to launch the Vannamei shrimp feed in India
© KCT GROUP 2017 | ALL RIGHTS RESERVED


Former Indonesian President Suharto, 86, is in a Jakarta hospital in critical condition. Doctors say he is suffering heart problems. Chad Bouchard reports from Jakarta.
The former president was admitted to a hospital Friday with swelling throughout his body. Doctors say Mr. Suharto's heart has become too weak to pump blood properly, and his kidneys are deteriorating.
Andi Mallarangeng, spokesman for current president Susilo Bambang Yudhoyono, says the president visited Mr. Suharto in the hospital Saturday and found him too weak to talk.
"He was conscious, he was not in a coma in that sense, he was conscious, but he was very weak," he said. "The presidential doctors informed the president that President Suharto is in [a] critical situation."
Suharto ruled Indonesia for 32 years, stepping down after widespread demonstrations and rioting during the Asian economic crisis of 1998.
He has since lived quietly in an upscale district of central Jakarta.
The anti-corruption organization Transparency International says Mr. Suharto embezzled more from Indonesia than any other modern leader, an estimated $15 billion to $35 billion. Mr. Suharto has steadfastly denied such accusations.


text:
Class 1 Integron-Borne, Multiple-Antibiotic Resistance Encoded by a 150-Kilobase Conjugative Plasmid in Epidemic Vibrio cholerae O1 Strains Isolated in Guinea-Bissau
Anders Dalsgaard, Anita Forslund, Andreas Petersen, Derek J. Brown, Francisco Dias, Serifo Monteiro, Kåre Mølbak, Peter Aaby, Amabelia Rodrigues, Anita Sandström
Anders Dalsgaard
Department of Veterinary Microbiology, The Royal Veterinary and Agricultural University, DK-1870 Frederiksberg C, and
Anita Forslund
Andreas Petersen
Derek J. Brown
Francisco Dias
Laboratório Nacional de Saúde Pública and
Serifo Monteiro
Kåre Mølbak
Projecto de Saúde, Bandim, Guinea-Bissau
Danish Epidemiology Science Centre, Statens Seruministitut, Copenhagen, Denmark, and
Peter Aaby
Amabelia Rodrigues
Department of Epidemiology, Ministry of Public Health,Bissau, and
Anita Sandström
DOI: 10.1128/JCM.38.10.3774-3779.2000
In the 1996–1997 cholera epidemic in Guinea-Bissau, surveillance for antimicrobial resistance showed the emergence of a multidrug-resistant strain of Vibrio cholerae O1 during the course of the epidemic. The strain was resistant to ampicillin, erythromycin, tetracycline, furazolidone, aminoglycosides, trimethoprim, and sulfamethoxazole. Concomitant with the emergence of this strain, we observed a resurgence in the number of registered cholera cases as well as an increase in the case fatality rate from 1.0% before the emergence of the multiple-drug-resistant strain to 5.3% after the emergence of the strain. Our study shows that the strain contained a 150-kb conjugative multiple-antibiotic resistance plasmid with class 1 integron-borne gene cassettes encoding resistance to trimethoprim (dhfrXII) and aminoglycosides [ant(3")-1a]). The finding of transferable resistance to almost all of the antibiotics commonly used to treat cholera is of great public health concern. Studies should be carried out to determine to what extent the strain or its resistance genes have been spread to other areas where cholera is endemic.
Following its arrival on the African continent in 1970, the seventh cholera pandemic, caused by Vibrio cholerae O1 biotype El Tor, spread throughout West Africa with devastating consequences (35). However, Guinea-Bissau, situated along the West African coast north of Guinea, first reported outbreaks in 1987 (42), which were followed by an epidemic in 1994 that extended into early 1995 (8). We previously determined by phenotypic and genotypic analyses how the epidemics in 1987 and 1994–1995 were caused by different O1 strains, with the 1994–1995 epidemic strain likely to have been introduced by fishermen or travelers from Guinea (8).
The most recent epidemic in Guinea-Bissau began in October 1996, extended throughout 1997, and included a total of 26,967 reported cases. The epidemic curve showed a bimodal pattern, with the first peak occurring in November 1996 and another peak extending over the summer of 1997 (Fig. 1). During the epidemic, monitoring of antimicrobial resistance was carried out by the National Public Health Laboratory in Bissau, Guinea-Bissau. This surveillance showed the emergence of a multidrug-resistant strain during 1997, concomitant with the second peak of the epidemic. At the same time, an increase in the case fatality rate was observed.
Number of cases of cholera and number of deaths reported during the cholera epidemic in Guinea-Bissau in 1996–1997.
Reports of toxigenic V. cholerae strains resistant to the antibiotics commonly used for treatment are appearing with increasing frequency, although the genetic mechanisms for the resistance are often not determined (2, 9, 13). Of particular interest and concern are the acquisition and transfer of resistance genes through mobile genetic elements, as such elements may spread among bacteria of different genera. Horizontal transferable plasmid-encoded resistance has been found on several occasions in V. cholerae O1 (4, 12, 13). Recently, Waldor et al. (41) showed that resistance genes among V. cholerae O1 and O139 strains in India were carried on a 62-kb self-transmissible, chromosomally integrating genetic element with properties similar to those of a conjugative transposon. A relatively new group of gene expression elements named integrons has been described as vehicles for the acquisition of antibiotic resistance genes (reviews have been provided by Hall and Stokes [16], Recchia and Hall [32], and Sundström [34]) (15, 33). So far three classes of integrons have been identified, and class 1 integrons have been found to be prevalent among clinical isolates. Class 1 elements were originally defined to be composed of two conserved segments, the 5′ conserved segment (5′-CS) and the 3′ conserved segment (3′-CS), and an internal variable region which contains gene cassettes encoding antibiotic resistance determinants (15, 33). The 5′-CS segment contains theintI1 gene, which encodes the type 1 integrase and which is responsible for site-specific insertion and excision of gene cassettes (3). Furthermore, the 5′-CS segment harbors theattI1 site, which is responsible for recombination. The 3′-CS segment contains the qacEΔ1 and sul1genes, which encode resistance to quaternary ammonium compounds and sulfonamides, respectively. It should be noted that class 1 integrons may not always contain the entire 3′-CS segment (32, 34).
The objective of the present study was to determine the genetic mechanisms of antibiotic resistance and the genotypes of V. cholerae O1 strains isolated during the cholera outbreak in Guinea-Bissau in 1996–1997.
Surveillance of the cholera epidemic and characterization of bacterial strains.Information about the number of cholera cases and the number of deaths was collected and reported to the Ministry of Health in Bissau for the period from October 1996 to February 1998. TenV. cholerae O1, biotype El Tor, serotype Ogawa strains recovered during the cholera outbreak in Guinea-Bissau in 1996–1997 were examined. Four and three O1 strains isolated during cholera outbreaks in 1987 (8) and 1994, respectively, were included for comparison purposes (V. cholerae O1 strains from the 1994 outbreak were kindly provided by Joy G. Wells and Fred Angulo, Centers for Disease Control and Prevention, Atlanta, Ga.) (Table1). Each stool specimen was enriched in alkaline peptone-water (pH 8.6) for 6 h, after which a loopful of the surface pellicle was streaked onto thiosulfate-citrate-bile salt-sucrose agar (Difco, Detroit, Mich.). Suspected V. cholerae isolates were tested by agglutination tests with polyvalent O1-specific and monospecific Ogawa and Inaba antisera (Difco). V. cholerae isolates and appropriate controls were tested for DNA sequences encoding cholera toxin (CT) in hybridization studies with an alkaline phosphatase-labeled 23-base oligonucleotide probe (44).
V. cholerae O1 strains isolated in Guinea-Bissau during cholera outbreaks in 1987, 1994, and 1996–1997
Antibiotic susceptibility testing and transfer of resistance.Each isolate was tested for susceptibility to 14 antibacterial agents representing different antimicrobial classes by the disk diffusion method with Mueller-Hinton agar (Difco) as described by the National Committee for Clinical Laboratory Standards (28). A nalidixic acid-resistant mutant strain ofEscherichia coli K-12, strain J53-1 (lac+pro met Nalr) (37), and a streptomycin- and rifampin-resistant mutant strain of V. cholerae O1, strain 1407, isolated during the epidemic in Guinea-Bissau in 1987 (8) were used as recipients in conjugation experiments. Following mating on nonselective L agar (Difco) incubated at 37°C for 4 to 6 h of growth, exconjugants were harvested and appropriate dilutions were streaked onto plates of Fluorocult E. coliO157:H7 agar (Merck, Virum, Denmark) supplemented with 50 μg of nalidixic acid per ml and adequate concentrations of selected drugs. Phenotypic appearance was used to differentiate possible spontaneous nalidixic acid-resistant donors from the exconjugants.
Plasmid analysis.Plasmid extraction was carried out by the method of Kado and Liu (22), modified by incubating the cells at elevated pH (12.54) for 30 min at 56°C during the lysis step. V. cholerae O1 strain 1075/25 carrying a 150-kb plasmid was used as a positive control (36). The enzymeEcoRI was used in restriction fragment length polymorphism (RFLP) analysis. Electrophoresis and visualization of plasmids were carried out essentially as described previously (29).
Ribotyping and CT genotyping.The genotype of V. cholerae O1 was established by ribotyping with the restriction enzyme BglI to digest chromosomal DNA (31). Ribotyping was performed by the procedure described by Dalsgaard et al. (5) with digoxigenin-labeled 16S and 23S rRNA probes. RFLP analysis of DNA sequences encoding CT genes (CT genotyping) was performed by hybridization of nylon membranes withBglI-digested DNA with a digoxigenin-labeled oligonucleotide CT probe (44). A 1-kb DNA molecular size standard (GIBCO BRL, Gaithersburg, Md.) was used as a size marker in ribotyping and CT genotyping.
PCR amplification and sequencing of class 1 integrons.First, we selected a specific class 1 integron primer set, qacEΔ1 and sul1, directed at the 3′-CS conserved segment of class 1 integrons (30, 32). As few class 1 integrons are known to not contain the qacEΔ1 and sul1 genes, these would not be detected with the selected primers. Figure2 demonstrates the general structure of class 1 integrons together with the amplicons sequenced and gene cassettes found. PCR was carried out as described by Dalsgaard et al. (7). DNAs from strains yielding a PCR product with the class 1 primers were subsequently amplified with the integron primers in-F and in-B (F is forward and B is backward) which amplify the region between the 5′-CS and 3′-CS conserved segments, generating products of variable sizes depending on the numbers and lengths of the inserted gene cassettes (Fig. 1 and Table 1). The in-B primer anneals at the 3′-CS conserved segment. Primers in-F and ant(3")-Ia-B (addA-B) were used to assess if the integron contained a gene cassette encoding resistance to streptomycin and spectinomycin (Fig. 2) (25). Appropriate control strains and molecular size markers were included, as described previously (7). Amplified DNA was purified before sequencing by using Microspin S-400HR columns purchased from Pharmacia Biotech (Hillerød, Denmark), and the nucleotide sequence was determined by cycle sequencing with the AmplitaqFS dye terminator kit and a Pharmacia Biotech ALF automated DNA sequencing apparatus. The identities of the sequences determined were analyzed by comparison of the sequences with the gene sequences in databases by using the BLAST software (1).
Integron structure and resistance gene cassettes found in V. cholerae O1. The PCR primers used and the amplicons obtained are shown below the integron structure, with the bold line representing the amplicon sequenced. att1 is a site which is responsible for recombination, and qacEΔ1 andsul1 encode resistance to disinfectants and sulphonamides, respectively. The individual gene cassettes are shown together with their recombination sites (59-bp element).
Statistical methods.To adjust for regional differences in mortality, we used multivariate logistic regression analysis to compare the case fatality rates in the first and the second parts of the epidemic.
Epidemic surveillance data.The epidemic in Guinea-Bissau, which started on October 6, 1996, and which ended on January 12, 1998, included a total of 26,967 reported cases (national attack rate, 24 cases per 1,000 inhabitants). The epidemic curve had a bimodal pattern (Fig. 1). During the period from October 1996 through March 1997, 10,386 of 10,574 cases (98%) were from the capital, Bissau, and the surrounding region, Biombo. During the period from April 1997 through January 1998, 11,020 (67%) of 16,393 cases were from the capital area and Biombo. Throughout the epidemic, the attack rate was highest in the capital area (68 cases per 1,000), but after April 1997, the epidemic spread to the interior regions of Cacheu, Oio, Bafatá, Quinara, and Tombali. Also, the archipelago of Bijagos was affected.
The case fatality rate increased from 1.0% (102 deaths per 10,574 cases) during the period from October 1996 through March 1997 to 5.3% (860 deaths per 16,393 cases) during the rest of the epidemic. There was a significantly higher rate of mortality among the case patients from the interior regions of Guinea-Bissau (P < 0.0001). Furthermore, the multivariate analysis showed that, independent of geography, there was an increase in the case fatality rate in the second epidemic period (adjusted odds ratio, 1.8; 95% confidence interval, 1.4 to 2.3; P < 0.0001). Among hospitalized case patients from Bissau, the case fatality rate increased from 3.2% (69 per 2,178) in the first period to 5.8% (155 per 2,680) in the second period (risk ratio, 1.9; P < 0.001).
Antibiotic susceptibility testing and plasmid analysis.Table1 shows the characteristics of the V. cholerae O1 strains isolated in Guinea-Bissau during the cholera outbreaks in 1987, 1994, and 1996–1997. Each of the strains hybridized with the CT probe. Seven strains, including all six strains recovered in 1997, isolated at different locations and different times during the epidemic in 1996–1997 had identical antibiograms that showed multiple-drug resistance, including resistance to furazolidone, sulphonamides, trimethoprim, and tetracycline (Table 1). In addition, each of these strains contained a 150-kb plasmid which was found to be identical in each case by RFLP analysis (profile D) with the restriction enzymeEcoRI (Fig. 3 and Table 1). Interestingly, three strains isolated when the epidemic began in December 1996 showed different resistance patterns, and one of these strains contained a 150-kb plasmid that showed a different RFLP profile (profile A). Strains isolated in 1987 were resistant to colistin only, did not contain plasmids, and showed a unique ribotype and CT genotype (8) (ribotypes are shown in Fig. 4).
RFLP analysis of the 150-kb plasmid isolated fromV. cholerae O1 strains isolated during cholera outbreaks in Guinea-Bissau. The explanations of the lanes include the strain designation and RFLP type unless stated otherwise. Lanes: a,HindIII-digested phage lambda DNA (weight marker); b, reference strain 1075/25; c, F2189, type E; d, F2196, type F; e, 9862, type A; f, 9868, type D; g, C230, type D; h, C231, type D; i, C233, type D; j, C235, type D; k, C236, type D; 1, C240, type D; m,HindIII-digested phage lambda DNA (weight marker).
Ribotyping and CT genotyping.Ribotyping showed that the 1996–1997 outbreak strains all belonged to the same type, type II, which was also shown for strains isolated in 1994 but which was clearly different from type Ia, as demonstrated with strains isolated in 1987 (Fig. 4). These findings were supported by CT genotyping, in which all strains from 1994 and 1996–1997 showed two fragments of 10.0 and 12.8 kb, whereas strains from 1987 demonstrated two fragments of 6.8 and 11.2 kb (Table 1).
Examples of BglI ribotypes of V. cholerae O1 strains associated with cholera outbreaks in Guinea-Bissau. Unless stated otherwise, the explanations of the lanes include the strain designation, ribotype, and year of isolation. Lanes: A, 1-kb molecular weight standard; B, 1407, ribotype Ia, 1987; C, C235, type II, 1997; D, 1445, type Ia, 1992; E, 1452, type Ia, 1992; F, 1482, type Ia, 1992; G, F2107, type II, 1994; H, F2196, type II, 1994; I, F2189, type II, 1994; J, 1-kb molecular weight standard.
Resistance transfer: PCR amplification and sequencing of class 1 integrons.As transferable plasmid-encoded multiple-drug resistance has been reported for V. cholerae O1, strain C230 was selected as a representative strain for transfer experiments intoE. coli K-12. One type of transconjugant that showed susceptibility patterns identical to that of the donor strain was found, except that the transconjugants were sensitive to colistin and furazolidone. Antibiotic susceptibility testing and plasmid analysis of the transconjugants, including RFLP analysis, revealed that the 150-kb plasmid was transferred and that the antibiotic resistance, except for resistance to colistin and furazolidone, was plasmid encoded. The 150-kb plasmid was subsequently transferred from the transconjugants back to strain 1407 and was further directly transferred from strain C230 to strain 1407. Again, the transfer of resistance was found as described above, with apparent higher frequencies in the direct transfer from V. cholerae strain C230 into strain 1407 than in transfer studies with E. coli K-12 (transfer frequencies were not calculated but were evaluated only by visual inspection of the agar plates on which mating occurred).
As the vast majority of strains containing class 1 integrons were resistant to sulphonamides, we investigated if the sulphonamide resistance was associated with the presence of class 1 integrons (30, 32). An approximately 800-bp 3′-CS PCR product was obtained from strain 9868 isolated in December 1996 and each of the six O1 strains isolated in 1997 with the primers qacEΔ1-F and sulI1-B (Fig. 2). None of the remaining strains yielded an amplicon with these primers and therefore did not seem to contain class 1 integrons (Table 1). The use of the in-F and in-B primers yielded a PCR product of 1,800 bp. DNA sequencing analysis of strain 9868 revealed that the 1,800-bp amplicon contained the dhfrXII(dfrA12) gene cassette encoding resistance to trimethoprim, an open reading frame, orfF, with an unknown function, and the gene cassette ant(3")-1a(aadA2), which confers resistance to streptomycin and spectinomycin (Fig. 2). The sequences of the dhfrXIIgene cassette and orfF have been published in GenBank under accession number Z21672 (18), while the sequence forant(3")-1a has the accession numberD43625 (25). The sequences of the three cassettes were identical to those that have been published previously. PCR with in-F and ant(3")-Ia-B primers also yielded an approximately 1,800-bp amplicon and therefore confirmed the presence and location of the aminoglycoside resistance gene cassetteant(3")-Ia (Fig. 2) (25). Furthermore, PCR of the cell lysates of the transconjugants E. coli K-12 and V. cholerae O1 strain 1407 with qacEΔ1-F and sul1-B and with in-F and in-B primers yielded 800- and 1,800-bp amplicons, respectively, showing that the class 1 integron and gene cassettes are located and transferred on the 150-kb plasmid.
The 1996–1997 cholera epidemic in Guinea-Bissau, which involved a reported 26,967 cases, was caused by a V. cholerae O1 that had a genotype identical to that of the 1994–1995 epidemic strain (8) but that contained a 150-kb conjugative multiple-antibiotic resistance plasmid with class 1 integron-borne gene cassettes encoding resistance to trimethoprim (dhfrXII) and aminoglycosides [ant(3")-1a].
Studies indicate that class 1 integrons may be widespread in multiple-drug-resistant clinical isolates of bacterial enteropathogens (17, 21, 24). However, few studies have investigated their presence and importance in V. cholerae. Dalsgaard et al. (7) reported that V. cholerae O1 strains isolated in Vietnam after 1990 were resistant to sulphonamides and streptomycin and harbored chromosomally located class 1 integrons containing the aminoglycoside gene cassetteant(3")-1a. Furthermore, an extensive study of the distribution and content of class 1 integrons in differentV. cholerae O serotypes strains isolated in Thailand revealed that O1 and non-O1 serotype strains mainly of clinical origin but not O139 serotypes contained antibiotic resistance genes located on class 1 integrons (6). Finally, class 1 integrons containing the aadA1 gene cassette, which encodes resistance to spectinomycin, were described among strains from a minor cholera outbreak in Albania and Italy in 1994 (10).
The 1996–1997 cholera epidemic was associated with a high case fatality rate, in particular during the second epidemic period. Cholera patients were managed according to standard guidelines, with emphasis on rehydration therapy. Antimicrobial treatment was, in principle, reserved for the patients with severe cases, but in practice, the therapeutic and prophylactic use of tetracycline was widespread. The increased mortality rate during the second period was partly due to the spread of the epidemic to the interior, where access to primary health care is more difficult and limited than in the capital area (14). However, the increase in the case fatality rate could not be attributed solely to the spread of the epidemic. Thus, an increased case fatality rate was observed among hospitalized patients from the capital and in the multivariate analysis by adjusting for regional differences. We suggest that the finding of transferable resistance to all of the antimicrobials most commonly used to treat cholera could contribute to the increased fatality rate. Although the primary objective of case management is the treatment and prevention of dehydration, it has been shown that appropriate antimicrobial therapy can reduce by half the volume of stool purged during illness, as well as shorten the duration of excretion of V. cholerae by the same amount (26). In addition, the presence of resistant strains may cause an increased risk of secondary transmission due to prolonged excretion. The widespread use of antimicrobials for any reason in the community may indeed favor the transmission of the resistant strain and may be one of the explanations for the second wave in the 1996–1997 epidemic.
In contrast to the strains from the earlier epidemics, the 1996–1997 strain was resistant not only to low-dose and high-dose tetracycline but also to other commonly used antimicrobials such as furazolidone and trimethoprim. It is likely that the multidrug-resistant outbreak strain in the 1996–1997 epidemic was derived from the same clone responsible for the 1994–1995 epidemic but had subsequently acquired the 150-kb resistance plasmid from an unknown source. The plasmid-containing strain may have come to dominate through the selective pressure caused by the prophylactic or therapeutic use of antimicrobials. This hypothesis is corroborated by the fact that some strains isolated early in the epidemic were less resistant and did not contain plasmids.
Although genes for resistance to eight antibiotics were shown to be located on the 150-kb plasmid, only a minor part of the resistance, namely, resistance to streptomycin, sulphonamides, and trimethoprim, were contained in gene cassettes of the class 1 integron. This corroborates the findings of the content of class 1 integrons inV. cholerae strains from Thailand (6) and in members of the family Enterobactericeae andPseudomonas spp., in which gene cassettes often encode resistance to aminoglycosides, trimethoprim, and β-lactamases. ThedhfrXII gene cassette product confers a level of resistance to trimethoprim which is about 10 times stronger than that conferred bydhfrI (18), the most widespread gene coding trimethoprim resistance (38, 39). It is believed that the spread of trimethoprim resistance genes among clinical isolates is mostly by the integron mechanism (20).
The cluster of three gene cassettes described in this paper has been found previously in five Escherichia coli isolates from patients with urinary tract infections and geriatric patients and in four patients with Shigella enteritis in Finland (18). The integrons were borne on a Tn21-like transposon located on a 70-kb plasmid (18), whereas our integron was located on a 150-kb self-transmissible plasmid. We did not investigate if the integron was inserted in a transposon, but this may be an explanation for the presence of the apparent identical integron on different plasmids. Although it is believed that the integrons themselves are mobile elements, this possibility remains to be verified experimentally. The presence of the same arrangement of gene cassettes in different bacterial species and on different plasmids suggests that the integron-associated mechanism of gene capture is a widespread and an important mechanism by which bacteria acquire novel resistance genes.
Surprisingly and in contrast to our earlier report of the cholera epidemic in 1994–1995, in which strains were found to be resistant to colistin, trimethoprim, and the vibriostatic agent O129, the three O1 strains isolated in 1994 showed increased antibiotic resistance, with strains F2196 and F2189 carrying a 150-kb plasmid (Table 1) (19). Two strains isolated early in the 1996–1997 epidemic did not carry plasmids, whereas another strain carried a 150-kb plasmid without class 1 integrons and was clearly different from the 1996–1997 epidemic strains, as determined by RFLP analysis. These findings suggest that several large plasmids are present in V. cholerae and that the 150-kb plasmid containing the integron may have been received by strains in the early stages of the epidemic.
Since the 1996–1997 epidemic in Guinea-Bissau, the eastern and southern parts of Africa have been hit severely by several cholera epidemics, with mortality rates as high as 20% being reported (43). Of the total of 147,425 cases of cholera reported to the World Health Organization in 1997, 80% occurred in Africa (43). Despite the tremendous negative impact on human health and economy caused by the cholera epidemics, little information has been published about the transmission patterns and characteristics of the V. cholerae O1 strains involved. Also, research suggests that V. cholerae is undergoing genetic changes at higher rates than previously thought. Manning et al. (27) described a new mega-integron genetic structure named VCR (V. choleraerepeat) which was proposed to represent a genetic hot spot in the capture of genes, thereby enabling more rapid adaptation and evolution in V. cholerae. Also, the recent evidence that the structural genes for CT and the toxin-coregulated pilus antigen are encoded by filamentous bacteriophages, which may integrate chromosomally or replicate as a plasmid following horizontal gene transfer, have provided new and intriguing information about the emergence of toxigenic V. cholerae (11, 23, 40). Thus, it is of utmost clinical importance that studies be carried out to characterize the current epidemic strains, including determination of the levels and mechanisms of antibiotic resistance and the extent which the 1996–1997 strain and its resistance genes have spread to other parts of Africa.
We thank Joy G. Wells and Fred Angulo of the Centers for Disease Control and Prevention for providing the strains from the cholera epidemic in 1994. We are grateful for the technical assistance provided by Anne-Mette Petersen.
Kåre Mølbak and Anders Dalsgaard were supported by the Danish Council for Development Research (DANIDA grants 104.Dan.8/1114 and 90928, respectively).
Returned for modification 14 May 2000.
Accepted 31 July 2000.
Altschul S. F.,
Gish W.,
Miller W.,
Myers E. W.,
Lipman D. J.
Basic local alignment search tool.J. Mol. Biol.2151990403410
Bag P. K.,
Maiti S.,
Sharma C.,
Ghosh A.,
Basu A.,
Mitra R.,
Bhattacharya S. K.,
Nakamura S.,
Yamasaki S.,
Takeda Y.,
Blair B. G.
Rapid spread of the new clone of Vibrio cholerae O1 biotype El Tor in cholera endemic areas in India.Epidemiol. Infect.1211998245251
Collis C. M.,
Grammaticopoulow G.,
Briton J.,
Stokes H. W.,
Hall R. M.
Site-specific insertion of gene cassettes into integrons.Mol. Microbiol.919934152
Coppo A.,
Colombo M.,
Pazzani C.,
Bruni R.,
Mohamud K. A.,
Omar K. H.,
Mastrandrea S.,
Salvia A. M.,
Rotigliano G.,
Maimone F.
Vibrio cholerae in the horn of Africa: epidemiology, plasmids, tetracycline resistance gene amplification, and comparison between O1 and non-O1 strains.Am. J. Trop. Med. Hyg.531995351359
Dalsgaard A.,
Echeverria P.,
Larsen J. L.,
Siebeling R.,
Serichantalergs O.,
Huss H. H.
Application of ribotyping for differentiating Vibrio cholerae non-O1 isolates from shrimp farms in Thailand.Appl. Environ. Microbiol.611995245251
Forslund A.,
Sandvang D.
Distribution and content of class 1 integrons in different Vibrio cholerae O-serotypes strains isolated in Thailand.Antimicrob. Agents Chemother.44200013151321
Tam N.,
Vinh D. X.,
Cam P. D.
Cholera in Vietnam: changes in genotypes and emergence of class I integrons containing aminoglycoside resistance gene cassettes in Vibrio cholerae O1 strains isolated from 1979 to 1996.J. Clin. Microbiol.371999734741
Mortensen H. F.,
Mølbak K.,
Dias F.,
Echeverria P.
Molecular characterization of Vibrio cholerae O1 strains isolated during cholera outbreaks in Guinea-Bissau.J. Clin. Microbiol.34199611891192
Dubon J. M.,
Palmer C. J.,
Ager A. L.,
Shor-Posner G.,
Baum M. K.
Emergence of multiple drug-resistant Vibrio cholerae O1 in San Pedro Sula, Honduras.Lancet3491997924
Falbo V.,
Carattoli A.,
Tosini F.,
Pezzella C.,
Dionisi A. M.,
Luzzi I.
Antibiotic resistance conferred by a conjugative plasmid and a class I integron in Vibrio cholerae O1 El Tor strains isolated in Albania and Italy.Antimicrob. Agents Chemother.431999693696
Faruque S.,
Asadulghani M.,
Saha M. N.,
Abdul Alim A. R. M.,
Albert M. J.,
Nasirul Islam K. M.,
Mekalanos J. J.
Analysis of clinical and environmental strains of nontoxogenic Vibrio cholerae for susceptibility to CTXφ: molecular basis for origination of new strains with epidemic potential.Infect. Immun.66199858195825
Finch M. J.,
Morris J. G. Jr.,
Kaviti J.,
Kagwanja W.,
Levine M. M.
Epidemiology of antimicrobial resistant cholera in Kenya and East Africa.Am. J. Trop. Med. Hyg.391988484490
Glass R. I.,
Huq I.,
Alim A. R. M. A.,
Yunus M.
Emergence of multiply antibiotic-resistant Vibrio cholerae in Bangladesh.J. Infect. Dis.1421980939943
Gunnlaugsson G.,
Angulo F. J.,
Einarsdottir J.,
Passa A.,
Tauxe R. V.
Epidemic cholera in Guinea-Bissau: the challenge of preventing deaths in rural West Africa.Int. J. Infect. Dis.42000813
Hall R. M.,
Collis C. M.
Mobile gene cassettes and integrons: capture and spread of genes by site-specific recombination.Mol. Microbiol.151995593600
Stokes H. W.
Integrons: novel DNA elements which capture genes by site-specific recombination.Genetica901993115132
Hannecart-Pokorni E.,
Depuydt F.,
Wit L. D.,
Bossuyt E. V.,
Content J.,
Vanhoof R.
Characterization of the 6′-N-aminoglycoside acetyltransferase gene aac(6′)-Il associated with a sulI-type integron.Antimicrob. Agents Chemother.411997314318
Heikkila E.,
Skurnik M.,
Sundström L.,
Huovinen P.
A novel dihydrofolate reductase cassette inserted in an integron borne on a Tn21-like element.Antimicrob. Agents Chemother.37199312971304
Holmberg S. D.,
Day D. E.,
Parker R. D. R.,
Rao N. U.,
Harris J. R.,
Hargrett N. T.,
Kansou N.,
Blake P. A.
Foodborne transmission of cholera in Micronesian households.Lancet111984325328
Huovinen P.,
Swedberg G.,
Sköld O.
Trimethoprim and sulfonamide resistance.Antimicrob. Agents Chemother.391995279289
Jones M. E.,
Peters E.,
Weersink A.-M.,
Fluit A.,
Verhoef J.
Widespread occurrence of integrons causing multiple antibiotic resistance in bacteria.Lancet349199717421743
Kado C. I.,
Liu S. T.
Rapid procedure for detection and isolation of large and small plasmids.J. Bacteriol.145198113651373
Karaolis D. K. R.,
Somara S.,
Maneval D. R. J.,
Johnson J. A.,
Kaper J. B.
A bacteriophage encoding a pathogenicity island, a type-IV pilus and a phage receptor in cholera bacteria.Nature3991999375379
Kazama H.,
Hamashima H.,
Sasatsu M.,
Arai T.
Distribution of the antiseptic resistance genes qacE and qacE1 in gram-negative bacteria.FEMS Microbiol. Lett.1591998173178
Kizu K.,
Iwasaki M.,
Isolation and structure of a new integron that includes a streptomycin resistance gene from the R plasmid of Pseudomonas aeroginosa.FEMS Microbiol. Lett.1341995137141
Mahalanabis D.,
Molla A. M.,
Sack D. A.
Clinical management of cholera Cholera. Barua D., Greenough W. B. I. 1992 253 283 Plenum Medical Book Company New York, N.Y
Manning P. A.,
Clark C. A.,
Focareta T.
Gene capture in Vibrio cholerae.Trends Microbiol.719999395
National Committee for Clinical Laboratory Standards Performance standards for antimicrobial susceptibility testing. Approved standard M100-S8 18 1998 National Committee for Clinical Laboratory Standards Wayne, Pa
Olsen J. E.,
Larsen J. L.
Restriction fragment length polymorphism of the Vibrio anguillarum serovar O1 virulence plasmid.Appl. Environ. Microbiol.56199031303132
Paulsen I. T.,
Littlejoh T. G.,
Rådström P.,
Sköld O.,
Skurray R. A.
The 3′ conserved segment of integrons contains a gene associated with multidrug resistance to antiseptics and disinfectants.Antimicrob. Agents Chemother.371993761768
Popovic T.,
Bopp C. A.,
Olsvik Ö.,
Wachsmuth K.
Epidemiologic application of a standardized ribotype scheme for Vibrio cholerae O1.J. Clin. Microbiol.31199324742482
Recchia G. D.,
Gene cassettes: a new class of mobile element.Microbiology141199530153027
A novel family of potentially mobile DNA elements encoding site-specific gene-integration functions: integrons.Mol. Microbiol.3198916691683
Sundström L.
The potential of integrons and connected programmed rearrangements for mediating horizontal gene transfer.APMIS8419983742
Swerdlow D. L.,
Isaäcson M.
The epidemiology of cholera in Africa Vibrio cholerae and cholera. Wachsmuth I. K., Blake P. A., Olsvik Ö 1994 297 307 ASM Press Washington, D.C.
Tabtieng R.,
Wattanasri S.,
Seriwatana J.,
Bodhidatta L.,
Chatkaeomorakot A.,
Rowe B.
An epidemic of V. cholerae El Tor Inaba resistant to several antibiotics with a conjugative group C plasmid coding for type II dihydrofolate reductase in Thailand.Am. J. Trop. Med. Hyg.411989680686
Taylor D. E.,
Levine J. G.,
Bradley D. E.
In vivo formation of a plasmid cointegrate expressing two incompatibility phenotypes.Plasmid51981233244
Towner K. J.,
Brennan A.,
Zhang Y.,
Holtham C. A.,
Brough J. L.,
Carter G. I.
Genetic structures associated with spread of the type Ia trimethoprim-resistant dihydrofolate reductase gene amongst Escherichia coli strains isolated in the Nottingham area of the United Kingdom.J. Antimicrob. Chemother.3319942532
Tsakris A.,
Johnson A. P.,
Legakis N. J.,
Tzouvelekis L. S.
Prevalence of the type I and type II DHFR genes in trimethoprim-resistant urinary isolates of Escherichia coli from Greece.J. Antimicrob. Chemother.311993665671
Waldor M. K.,
Lysogenic conversion by a filamentous phage encoding cholera toxin.Science272199619101914
Tschäpe H.,
A new type of conjugative transposon encodes resistance to sulfamethoxazole, trimethoprim, and streptomycin in Vibrio cholerae O139.J. Bacteriol.178199641574165
World Health Organization Cholera in 1986.Weekly Epidemiol. Rec.621987141142
Wright A. C.,
Guo Y.,
Nataro J. P.,
Morris J. G. Jr.
Development and testing of a non-radioactive DNA oligonucleotide probe that is specific for Vibrio cholerae cholera toxin.J. Clin. Microbiol.30199223022306
Journal of Clinical Microbiology Oct 2000, 38 (10) 3774-3779; DOI: 10.1128/JCM.38.10.3774-3779.2000
You are going to email the following Class 1 Integron-Borne, Multiple-Antibiotic Resistance Encoded by a 150-Kilobase Conjugative Plasmid in Epidemic Vibrio cholerae O1 Strains Isolated in Guinea-Bissau
Drug Resistance, Microbial
R Factors


text:
Kaiserin-Augusta-Stift steht für:

 Kaiserin-Augusta-Stift (Potsdam)
 Kaiserin-Augusta-Stift (Wuppertal)

redpajama_set_name: RedPajamaWikipedia


Brighton is a unique, stylish, cosmopolitan and eccentric city by-the-sea offering amazing shopping.
The extraordinary Royal Palace – The Royal Pavilion – the historic lands, the seaside Pier, the fascinating museums, the elegant Regency architecture, a bustling marine, arts and culture galore and the colourful year-round events all combine to create a vibrant and special city.
A sparkling mix of sea and city, Brighton is a glamorous heritage city with fabulous leisure facilities and glittering.
Brighton College with its attractive enclosed campus is situated just a few minutes walk from the beaches and the town centre which is famous as one of Britain's most exciting and vibrant seaside towns. Beach volleyball, windsurfing or fun on the famous Brighton Pier are just a few of the many things to do.
Brighton is also an historic city – The Royal Pavilion dominates with its Regency splendour – with many fascinating places to visit.
Or why not wander through The Lanes – the famous and exclusive shopping district? Without doubt, Brighton is one of the most attractive venues for a teenage language student!
The college accommodation has recently been renovated and refurbished.
• Excellent dining room offers a choice of continental or English breakfast, self service two course lunch, two course dinner. The "Café de Paris" is also open during break times for students to purchase refreshments.
• Full day excursions include London and Canterbury . Half days may include Portsmouth , Hastings or Arundel.


The Park City Museum is looking for new volunteers and we would love your help! Join us for our New Volunteer Orientation scheduled for Thursday, February 15, 2018 from 3-5PM to learn about the many opportunities to help preserve, protect, and promote Park City's history. The Park City Museum seeks volunteers to help install and take down traveling exhibits, help with special events, lead both adult tours and school group programs, and assist as gallery guides. All are invited to attend. If you have any questions, please contact Diane Knispel at education@parkcityhistory.org or 435-649-7457 ext. 102.


text:
$(document).ready(function() {
    $('#ojs_admin_quickswitch_switch').click(function(e) {
        e.preventDefault();
        var id = $('#ojs_admin_quickswitch_journal').val();
        window.location.href = Routing.generate('ojs_journal_dashboard_index', {'journalId': id});
    });
});

redpajama_set_name: RedPajamaGithub


text:
Gazelle in the House by Lisa Williams
HomeGazelle in the House by Lisa Williams
Gazelle in the House
A Green Rose Book
Lisa Williams's new collection, Gazelle in the House, is truly a book of stanzas: poetic rooms in which to dwell. Some of these dwellings have the uncanny familiarity of ordinary domestic space and others are as mysterious and disorienting as the depths of the sea. Painting with colors at times opaque, at times transparent, moving between shallows, tide-pools, and the abysses of dreams, Williams's voice is solitary, meditative, intimate—and in the end a means of revelation.
—Susan Stewart
Like the photographer who "wants that dialogue between a singer's gesture and what slants it," Lisa Williams elegantly slants the space between sight and sound in her striking third collection. She plumbs the worlds of eels and deep sea bells as deftly as she conjures unsparing snapshots of female adolescence.
—Rebecca Morgan Frank
"Go / become again a threshold," commands one of the speakers in Lisa Williams' newest collection, Gazelle in the House. Williams' poems consistently compel the reader to become a door; to pass through the most difficult of emotional landscapes. This collection asks us to traverse a burning landscape and to come out on the other side singed brighter and better. Williams moves us through this landscape with an intellectual and aesthetic rigor.
—Roger Reeves
Originally from Nashville, Lisa Williams teaches at Centre College in Danville, Kentucky. She is the author of Woman Reading to the Sea (W.W. Norton 2008), which won the Barnard Women Poets Prize, and The Hammered Dulcimer (Utah State University Press, 1998), which won the May Swenson Poetry Award. Her poems have been published or are forthcoming in Blackbird,Poetry, The Oxford American, and other magazines, and have been featured on Poetry Daily andVerse Daily, as well as in anthologies including Best American Poetry 2009, Bright Wings: An Illustrated Anthology of Poems About Birds, and American Poetry: Next Generation. Her essays on contemporary women poets have appeared in The Hollins Critic, The Cincinnati Review, and on Poetry Daily. Williams is the recipient of a 2011 Brown Foundation Fellowship awarded by the Brown Foundation and the Museum of Fine Arts, Houston; a 2010 Al Smith Individual Artist Fellowship awarded by the Kentucky Arts Council; and a 2004 Rome Prize in Literature, awarded by the American Academy of Arts and Letters.
Spilled Milk on Banjo
Read by Lisa Williams
Video by Adriane Little
Publication of this book and production of this video were made possible in part by a grant from the National Endowment for the Arts.
To find out more about how NEA grants impact individuals and communities, visit www.arts.gov.


text:
CITIZENSHIP OF THE CHILD BORN ABROAD
A child born abroad may automatically acquire U.S. citizenship of one or both parents. In order to transfer citizenship the parent who is U.S. citizen must reside in the U.S. for certain time. An individual who is a naturalized citizen can count time before and after naturalization towards required time. A child who acquires citizenship at birth does not need to file Form N-600, an an Application for Certificate of Citizenship. A person who seeks documentation of such status, however, must submit an application to obtain a Certificate of Citizenship from USCIS. A person may also apply for a U.S. Passport with the Department of State to serve as evidence of his or her U.S. citizenship.
PARENTS ARE MARRIED
1. Child of Two U.S. Citizen Parents
A child born outside of the United States automatically becomes a U.S. citizen at birth if at the time of birth:
Both of the child's parents are U.S. citizens; and
At least one parent had resided in the U.S. at the time of birth.
A child needs only demonstrate that only one of the parents lived in the U.S. for one day to receive a consular report of birth abroad.
2. Child of U.S. Citizen Parent and U.S. National
One parent is a U.S. citizen and the other parent is a U.S. national; and
The U.S. citizen parent was physically present in the United States for a continuous period of at least one year prior to the birth of the child.
3. Child of U.S. Citizen Parent and Foreign National Parent
One parent is a foreign national and the other parent is a U.S. citizen; and
The U.S. citizen parent was physically present in the United States for at least 5 years, including at least 2 years after 14 years of age.
 Time abroad counts as physical presence in the United States if the time abroad was:
As a member of the U.S. armed forces in honorable status;
Under the employment of the U.S. government or other qualifying organizations; or
As a dependent unmarried son or daughter of such persons.
There is no requirement that the child live in the U.S. later in life to retain U.S. Citizenship.
4. Child of a U.S. Citizen Mother and Foreign National Father
A child born outside of the United States and its outlying possessions automatically becomes a U.S. citizen at birth if:
The child was born before noon (Eastern Standard Time) May 24, 1934;
The child's father is a foreign national;
The child's mother was a U.S. citizen at the time of the child's birth; and
The child's U.S. citizen mother resided in the United States prior to the child's birth.
UNMARRIED PARENTS
Child Status Protection Act of 2000 (CSPA) provided for automatic derivation of US Citizenship after birth.
1. Child of a U.S. Citizen Father
Same rules listed above for a child born when his or her parents are married apply to a child born out of wedlock outside of the United States claiming citizenship through a U.S. citizen father if:
A blood relationship between the child and the father is established by clear and convincing evidence;
The child's father was a U.S. citizen at the time of the child's birth either by birth or by naturalization;
The child's father (unless deceased) has agreed in writing to provide financial support for the child until the child reaches 18 years of age; and
One of the following criteria is met before the child reaches 18 years of age:
•The child is legitimated under the law of his or her residence or domicile;
•The father acknowledges in writing and under oath the paternity of the child; or
•The paternity of the child is established by adjudication of a competent court.
 In addition, the residence or physical presence requirements contained in the relevant paragraph of INA 301 continue to apply to children born out of wedlock claiming citizenship through their fathers.
2. Child of a U.S. Citizen Mother
 A child born out of wedlock outside of the United States and its outlying possessions acquires citizenship at birth if:
•The child was born after December 23, 1952;
•The child's mother was a U.S. citizen at the time of the child's birth; and
•The child's U.S. citizen mother was physically present in the United States or outlying possession for one continuous year prior to the child's birth.
The child's parents should contact the nearest U.S. embassy or consulate to apply for a Consular Report of Birth Abroad of a Citizen of the United States of America (CRBA) to document that the child is a U.S. citizen. If the U.S. embassy or consulate determines that the child acquired U.S. citizenship at birth, a consular officer will approve the CRBA application and the Department of State will issue a CRBA, also called a Form FS-240, in the child's name. CRBA is proof of U.S. citizenship and may be used to obtain a U.S. passport and all other purposes.
The child's parents may choose to apply for a U.S. passport for the child at the same time that they apply for a CRBA. Parents may also choose to apply only for a U.S. passport for the child. Like a CRBA, a full validity, unexpired U.S. passport is proof of U.S. citizenship. All applicants applying outside of the United States should apply for passport in person at a U.S. Embassy or Consulate.
Parents of a child born abroad to a U.S. citizen or citizens should apply for a CRBA and/or a U.S. passport for the child as soon as possible. By law, U.S. citizens, including dual nationals, must use a U.S. passport to enter and leave the United States.
If you need help obtaining US Passport for yourself or your child born abroad please contact Washington DC Metropolitan Area Immigration Lawyer Irina Vinogradsky!


text:
Die Leitzach ist ein 33,5 km langer, rechter bzw. südlicher Zufluss der Mangfall in den Bayerischen Voralpen, Bayern (Deutschland). Die Leitzach ist namengebend für das Leitzachtal, von dem ein Teil kurz vor der Einmündung in die Mangfall Goldenes Tal heißt.

Verlauf
Der Fluss entsteht aus dem Zusammenfluss vieler Quellbäche im Mangfallgebirge unterhalb des Ursprungpasses. Hauptquellbach ist der Sillbach, der am Osthang der Maroldschneid bei der Wirthsalm entspringt. Er fließt dann im Ursprungtal nach Norden, ab der Vereinigung mit dem Wackbach unter dem Namen Aubach, und wendet sich bei Bayrischzell nach Nordwesten. Ab hier heißt der Fluss Leitzach. Er fließt durch das Leitzachtal nach Feldkirchen-Westerham, wo er in die Mangfall mündet.

Wasserwirtschaft
Bevor die Leitzach in die Mangfall mündet, wird ihr bereits seit dem Jahr 1913 in der Mühlau Wasser entzogen und in einem 6,7 km langen Stollen in den Seehamer See geleitet, dessen Reservoir für den Betrieb der Leitzachwerke genutzt wird. Das Wasser wird durchschnittlich nach 24 Stunden in die Mangfall geleitet, der es ohnehin zufließen würde.

Die Leitzach ist trotz dieser Verbauungsmaßnahmen und der Wasserkraftnutzung ein weitgehend naturnaher Fluss geblieben. Dennoch stellt die Entnahme des Wassers nach Ansicht der wasserwirtschaftlichen Behörden einen schweren Eingriff in die Natur dar. Der Leitzach werden im Sommer bis zu 7 m³/s, im Winter bis zu 4,6 m³/s entnommen, um den Füllstand des Seehamer Sees mit seinen rund 6.000.000 m³ zu gewährleisten. Erst seit 1978 wurde vertraglich geregelt, dass die Leitzachwerke mindestens 0,5 m³/s im Fluss belassen müssen.

Ortschaften
Zu den Ortschaften an der Leitzach gehören Bayrischzell, Fischbachau, Wörnsmühl, Naring und Westerham.

Literatur
 Josef Brunhuber: Chronik des oberen Leizachtales. Fischbachau / Hundham 1928, Nachdruck 1973, .

Weblinks

 Befahrungsbericht auf www.kajaktour.de
 Wasserstand der Leitzach bei Erb (kurz vor der Mündung)

Einzelnachweise

Fließgewässer im Landkreis Miesbach
Fließgewässer im Landkreis Rosenheim
Geographie (Feldkirchen-Westerham)


text:
Jun 13, 2016 · by TSB	· in Robert Adam |.	·
Oxford University – Bodleian Libraries
"a conversation between the architecture of Ancient Rome, fifth-century Greece and 20th-century Sweden." Jeremy Melvin
A research library for Archaeology, Art History and Classics (Ancient History and Literature) (Adam Architecture, 2001).
Entrance pavilion
A circular pavilion on St John Street forms the entrance to this library, which comprises a small group of buildings dominated by an imposing classical rotunda. The library has incorporated the collections of the former Ashmolean Library, and provides reading areas, a rare books room, and a classics lending library.
Completed at the start of the new millennium, the Sackler Library references the architecture of classical Rome. The pavilion was specifically identified by Robert Adam as based on the Temple of Apollo at Bassae, built between 450 and 400 BC, which was excavated by the architect of the adjacent Ashmolean Museum, C R Cockerell. Shortly after completing this building, Robert Adam founded the Traditional Architecture Group, affiliated to the RIBA, to promote classical and traditional architecture and urbanism in Britain.
1 St John Street, Oxford OX1 2LG
References: Jeremy Melvin, 'Building Study – Sackler Library, University of Oxford – Classical education,' RIBA Journal, 17 Jan 2002 [WWW].
← Saïd Business School
Blavatnik School of Government →


1 The word "queue" is the only word in the English language that is still pronounced the same way when the last four letters are removed.
3 Of all the words in the English language, the word 'set' has the most definitions!
4 What is called a "French kiss" in the English speaking world is known as an "English kiss" in France.
5 "Almost" is the longest word in the English language with all the letters in alphabetical order.
6 "Rhythm" is the longest English word without a vowel.
11 There is a city called Rome on every continent.
12 It's against the law to have a pet dog in Iceland!
13 Your heart beats over 100,000 times a day!
14 Horatio Nelson, one of England's most illustrious admirals was throughout his life, never able to find a cure for his sea-sickness.
17 Your ribs move about 5 million times a year, everytime you breathe!
18 The elephant is the only mammal that can't jump!
19 One quarter of the bones in your body, are in your feet!
20 Like fingerprints, everyone's tongue print is different!
22 Fingernails grow nearly 4 times faster than toenails! Most dust particles in your house are made from dead skin!
23 The present population of 5 billion plus people of the world is predicted to become 15 billion by 2080.
24 Women blink nearly twice as much as men.
25 Adolf Hitler was a vegetarian, and had only ONE testicle.
26 Honey is the only food that does not spoil. Honey found in the tombs of Egyptian pharaohs has been tasted by archaeologists and found edible.
27 Months that begin on a Sunday will always have a "Friday the 13th."
28 Coca-Cola would be green if colouring weren't added to it.
29 On average a hedgehog's heart beats 300 times a minute.
30 More people are killed each year from bees than from snakes.
31 The average lead pencil will draw a line 35 miles long or write approximately 50,000 English words.
32More people are allergic to cow's milk than any other food.
33Camels have three eyelids to protect themselves from blowing sand.
34 The placement of a donkey's eyes in its' heads enables it to see all four feet at all times!
35The six official languages of the United Nations are: English, French, Arabic, Chinese, Russian and Spanish.
36 Earth is the only planet not named after a god.
37 It's against the law to burp, or sneeze in a church in Nebraska, USA.
38 You're born with 300 bones, but by the time you become an adult, you only have 206.
39 Some worms will eat themselves if they can't find any food!
40 Dolphins sleep with one eye open!
42 The worlds oldest piece of chewing gum is 9000 years old!
45 Slugs have 4 noses.
46 Owls are the only birds who can see the colour blue.
47 A man named Charles Osborne had the hiccups for 69 years!
48 A giraffe can clean its ears with its 21-inch tongue!
49 The average person laughs 10 times a day!